ALBERT

Description: Microbes living in plant tissues—endophytes—are mainly studied in crop plants where they typically colonize the root apoplast. Trees—a large carbon source with a high capacity for photosynthesis—provide a variety of niches for endophytic colonization. We have earlier identified a new type of plant–endophyte interaction in buds of adult Scots pine, where Methylorubrum species live inside the meristematic cells. The endosymbiont Methylorubrum extorquens DSM13060 significantly increases needle and root growth of pine seedlings without producing plant hormones, but by aggregating around host nuclei. Here, we studied gene expression and metabolites of the pine host induced by M. extorquens DSM13060 infection. Malic acid was produced by pine to potentially boost M. extorquens colonization and interaction. Based on gene expression, the endosymbiont activated the auxin- and ethylene (ET)-associated hormonal pathways through induction of CUL1 and HYL1, and suppressed salicylic and abscisic acid signaling of pine. Infection by the endosymbiont had an effect on pine meristem and leaf development through activation of GLP1-7 and ALE2, and suppressed flowering, root hair and lateral root formation by downregulation of AGL8, plantacyanin, GASA7, COW1 and RALFL34. Despite of systemic infection of pine seedlings by the endosymbiont, the pine genes CUL1, ETR2, ERF3, HYL, GLP1-7 and CYP71 were highly expressed in the shoot apical meristem, rarely in needles and not in stem or root tissues. Low expression of MERI5, CLH2, EULS3 and high quantities of ononitol suggest that endosymbiont promotes viability and protects pine seedlings against abiotic stress. Our results indicate that the endosymbiont positively affects host development and stress tolerance through mechanisms previously unknown for endophytic bacteria, manipulation of plant hormone signaling pathways, downregulation of senescence and cell death-associated genes and induction of ononitol biosynthesis.

Description: Boreal trees are capable of taking up organic nitrogen (N) as effectively as inorganic N. Depending on the abundance of soil N forms, plants may adjust physiological and morphological traits to optimize N uptake. However, the link between these traits and N uptake in response to soil N sources is poorly understood. We examined Pinus sylvestris seedlings’ biomass growth and allocation, transpiration, and N uptake in response to additions of organic (the amino acid arginine) or inorganic N (ammonium-nitrate). We also monitored in-situ soil N fluxes in the pots following an addition of N, using a microdialysis system. Supplying organic N resulted in a stable soil N flux, whereas the inorganic N resulted in a sharp increase of nitrate flux followed by a rapid decline, demonstrating a fluctuating N supply and a risk for loss of nitrate from the growth medium. Seedlings supplied with organic N achieved a greater biomass with a higher N content, thus reaching a higher N recovery compared with those supplied inorganic N. In spite of a higher N concentration in organic N seedlings, root-to-shoot ratio and transpiration per unit leaf area were similar to those of inorganic N seedlings. We conclude that enhanced seedlings’ nutrition and growth under the organic N source may be attributed to a stable supply of N, owing to a strong retention rate in the soil medium.

Description: Quantifying inter-specific variations of tree resilience to drought and revealing the underlying mechanisms are of great importance to the understanding of forest functionality particularly in water-limited regions. So far, comprehensive studies incorporating investigations in inter-specific variations of long-term growth patterns of trees and the underlying physiological mechanisms are very limited. Here, in a semi-arid site of northern China, tree radial growth rate, inter-annual tree-ring growth responses to climate variability, as well as physiological characteristics pertinent to xylem hydraulics, carbon assimilation and drought tolerance were analyzed in seven pine species growing in a common environment. Considerable inter-specific variations in radial growth rate, growth response to drought and physiological characteristics were observed among the studied species. Differently, the studied species exhibited similar degrees of resistance to drought-induced branch xylem embolism with water potential corresponding to 50% loss hydraulic conductivity ranged from −2.31 to −2.96 MPa. We found that higher branch hydraulic efficiency is related to greater leaf photosynthetic capacity, smaller hydraulic safety margin and lower woody density (P  0.05). Rather, species with higher hydraulic conductivity and photosynthetic capacity was more sensitive to drought stress and tended to show weaker growth resistance to extreme drought events as quantified by tree-ring analyses, which is at least partially due to a trade-off between hydraulic efficiency and safety across species. This study thus demonstrates the importance of drought resilience rather than instantaneous water and carbon flux capacity in determining tree growth in water-limited environments.

Description: Canopy structure—the size and distribution of tree crowns and the spatial and temporal distribution of leaves within them—exerts dominant control over primary productivity, transpiration and energy exchange. Stand structure—the spatial arrangement of trees in the forest (height, basal area and spacing)—has a strong influence on forest growth, allocation and resource use. Forest response to elevated atmospheric CO2 is likely to be dependent on the canopy and stand structure. Here, we investigated elevated CO2 effects on the forest structure of a Liquidambar styraciflua L. stand in a free-air CO2 enrichment experiment, considering leaves, tree crowns, forest canopy and stand structure. During the 12-year experiment, the trees increased in height by 5 m and basal area increased by 37%. Basal area distribution among trees shifted from a relatively narrow distribution to a much broader one, but there was little evidence of a CO2 effect on height growth or basal area distribution. The differentiation into crown classes over time led to an increase in the number of unproductive intermediate and suppressed trees and to a greater concentration of stand basal area in the largest trees. A whole-tree harvest at the end of the experiment permitted detailed analysis of canopy structure. There was little effect of CO2 enrichment on the relative leaf area distribution within tree crowns and there was little change from 1998 to 2009. Leaf characteristics (leaf mass per unit area and nitrogen content) varied with crown depth; any effects of elevated CO2 were much smaller than the variation within the crown and were consistent throughout the crown. In this young, even-aged, monoculture plantation forest, there was little evidence that elevated CO2 accelerated tree and stand development, and there were remarkably small changes in canopy structure. Questions remain as to whether a more diverse, mixed species forest would respond similarly.

Description: The health of Corymbia calophylla (marri), a keystone tree species in the native forests of southwest Western Australia, has been in decline for the past few decades. Phytophthora root disease and waterlogging have often been cited as contributing to this decline. Traditional methods (i.e., field surveys and sampling) of mapping Phytophthora root infection in the field are time-consuming and expensive; thus, the potential of reflectance spectroscopy to characterize marri response to Phytophthora and waterlogging stress was investigated. Twelve-month old marri plants were infected with either P. cinnamomi or P. multivora in two glasshouse trials and waterlogged for 24 h each fortnight. Spectral measurements with a portable high-resolution spectroradiometer were taken weekly. Plant biophysical measurements were taken at harvest time. Normalized difference spectral index (NDSI) was calculated for every combination of reflectance values between 400 and 2500 nm for all time points, correlated with the treatment effects and displayed as heat maps. Narrowband vegetation indices (VIs), utilizing different wavelengths of the electromagnetic spectrum, were also calculated from the spectral data. The Phytophthora treatments did not cause significant differences with the biophysical measurements in both trials. In the second trial, the waterlogging treatment significantly lowered plant top dry weight (P = 0.016) and diameter (P = 0.044). Reflectance values plotted against wavelength displayed differences between treatments as well as a seasonal trend. The NDSI heat maps indicated that the Phytophthora and waterlogging treatment effects were strongest correlated with bandwidths in the visible and near-infrared portions of the electromagnetic spectrum (538–558 nm and 701–709 nm). Six of the VIs (normalized difference nitrogen index 2, anthocyanin reflectance index 1, photochemical reflectance index, Carter index 1, Vogelman index 3 and water band index) were able to track the biochemical changes in the leaves over the 10 weeks, confirming the seasonal trend. The interaction effect between P. cinnamomi, waterlogging and elapsed time in the first trial was significant for water band index (P = 0.010). This study demonstrates that reflectance spectroscopy holds promise for characterizing marri response but more work needs to be done to identify the optimum wavelengths for identifying Phytophthora and waterlogging stress with marri.

Description: Light and water availability are likely to vary over the lifespan of closed-canopy forest trees with understory trees experiencing greater limitations to growth by light and canopy trees greater limitation due to drought. As drought and shade have opposing effects on isotope discrimination (Δ13C), paired measurement of ring width and Δ13C can potentially be used to differentiate between water and light limitations on tree growth. We tested this approach for Cedrela trees from three tropical forests in Bolivia and Mexico that differ in rainfall and canopy structure. Using lifetime ring width and Δ13C data for trees of up to and over 200 years old, we assessed how controls on tree growth changed from understory to the canopy. Growth and Δ13C are mostly anti-correlated in the understory but this anti-correlation disappeared or weakened when trees reached the canopy, especially at the wettest site. This indicates that understory growth variation is controlled by photosynthetic carbon assimilation (A) due to variation in light levels. Once trees reached the canopy, inter-annual variation in growth and Δ13C at one of the dry sites showed positive correlations, indicating inter-annual variation in growth is driven by variation in water stress affecting stomatal conductance (gs). Paired analysis of ring widths and carbon isotopes provides significant insight to discerning between environmental factors controlling growth over trees’ life; strong light limitations for understory trees in closed-canopy moist forests switched to drought stress for (sub)canopy trees in dry forests. We show that combined isotope and ring width measurements can significantly improve our insights in tree functioning and be used to disentangle limitations due to shade from those due to drought.

Description: Non-structural carbohydrate (NSC) pools fluctuate based on the interplay between photosynthesis, demand from various carbon (C) sinks and tree hydraulic status. Thus, it has been hypothesized that tree species with isohydric stomatal control (i.e., trees that close stomata rapidly in response to drought) rely heavily on NSC pools to sustain metabolism, which can lead to negative physiological consequences such as C depletion. Here, we seek to use a species’ degree of isohydry or anisohydry as a conceptual framework for understanding the interrelations between photosynthetic C supply, hydraulic damage and fluctuations in NSC pools. We conducted a 6-week experimental drought, followed by a 6-week recovery period, in a greenhouse on seven tree species that span the spectrum from isohydric to anisohydric. Throughout the experiment, we measured photosynthesis, hydraulic damage and NSC pools. Non-structural carbohydrate pools were remarkably stable across species and tissues—even highly isohydric species that drastically reduced C assimilation were able to maintain stored C. Despite these static NSC pools, we still inferred an important role for stored C during drought, as most species converted starches into sugars during water stress (and back again post-drought). Finally, we did not observe any linkages between C supply, hydraulic damage and NSC pools, indicating that NSC was maintained independent of variation in photosynthesis and hydraulic function. Our results advance the idea that C depletion is a rare phenomenon due to either active maintenance of NSC pools or sink limitation, and thus question the hypothesis that reductions in C assimilation necessarily lead to C depletion.

Description: AGAMOUS/SEEDSTICK (AG/STK) subfamily genes play crucial roles in the reproductive development of plants. However, most of our current knowledge of AG/STK subfamily genes is restricted to core eudicots and grasses, and the knowledge of ancestral exon–intron structures, expression patterns, protein–protein interaction patterns and functions of AG/STK subfamily genes remains unclear. To determine these, we isolated AG/STK subfamily genes (MawuAG1, MawuAG2 and MawuSTK) from a woody basal angiosperm Magnolia wufengensis (Magnoliaceae). MawuSTK arose from the gene duplication event occurring before the diversification of extant angiosperms, and MawuAG1 and MawuAG2 may result from a gene duplication event occurring before the divergence of Magnoliaceae and Lauraceae. Gene duplication led to apparent diversification in their expression and interaction patterns. It revealed that expression in both stamens and carpels likely represents the ancestral expression profiles of AG lineage genes, and expression of STK-like genes in stamens may have been lost soon after the appearance of the STK lineage. Moreover, AG/STK subfamily proteins may have immediately established interactions with the SEPALLATA (SEP) subfamily proteins following the emergence of the SEP subfamily; however, their interactions with the APETALA1/FRUITFULL subfamily proteins or themselves differ from those found in monocots and basal and core eudicots. MawuAG1 plays highly conserved roles in the determinacy of stamen, carpel and ovule identity, while gene duplication contributed to the functional diversification of MawuAG2 and MawuSTK. In addition, we investigated the evolutionary history of exon–intron structural changes of the AG/STK subfamily, and a novel splice-acceptor mode (GUU-AU) and the convergent evolution of N-terminal extension in the euAG and PLE subclades were revealed for the first time. These results further advance our understanding of ancestral AG/STK subfamily genes in terms of phylogeny, exon–intron structures, expression and interaction patterns, and functions, and provide strong evidence for the significance of gene duplication in the expansion and evolution of the AG/STK subfamily.

Description: Plants have evolved energy dissipation pathways to reduce photooxidative damage under drought when photosynthesis is hampered. Non-volatile and volatile isoprenoids are involved in non-photochemical quenching of excess light energy and scavenging of reactive oxygen species. A better understanding of trees’ ability to cope with and withstand drought stress will contribute to mitigate the negative effects of prolonged drought periods expected under future climate conditions. Therefore we investigated if Douglas-fir (Pseudotsuga menziesii(Mirb.)) provenances from habitats with contrasting water availability reveal intraspecific variation in isoprenoid-mediated energy dissipation pathways. In a controlled drought experiment with 1-year-old seedlings of an interior and a coastal Douglas-fir provenance, we assessed the photosynthetic capacity, pool sizes of non-volatile isoprenoids associated with the photosynthetic apparatus, as well as pool sizes and emission of volatile isoprenoids. We observed variation in the amount and composition of non-volatile and volatile isoprenoids among provenances, which could be linked to variation in photosynthetic capacity under drought. The coastal provenance exhibited an enhanced biosynthesis and emission of volatile isoprenoids, which is likely sustained by generally higher assimilation rates under drought. In contrast, the interior provenance showed an enhanced photoprotection of the photosynthetic apparatus by generally higher amounts of non-volatile isoprenoids and increased amounts of xanthophyll cycle pigments under drought. Our results demonstrate that there is intraspecific variation in isoprenoid-mediated energy dissipation pathways among Douglas-fir provenances, which may be important traits when selecting provenances suitable to grow under future climate conditions.

Description: Abnormal fruitlet abscission is a limiting factor in the production of litchi, an economically important fruit in Southern Asia. Both ethylene and abscisic acid (ABA) induce organ abscission in plants. Although ACS/ACO and NCED genes are known to encode key enzymes required for ethylene and ABA biosynthesis, respectively, the transcriptional regulation of these genes is unclear in the process of plant organ shedding. Here, two polygalacturonase (PG) genes (LcPG1 and LcPG2) and two novel homeodomain-leucine zipper I transcription factors genes (LcHB2 and LcHB3) were identified as key genes associated with the fruitlet abscission in litchi. The expression of LcPG1 and LcPG2 was strongly associated with litchi fruitlet abscission, consistent with enhanced PG activity and reduced homogalacturonan content in fruitlet abscission zones (FAZs). The promoter activities of LcPG1/2 were enhanced by ethephon and ABA. In addition, the production of ethylene and ABA in fruitlets was significantly increased during fruit abscission. Consistently, expression of five genes (LcACO2, LcACO3, LcACS1, LcACS4 and LcACS7) related to ethylene biosynthesis and one gene (LcNCED3) related to ABA biosynthesis in FAZs were activated. Further, electrophoretic mobility shift assays and transient expression experiments demonstrated that both LcHB2 and LcHB3 could directly bind to the promoter of LcACO2/3, LcACS1/4/7 and LcNCED3 genes and activate their expression. Collectively, we propose that LcHB2/3 are involved in the litchi fruitlet abscission through positive regulation of ethylene and ABA biosynthesis.

Description: Maintaining xylem water transport under drought is vital for plants, but xylem failure does occur when drought-induced embolisms form and progressively spread through the xylem. The hydraulic method is widely considered the gold standard to quantify drought-induced xylem embolism. The method determines hydraulic conductivity (Kh) in cut branch samples, dehydrated to specific drought levels, by pushing water through them. The technique is widely considered for its reliable Kh measurements, but there is some uncertainty in the literature over how to define stable Kh and how that relates to the degree of xylem embolism formation. Therefore, the most common setup for this method was extended to measure four parameters: (i) inlet Kh, (ii) outlet Kh, (iii) radial flow from xylem to surrounding living tissue and (iv) the pressure difference across the sample. From a strictly theoretical viewpoint, hydraulic steady state, where inflow equals outflow and radial flow is zero, will result in stable Kh. Application of the setup to Malus domestica Borkh. branches showed that achieving hydraulic steady state takes considerable time (up to 300 min) and that time to reach steady state increased with declining xylem water potentials. During each experimental run, Kh and xylem water potentials dynamically increased, which was supported by X-ray computed microtomography visualizations of embolism refilling under both high- (8 kPa) and low-pressure (2 kPa) heads. Supplying pressurized water can hence cause artificial refilling of vessels, which makes it difficult to achieve a truly stable Kh in partially embolized xylem.

Description: Tree autotrophic respiratory processes, especially stem respiration or stem CO2 efflux (Estem), are important components of the forest carbon budget. Despite the efforts to investigate the controlling processes of Estem in the last years a considerable lack in our knowledge remains on the abiotic and biotic drivers affecting Estem dynamics. It has been strongly advocated that long-term measurements would shed light into those processes. The expensive scientific instruments needed to measure gas exchange has prevented from applying Estem measurements on a larger temporal and spatial scale. Here, we present an automated closed dynamic chamber system based on inexpensive and industrially broadly applied CO2 sensors reducing the costs for the sensing system to a minimum. The CO2 sensor was cross-calibrated with a commonly used gas exchange system in the laboratory and in the field, and we found very good accordance of these sensors. We tested the system under harsh tropical climatic conditions, characterized by heavy tropical rainfall events, extreme humidity, and temperatures, in a moist lowland forest in Malaysia. We recorded Estem of three Dyera costulata trees with our prototype over various days. The variation of Estem was large among the three tree individuals and varied by 7.5-fold. However, clear diurnal changes in Estem were present in all three tree individuals. One tree showed high diurnal variation in Estem and the relationship between Estem and temperature was characterized by a strong hysteresis. The large variations found within one single tree species highlights the importance of continuous measurement to quantify ecosystem carbon fluxes.

Description: Coordination between sapwood-specific hydraulic conductivity (Ks) and stomatal conductance (gs) has been identified in previous studies; however, coordination between leaf hydraulic conductance (Kleaf) and gs, as well as between Kleaf and Ks is not always consistent. This suggests that there is a need to improve our understanding of the coordination among hydraulic and gas exchange traits. In this study, hydraulic traits (e.g., Ks and Kleaf) and gas exchange traits, including gs, transpiration (E) and net CO2 assimilation (Aarea), were measured across 33 co-occurring subtropical woody species. Kleaf was divided into two components: leaf hydraulic conductance inside the xylem (Kleaf-x) and outside the xylem (Kleaf-ox). We found that both Kleaf-x and Kleaf-ox were coordinated with gs and E, but the correlations between Kleaf-ox and gs (or E) were substantially weaker, and that Ks was coordinated with Kleaf-x, but not with Kleaf-ox. In addition, we found that Ks, Kleaf-x and Kleaf-ox together explained 63% of the variation in gs and 42% of the variation in Aarea across species, with Ks contributing the largest proportion of explanatory power, whereas Kleaf-ox contributed the least explanatory power. Our results demonstrate that the coordination between leaf water transport and gas exchange, as well as the hydraulic linkage between leaf and stem, were weakened by Kleaf-ox. This highlights the possibility that water transport efficiencies of stem and leaf xylem, rather than that of leaf tissues outside the xylem, are important determinants of stomatal conductance and photosynthetic capacity across species.

Description: With the prospect of climate change and more frequent El Niño-related dry spells, the drought tolerance of oil palm (Elaeis guineensis Jacq.), one of the most important tropical crop species, is of major concern. We studied the influence of soil water availability and palm height on the plasticity of xylem anatomy of oil palm fronds and their embolism resistance at well-drained and seasonally flooded riparian sites in lowland Sumatra, Indonesia. We found overall mean P12 and P50 values, i.e., the xylem pressures at 12% or 50% loss of hydraulic conductance, of −1.05 and − 1.86 MPa, respectively, indicating a rather vulnerable frond xylem of oil palm. This matches diurnal courses of stomatal conductance, which in combination with the observed low xylem safety evidence a sensitive water loss regulation. While the xylem anatomical traits vessel diameter (Dh), vessel density and potential hydraulic conductivity (Kp) were not different between the sites, palms in the moister riparian plots had on average by 0.4 MPa higher P50 values than plants in the well-drained plots. This could largely be attributed to differences in palm height between systems. As a consequence, palms of equal height had 1.3 MPa less negative P50 values in the moister riparian plots than in the well-drained plots. While palm height was positively related to P50, Dh and Kp decreased with height. The high plasticity in embolism resistance may be an element of the drought response strategy of oil palm, which, as a monocot, has a relatively deterministic hydraulic architecture. We conclude that oil palm fronds develop a vulnerable water transport system, which may expose the palms to increasing drought stress in a warmer and drier climate. However, the risk of hydraulic failure may be reduced by considerable plasticity in the hydraulic system and the environmental control of embolism resistance, and a presumably large stem capacitance.

Description: Canker pathogens cause necrosis of the phloem, but in many host/pathogen systems, they also cause canopy dieback, which implicates xylem, not phloem dysfunction. We hypothesize that this dieback distal to the canker is caused by water stress resulting from the lack of a phloem-to-xylem connection, which in a healthy plant would allow delivery of nonstructural carbohydrates (NSCs) and water inward to aid in xylem embolism refilling. We tested several components of this hypothesis in the host/pathogen system Corylus avellana L./Anisogramma anomala (Peck) E. Müll (Eastern filbert blight). Cankers were non-girdling and usually ≥0.1 m long. As expected, healthy controls had higher specific conductivity (Ks) than diseased stems, but unexpectedly, had similar moisture content (m.c.), showing that the lower Ks did not result from more embolisms in the diseased stems. Moreover, manipulations that removed cambium and phloem to simulate a canker, or that shaded stems to lower NSCs, did not result in lower Ks or m.c. than controls. The outer millimeter of xylem adjacent to a canker had infrequent tyloses and/or fungal hyphae in many but not all samples, and dye studies showed little xylem water transport in that region, but the incidence of these blockages was insufficient to cause the observed 19% decrease in Ks. Healthy stems had higher m.c. than diseased stems above the canker (or analogous) location and were longer for the same leaf weight, suggestive of water stress in the upper portion of diseased stems. These results suggest that dieback distal to cankers in this system results from the bottleneck in water transport in the region adjacent to a canker, but did not find evidence to support the requirement of a phloem-to-xylem connection for continued water transport.

Description: In plants, R2R3 MYB transcription factors (TFs) consist of one large gene family and are involved in the regulation of many developmental processes and various stresses. However, the functions of most of MYB TFs in woody plants remain unknown. Here, PtrMYB94, an R2R3 MYB TF from Populus trichocarpa, is characterized to be involved in the regulation of drought responses and abscisic acid (ABA) signaling. PtrMYB94 encodes a nuclear-localized R2R3 MYB TF. RT-PCR results showed that the PtrMYB94 transcripts were relatively abundant in leaves and stems, and were induced rapidly in response to dehydration stress. Overexpression of PtrMYB94 improved plant drought responses, suggesting that this MYB TF may functionally regulate poplar adaptability to drought stress. Furthermore, the analysis of transcriptional expression and PtrMYB94 promoter: GUS activity showed that PtrMYB94 responded to ABA induction. PtrMYB94-overexpressing plants exhibited the inhibition of seed germination compared with the wild-type (WT) control under ABA exposure condition. The ABA content was evidently increased in the PtrMYB94-overexpressing plants relative to the WT plants. In addition, transcript levels of several ABA- and drought-responsive genes, such as ABA1 and DREB2B, were up-regulated. Taken together, our results suggest that PtrMYB94 is involved in an ABA-dependent drought stress regulation in Populus.

Description: Within the North American boreal forest, a widespread outbreak of the epidermal leaf miner Phyllocnistis populiella Cham. has damaged quaking aspen (Populus tremuloides Michx.) for nearly 20 years. In a series of experiments, we tested the effects of feeding damage by P. populiella on leaf water relations and gas exchange. Relative to insecticide-treated trees, the leaves of naturally mined trees had lower photosynthesis, stomatal conductance to water vapor, transpiration, water-use efficiency, predawn water potential and water content, as well as more enriched foliar δ13C. The magnitude of the difference between naturally mined and insecticide-treated trees did not change significantly throughout the growing season, suggesting that the effect is not caused by accumulation of incidental damage to mined portions of the epidermis over time. The contributions of mining-related stomatal malfunction and cuticular transpiration to these overall effects were investigated by restricting mining damage to stomatous abaxial and astomatous adaxial leaf surfaces. Mining of the abaxial epidermis decreased photosynthesis and enriched leaf δ13C, while increasing leaf water potential and water content relative to unmined leaves, effects consistent with stomatal closure due to disfunction of mined guard cells. Mining of the adaxial epidermis also reduced photosynthesis but had different effects on water relations, reducing midday leaf water potential and water content relative to unmined leaves, and did not affect δ13C. In the laboratory, extent of mining damage to the adaxial surface was positively related to the rate of water loss by leaves treated to prevent water loss through stomata. We conclude that overall, despite water savings due to closure of mined stomata, natural levels of damage by P. populiella negatively impact water relations due to increased cuticular permeability to water vapor across the mined portions of the epidermis. Leaf mining by P. populiella could exacerbate the negative effects of climate warming and water deficit in interior Alaska.

Description: The ability of a plant to form roots from its non-root tissues is ecologically advantageous during rapid adaptation to a changing environment. Although this biological phenomenon has been widely utilized for cuttings in many economically important agronomic and tree species, its genetic and developmental mechanisms have been poorly understood. In this study, we conducted an association analysis of small RNAs, the degradome and the transcriptome of adventitious rooting in poplar softwood cuttings, which revealed that 373 miRNA-target pairs were detected. Of these, 72 significantly differentially expressed targets were screened as likely to modulate adventitious root (AR) development, in conjunction with plant hormone signal transduction. Poplar miR167a and its targets PeARF6s and PeARF8s were subjected to functional verification of their ability to mediate plant growth and hormone signal transduction. Overexpression of miR167a inhibited target transcripts and improved lateral root (LR) development in poplar, while overexpressing PeARF8.1mut increased AR numbers and slightly inhibited LR development. Taken together, these results suggest that miR167a-PeARF8.1 modules play crucial roles in regulating AR and LR development in poplar and improve the adaptation of poplar to more complex environments.

Description: The magnitude and frequency of insect outbreaks are predicted to increase in forests, but how trees cope with severe outbreak defoliation is not yet fully understood. Winter deciduous trees often produce a secondary leaf flush in response to defoliation (i.e., compensatory leaf regrowth or refoliation), which promotes fast replenishment of carbon (C) storage and eventually tree survival. However, secondary leaf flushes may imply a high susceptibility to insect herbivory, especially in the event of an ongoing outbreak. We hypothesized that in winter deciduous species adapted to outbreak-driven defoliations, secondary leaves are both more C acquisitive and more herbivore resistant than primary leaves. During an outbreak by Ormiscodes amphimone F. affecting Nothofagus pumilio (Poepp. & Endl.) Krasser forests, we (i) quantified the defoliation and subsequent refoliation by analyzing the seasonal dynamics of the normalized difference vegetation index (NDVI) and (ii) compared the physiological traits and herbivore resistance of primary and secondary leaves. Comparisons of the NDVI of the primary and second leaf flushes relative to the NDVI of the defoliated forest indicated 31% refoliation, which is close to the leaf regrowth reported by a previous study in juvenile N. pumilio trees subjected to experimental defoliation. Primary leaves had higher leaf mass per area, size, carbon:nitrogen ratio and soluble sugar concentration than secondary leaves, along with lower nitrogen and starch concentrations, and similar total polyphenol and phosphorus concentrations. In both a choice and a non-choice bioassay, the leaf consumption rates by O. amphimone larvae were significantly higher (〉50%) for primary than for secondary leaves, indicating higher herbivore resistance in the latter. Our study shows that secondary leaf flushes in outbreak-adapted tree species can be both C acquisitive and herbivore resistant, and suggests that these two features mediate the positive effects of the compensatory leaf regrowth on the tree C balance and forest resilience.

Description: Tapiscia sinensis Oliv. (Tapisciaceae) has been proven to be a functional androdioecious species with both male and hermaphroditic individuals, and the pollen viability of males is far higher than that of hermaphrodites. To better understand the causes of the low pollen viability in hermaphroditic flowers, different stages of anther development were observed. We found that hermaphroditic flowers exhibit abnormal tapetum development, resulting in low pollen viability. To clarify the underlying molecular mechanism of abnormal tapetum development in hermaphrodites, quantitative real-time PCR analyses were performed. The results revealed that the expression levels of an important transcription factor for tapetum development and function, T. sinensis DYSFUNCTIONAL TAPETUM1 (TsDYT1), and its potential downstream regulatory genes T. sinensis DEFECTIVE in TAPETAL DEVELOPMENT and FUNCTION1 (TsTDF1), T. sinensis ABORTED MICROSPORE (TsAMS) and T. sinensis MALE STERILITY 1 (TsMS1) were all significantly downregulated in hermaphrodites compared with males at some key stages of anther development. The amino acid sequence similarity, expression pattern, gene structure and subcellular localization of these genes were analyzed, and the results indicated functional conservation between T. sinensis and homologues in Arabidopsis thaliana. Next, rapid amplification of cDNA end and thermal asymmetric interlaced PCR were employed to clone the full-length cDNA and promoter sequences of these genes, respectively. In addition, results of yeast two-hybrid analysis showed that TsDYT1 can form heterodimers with TsAMS, and yeast one-hybrid analysis demonstrated that TsDYT1 directly binds to the promoter regions of TsTDF1 and TsMS1. TsTDF1 can directly regulate expression of TsAMS, suggesting that a functionally conserved pathway exists between A. thaliana and T. sinensis to regulate tapetum development. In conclusion, the results suggest that abnormal expression of core transcription factors for tapetum development, including TsDYT1, TsTDF1, TsAMS and TsMS1, plays an important role in the abnormal development of the tapetum in T. sinensis hermaphrodites. Furthermore, a hermaphroditic tapetum with abnormal function causes the low pollen viability of hermaphroditic trees. Our results provide new insight into our understanding of the underlying mechanism of why pollen viability is much higher in males than hermaphrodites of the androdioecious tree T. sinensis.

Description: Given the importance of carbon allocation for plant performance and fitness, it is expected that competition and abiotic stress influence respiratory costs associated with stem wood biomass production and maintenance. In this study, stem respiration (R) was measured together with stem diameter increment in adult trees of eight co-occurring species in a sub-Mediterranean forest stand for 2 years. We estimated growth R (Rg), maintenance R (Rm) and the growth respiration coefficient (GRC) using two gas exchange methods: (i) estimating Rg as the product of growth and GRC (then Rm as R minus Rg) and (ii) estimating Rm from temperature-dependent kinetics of basal Rm at the dormant season (then Rg as R minus Rm). In both cases, stem basal-area growth rates governed intra-annual variation in R, Rg and Rm. Maximum annual Rm occurred slightly before or after maximum Rg. The mean contribution of Rm to R during the growing season ranged from 56% to 88% across species using method 1 and from 23% to 66% using method 2. An analysis accounting for the phylogenetic distance among species indicated that more shade-tolerant, faster growing species exhibited higher Rm and Rg than less shade-tolerant, slower growing ones, suggesting a balance between carbon supply and demand mediated by growth. However, GRC was not related to species growth rate, wood density, or drought and shade tolerance across the surveyed species nor across 27 tree species for which GRC was compiled. The GRC estimates based on wood chemical analysis were lower (0.19) than those based on gas exchange methods (0.35). These results give partial support to the hypothesis that wood production and maintenance costs are related to species ecology and highlight the divergence of respiratory parameters widely used in plant models according to the methodological approach applied to derive them.

Description: Nonstructural carbohydrate (NSC) remobilization remains poorly understood in trees. In particular, it remains unclear (i) which tissues (e.g., living bark or xylem) and compounds (sugars or starch) in woody plants are the main sources of remobilized carbon, (ii) to what extent these NSC pools can be depleted and (iii) whether initial NSC mass or concentration is a better predictor of regrowth potential following disturbance. To address these questions, we collected root segments from a large mature trembling aspen stand; we then allowed them to resprout (sucker) in the dark and remobilize NSC until all sprouts had died. We found that initial starch mass, not concentration, was the best predictor of subsequent sprout mass. In total, more NSC mass (~4×) was remobilized from the living inner bark than the xylem of the roots. After resprouting, root starch was generally depleted to

Description: First-year tree seedlings represent a critical demographic life stage, functioning as a bottleneck to forest regeneration. Knowledge of how mortality is related to whole-seedling carbon and water relations is deficient and is required to understand how forest compositions will be altered in future climatic conditions. We performed a greenhouse drought experiment using first-year seedlings of two common pine species found in the Intermountain West, USA. Gas exchange, biomass gain, allometry and xylem water potentials were compared between well-watered and droughted seedlings from emergence until drought-induced mortality. In both species, morphological adjustments to confer drought tolerance, such as increased leaf mass per unit area, were not observed in seedlings exposed to drought, and droughted seedlings maintained photosynthesis and whole-seedling carbon gain well into the experiment. Yet, there were important differences between species in terms of carbon budgets, physiological responses and mortality patterns. In Pinus ponderosa P. & C. Lawson, physiological acclimation to drought was much greater, evident through stronger stomatal regulation and increased water-use efficiency. Photosynthesis and carbon budgets in P. ponderosa were greater than in Pinus contorta Dougl. ex. Loud., and survival was 100% until critical hydraulic thresholds in leaf water content and seedling water potentials were crossed. In P. contorta, physiological adjustments to drought were less, and mortality occurred much sooner and well before injurious hydraulic thresholds were approached. First-year conifer seedlings appear canalized for a suite of functional traits that prioritize short-term carbon gain over long-term drought tolerance, suggesting that conifer seedling survival is linked with carbon limitations, even during drought, with survival in species having narrower carbon survival margins being more hampered by carbon limitations.

Description: A major environmental pollution problem is the release into the atmosphere of particulate matter, including nanoparticles (NPs), which causes serious hazards to human and ecosystem health, particularly in urban areas. However, knowledge about the uptake, translocation and accumulation of NPs in plant tissues is almost completely lacking. The uptake of silver nanoparticles (Ag-NPs) and their transport and accumulation in the leaves, stems and roots of three different tree species, downy oak (Quercus pubescens Willd.), Scots pine (Pinus sylvestris L.) and black poplar (Populus nigra L.), were assessed. In the experiment, Ag-NPs were supplied separately to the leaves (via spraying, the foliar treatment) and roots (via watering, the root treatment) of the three species. Uptake, transport and accumulation of Ag were investigated through spectroscopy. The concentration of Ag in the stem was higher in the foliar than in the root treatment, and in poplar more than in oak and pine. Foliar treatment with Ag-NPs reduced aboveground biomass and stem length in poplars, but not in oaks or pines. Species-specific signals of oxidative stress were observed; foliar treatment of oak caused the accumulation of H2O2 in leaves, and both foliar and root treatments of poplar led to increased O2− in leaves. Ag-NPs affected leaf and root bacteria and fungi; in the case of leaves, foliar treatment reduced bacterial populations in oak and poplar and fungi populations in pine, and in the case of roots, root treatment reduced bacteria and increased fungi in poplar. Species-specific mechanisms of interaction, transport, allocation and storage of NPs in trees were found. We demonstrated definitively that NPs enter into the tree stem through leaves faster than through roots in all of the investigated tree species.

Description: Plants frequently exhibit tradeoffs between reproduction and growth when resources are limited, and often change these allocation patterns in response to stress. Shorter-lived plants such as annuals tend to allocate relatively more resources toward reproduction when stressed, while longer-lived plants tend to invest more heavily in survival and stress defense. However, severe stress may affect the fitness implications of allocating relatively more resources to reproduction versus stress defense. Increased drought intensity and duration have led to widespread mortality events in coniferous forests. In this review, we ask how potential tradeoffs between reproduction and survival influence the likelihood of drought-induced mortality and species persistence. We propose that trees may exhibit what we call ‘fight or flight’ behaviors under stress. ‘Fight’ behaviors involve greater resource allocation toward survival (e.g., growth, drought-resistant xylem and pest defense). ‘Flight’ consists of higher relative allocation of resources to reproduction, potentially increasing both offspring production and mortality risk for the adult. We hypothesize that flight behaviors increase as drought stress escalates the likelihood of mortality in a given location.

Description: Leaf senescence (LS) affects tree fitness, species distribution and ecosystem structure and functioning. The drivers of LS and the processes underlying it have been studied, but the studies have mainly focused on environmental cues and have mainly been based on statistical analyses using in situ data sets. Experimental investigation and field verification of the processes and drivers are thus urgently needed. We conducted a nutrient-addition experiment after a spring-warming experiment in which an ~40-day range of leaf-out (LO) dates was induced in horse chestnut (Aesculus hippocastanum) and beech (Fagus sylvatica) saplings. We found that both increased nutrient supply and advanced LO date significantly affected the timing of LS, but their effects were opposite, as the former delayed and the latter advanced the senescence. The effects of nutrient supply and LO interacted species specifically. In chestnut, the delay of senescence caused by fertilization increased with the delay of LO and was thus stronger for individuals that flushed late in the spring. On the contrary, in beech the delay of senescence caused by fertilization decreased with the delay of LO and was insignificant for individuals with the latest LO. The experimental findings for beech were confirmed with mature trees at a regional scale. The interactive effect between nutrients and LO on senescence may be associated with variable sensitivity to photoperiod, growth sink limitation and/or direct effect of foliar nutrition on the timing of senescence. Our novel results show that the interactive effects of LO and nutrient supply on the timing of LS should be further addressed experimentally in forthcoming studies. It would also be interesting to consider our results in the further development of phenological models used in assessing the effects of climatic change. The differences found in the present study between horse chestnut and beech suggest that the results found for one species cannot necessarily be generalized to other species, so studies with different temperate tree species are called for.

Description: Modeling stomatal control is critical for predicting forest responses to the changing environment and hence the global water and carbon cycles. A trait-based stomatal control model that optimizes carbon gain while avoiding hydraulic risk has been shown to perform well in response to drought. However, the model’s performance against changes in atmospheric CO2, which is rising rapidly due to human emissions, has yet to be evaluated. The present study tested the gain–risk model’s ability to predict the stomatal response to CO2 concentration with potted water birch (Betula occidentalis Hook.) saplings in a growth chamber. The model’s performance in predicting stomatal response to changes in atmospheric relative humidity and soil moisture was also assessed. The gain–risk model predicted the photosynthetic assimilation, transpiration rate and leaf xylem pressure under different CO2 concentrations, having a mean absolute percentage error (MAPE) of 25%. The model also predicted the responses to relative humidity and soil drought with a MAPE of 21.9% and 41.9%, respectively. Overall, the gain–risk model had an MAPE of 26.8% compared with the 37.5% MAPE obtained by a standard empirical model of stomatal conductance. Importantly, unlike empirical models, the optimization model relies on measurable physiological traits as inputs and performs well in predicting responses to novel environmental conditions without empirical corrections. Incorporating the optimization model in larger scale models has the potential for improving the simulation of water and carbon cycles.

Description: Evergreen tree species that maintain positive carbon balance during the late growing season may subsidize extra carbon in a mixed forest. To test this concept of ‘carbon subsidy’, leaf gas exchange characteristics and related leaf traits were measured for three gymnosperm evergreen species (Chamaecyparis thyoides, Tsuga canadensis and Pinus strobus) native to the oak-hickory deciduous forest in northeast USA from March (early Spring) to October (late Autumn) in a single year. All three species were photosynthetically active in Autumn. During the Summer–Autumn transition, photosynthetic capacity (Amax) of T. canadensis and P. strobus increased (T-test, P 〈 0.001) and was maintained in C. thyoides (T-test, P = 0.49), while dark respiration at 20 °C (Rn) and its thermal sensitivity were generally unchanged for all species (one-way ANOVA, P 〉 0.05). In Autumn, reductions in mitochondrial respiration rate in the daylight (RL) and the ratio of RL to Rn (RL/Rn) were observed in P. strobus (46.3% and 44.0% compared to Summer, respectively). Collectively, these physiological adjustments resulted in higher ratios of photosynthesis to respiration (A/Rnand A/RL) in Autumn for all species. Across season, photosynthetic biochemistry and respiratory variables were not correlated with prevailing growth temperature. Physiological adjustments allowed all three gymnosperm species to maintain positive carbon balance into late Autumn, suggesting that gymnosperm evergreens may benefit from Autumn warming trends relative to deciduous trees that have already lost their leaves.

Description: Forests are sensitive to droughts, which increase the mortality rate of tree species. Various processes have been proposed to underlie drought-induced tree mortality, including hydraulic failure, carbon starvation and increased susceptibility to natural enemies. To give insights into these processes, we assessed the metabolic effects of a mortality-inducing drought on seedlings of Pinus sylvestris L. (Scots Pine), a widespread and important Eurasian species. We found divergence over time in the foliar metabolic composition of droughted vs well-watered seedlings, with the former showing increased abundance of aromatic amino acids and decreases in secondary metabolism associated with defence. We observed no significant differences amongst provenances in these effects: seedlings from drought-prone areas showed the same foliar metabolic changes under drought as seedlings from moist environments, although morphological effects of drought varied by provenance. Overall, our results demonstrate how severe drought prior to death may target particular primary and secondary metabolic pathways, weakening defences against natural enemies and contributing to the risk of drought-induced mortality in P. sylvestris.

Description: Trees use many mechanisms to adapt and respond to stressful conditions. The phenylpropanoid pathway in particular is known to be associated with a diverse suite of plant stress responses. In this study, we explored the relationship between the phenylpropanoid pathway metabolite production, gene expression and adaptive trait variation associated with floral bud reactivation during and following dormancy in Prunus armeniaca L. (apricot). Concentrations of eight phenylpropanoid metabolites were measured during chill accumulation and at developmental stages corresponding to the emergence of sepals and petals in floral buds of varieties that differ phenotypically in bloom date (BD). A significant interaction effect of chill hours and BD phenotype on the concentration of each of the compounds was observed (mixed analysis of variance, P 

Description: Transcription factors (TFs) play crucial roles in the regulation of photosynthesis; elucidating these roles will facilitate our understanding of photosynthesis and thus accelerate its improvement for enhancing crop yield. Promoter analysis of 52 nuclear-encoded Populus tomentosa Carr. genes involved in the Calvin–Benson–Bassham (CBB) cycle revealed 706 motifs and 326 potentially interacting TFs. A backward elimination random forest (BWERF) algorithm reduced the number of TFs to 40, involved in a three-layer gene regulatory network (GRN) including 46 photosynthesis genes (bottom layer), 25 TFs (second layer) and 15 TFs (top layer). Phenotype–genotype association identified 248 single-nucleotide polymorphisms (SNPs) within 72 genes associated with 11 photosynthesis traits. Of the regulatory pairs identified by the BWERF (202 pairs), 77 TF–target combinations harbored SNPs associated with the same trait, supporting similar mechanisms of phenotype modulation. We used expression quantitative trait nucleotide (eQTN) analysis to identify causal SNPs affecting gene expression, identifying 1851 eQTN signals for 50 eGenes (genes whose expressions are regulated by eQTNs). Distribution patterns identified 14 eQTNs from seven TFs associated with eight expression levels of their downstream targets (defined in the GRN), whereas seven TF–target pairs were also identified by phenotype–genotype associations. To further validate the roles of TFs at the metabolic level, we selected 6764 SNPs from 55 genes (identified by GRN–association or GRN–eQTN pairs or both) for metabolic association, identifying variants within 10 TFs affecting metabolic processes underlying the CBB cycle. Our study provides new insights into the photosynthesis pathway in poplar and may facilitate understanding of processes underlying photosynthesis improvement.

Description: Reactivation of axial water flow in ring-porous species is a complex process related to stem water content and developmental stage of both earlywood-vessel and leaf formation. Yet empirical evidence with non-destructive methods on the dynamics of water flow resumption in relation to these mechanisms is lacking. Here we combined in vivo magnetic resonance imaging and wood-anatomical observations to monitor the dynamic changes in stem water content and flow during spring reactivation in 4-year-old pedunculate oaks (Quercus robur L.) saplings. We found that previous year latewood vessels and current year developing earlywood vessels form a functional unit for water flow during growth resumption. During spring reactivation, water flow shifted from latewood towards the new earlywood, paralleling the formation of earlywood vessels and leaves. At leaves' full expansion, volumetric water content of previous rings drastically decreased due to the near-absence of water in fibre tissue. We conclude (i) that in ring-porous oak, latewood vessels play an important hydraulic role for bridging the transition between old and new water-conducting vessels and (ii) that fibre and parenchyma provides a place for water storage.

Description: To study the effects of slightly elevated temperature and ozone (O3) on leaf structural characteristics of silver birch (Betula pendula Roth), saplings of four clonal genotypes of this species were exposed to elevated temperature (ambient air temperature +0.8–1.0 °C) and elevated O3 (1.3–1.4× ambient O3), alone and in combination, in an open-air exposure field over two growing seasons (2007 and 2008). So far, the impacts of moderate elevation of temperature or the combination of elevated temperature and O3 on leaf structure of silver birch have not been intensively studied, thus showing the urgent need for this type of studies. Elevated temperature significantly increased leaf size, reduced non-glandular trichome density, decreased epidermis thickness and increased plastoglobuli size in birch leaves during one or both growing seasons. During the second growing season, O3 elevation reduced leaf size, increased palisade layer thickness and decreased the number of plastoglobuli in spongy cells. Certain leaf structural changes observed under a single treatment of elevated temperature or O3, such as increase in the amount of chloroplasts or vacuole, were no longer detected at the combined treatment. Leaf structural responses to O3 and rising temperature may also depend on timing of the exposure during the plant and leaf development as indicated by the distinct changes in leaf structure along the experiment. Genotype-dependent cellular responses to the treatments were detected particularly in the palisade cells. Overall, this study showed that even a slight but realistic elevation in ambient temperature can notably modify leaf structure of silver birch saplings. Leaf structure, in turn, influences leaf function, thus potentially affecting acclimation capacity under changing climate.

Description: Scots pine (Pinus sylvestris L.) is one of the most important conifers in Northern Europe. In boreal forests, over one-third of net primary production is allocated to roots. Pioneer roots expand the horizontal and vertical root systems and transport nutrients and water from belowground to aboveground. Fibrous roots, often colonized by mycorrhiza, emerge from the pioneer roots and absorb water and nutrients from the soil. In this study, we installed three flatbed scanners to detect the daily growth of both pioneer and fibrous roots of Scots pine during the growing season of 2018, a year with an unexpected summer drought in Southern Finland. The growth rate of both types of roots had a positive relationship with temperature. However, the relations between root elongation rate and soil moisture differed significantly between scanners and between root types indicating spatial heterogeneity in soil moisture. The pioneer roots were more tolerant to severe environmental conditions than the fibrous roots. The pioneer roots initiated elongation earlier and ceased it later than the fibrous roots. Elongation ended when the temperature dropped below the threshold temperature of 4 °C for pioneer roots and 6 °C for fibrous roots. During the summer drought, the fibrous roots halted root surface area growth at the beginning of the drought, but there was no drought effect on the pioneer roots over the same period. To compare the timing of root production and the aboveground organs’ production, we used the CASSIA model, which estimates the aboveground tree carbon dynamics. In this study, root growth started and ceased later than growth of aboveground organs. Pioneer roots accounted for 87% of total root productivity. We suggest that future carbon allocation models should separate the roots by root types (pioneer and fibrous), as their growth patterns are different and they have different reactions to changes in the soil environment.

Description: Tropical dry forests (TDFs) experience a long dry season in which plant species are subject to several months of water deficit. However, TDFs maintain a diverse group of plant life forms, growth forms and leaf phenology, and it is not clear how they vary in their mechanisms for coping with seasonal drought. We studied seasonal changes in leaf water potential (Ψleaf), gas exchange, photochemical activity and functional traits in evergreen and drought-deciduous species from a TDF to determine if leaf phenology mediates plant responses to drought. We found seasonal decreases in Ψleaf, stomatal conductance (gs) and transpiration rate (E), and increases in both intrinsic and instantaneous water-use efficiency. We did not find seasonal differences in photosynthetic rate (Aarea) and carbon isotope composition (δ13C); however, these traits differed between leaf phenology groups, with drought-deciduous plants having higher Aarea and δ13C than evergreen plants. We also found that plants with high leaf nitrogen concentration (Narea) also had low mass-based photosynthetic rate (Amass), photosynthetic-nitrogen-use efficiency and specific leaf area, contrary to the expected relationships given by the leaf economics spectrum. Despite higher Narea, sclerophyllous leaves maintained lower Amass, and this increased structural toughness of leaves may be imposing a stronger limitation for CO2 diffusion and hence photosynthesis. Overall, we found more water-conservative traits in deciduous than in evergreen plants, contrary to what is known about these two leaf phenology groups in other seasonal sites both at tropical and temperate latitudes.

Description: Iron (Fe) is an essential micronutrient for plant survival and proliferation. Plants have evolved complex mechanisms to maintain Fe homeostasis in response to Fe deficiency. In this study, we evaluated the physiological, biochemical and transcriptomic differences between poplars grown under Fe-sufficient and Fe-deficient conditions to elucidate the mechanistic responses of poplars to Fe deficiency. Our results revealed that chlorophyll synthesis and photosynthesis were inhibited under Fe-deficient conditions. The inhibition of these pathways caused chlorosis and reduced shoot growth. Although both photosynthetic systems (PSI and PSII) were inhibited under Fe limitation, PSI was affected more severely and earlier than PSII. Fe deficiency also promoted root growth and increased the accumulation of divalent metal ions in roots. IRT1 and NRAMP1 are both Fe2+ transporters for iron uptake in Arabidopsis. In this study, however, only NRAMP1 was induced to promote Fe2+ uptake in roots at the late stage of Fe deficiency response. It indicated that NRAMP1, rather than the more well-known IRT1, might be a major Fe2+ transporter at the late stage of Fe-deficiency in poplars.

Description: Temperate forests are widely invaded by shade-tolerant shrubs and trees, including those of Eastern North America (ENA). However, it remains unknown whether these invaders are ‘preadapted’ for success in their new ranges due to unique aspects of their evolutionary history or whether selection due to enemy release or other postintroduction processes have driven rapid evolution in the invaded range. We sampled leaf traits of populations of woody understory invaders across light gradients in their native range in Japan and in their invaded ENA range to examine potential phenotypic shifts related to carbon gain and nitrogen use between ranges. We also measured leaf traits in three co-occurring ENA native shrub species. In their invaded range, invaders invested significantly less in leaf chlorophyll content (both per unit leaf mass and area) compared with native range populations of the same species, yet maintained similar rates of photosynthesis in low light. In addition, compared with ENA natives, ENA invaders displayed greater trait variation in response to increasing light availability (forest edges, gaps), giving them a potential advantage over ENA natives in a variety of light conditions. We conclude that, for this group of species, newly evolved phenotypes in the invaded range are more important than preadaptation for their success as shade-tolerant forest invaders.

Description: Electrical resistivity (ER) tomography is a promising technique to minimally invasively study stems of living trees. It allows insights into xylem properties based on the cross-sectional distribution of ER that is governed by the wood’s electrical conductance. In this study, ER measurements were carried out on four forest tree species, Betula pendula, Fagus sylvatica, Picea abies and Pinus sylvestris, to demonstrate interspecific, intraspecific and within-tree variation of ER tomograms. Further, ER patterns were linked to xylem moisture content (MC), electrolyte content and density obtained from wood core analyses. The ER patterns of both coniferous species, P. abies and P. sylvestris, were found to be more homogenous and concentric compared with the complex tomograms of angiosperms. However, the ER range of coniferous trees showed considerable intraspecific variation. Measurements near ground level showed pronounced effects on ER tomograms, highlighting the importance of the chosen measurement height. A strong relation between ER and wood density was found in F. sylvatica while ER patterns of conifers were mainly influenced by MC. Results demonstrate a high species specificity of ER tomograms and of respective influencing xylem traits. They underline the importance of reference measurements for a correct interpretation of ER studies.

Description: To elucidate the role of phytohormones during bud dormancy progression in the Japanese pear (Pyrus pyrifolia Nakai), we investigated changes in phytohormone levels of indole acetic acid (IAA), gibberellic acid (GA), abscisic acid (ABA) and trans-zeatin (tZ). Using ultra-performance liquid chromatography/mass spectrometry/mass spectrometry, we monitored phytohormone levels in the buds of field-grown and potted trees that were artificially heated to modify the timing of dormancy and flowering (spring flush) progression. We also analyzed the expression of GA- and ABA-metabolic genes during dormancy. Indole acetic acid and tZ levels were low during dormancy and increased toward the flowering stage. Gibberellic acid levels were maintained at relatively high concentrations during the dormancy induction stage, then decreased before slightly increasing prior to flowering. The low GA concentration in potted trees compared with field-grown trees indicated that GA functions in regulating tree vigor. Abscisic acid levels increased from the dormancy induction stage, peaked near endodormancy release and steadily decreased before increasing again before the flowering stage. The ABA peak levels did not always coincide with endodormancy release, but peak height correlated with flowering uniformity, suggesting that a decline in ABA concentration was not necessary for resumption of growth but the abundance of ABA might be associated with dormancy depth. From monitoring the expression of genes related to GA and ABA metabolism, we inferred that phytohormone metabolism changed significantly during dormancy, even though the levels of bioactive molecules were consistently low. Phytohormones regulate dormancy progression not only upon the reception of internal signals but also upon sensing ambient conditions.

Description: Metabolic scaling theory allows us to link plant hydraulic structure with metabolic rates in a quantitative framework. In this theoretical framework, we considered the hydraulic structure of current-year shoots in Pinus sylvestris and Picea abies, focusing on two properties unaccounted for by metabolic scaling theories: conifer needles are attached to the entire length of shoots, and the shoot as a terminal element does not display invariant properties. We measured shoot length and diameter as well as conduit diameter and density in two locations of 14 current-year non-leader shoots of pine and spruce saplings, and calculated conductivities of shoots from measured conduit properties. We evaluated scaling exponents for the hydraulic structure of shoots at the end of the water transport pathway from the data and applied the results to simulate water potential of shoots in the crown. Shoot shape was intermediate between cylindrical and paraboloid. Contrary to previous findings, we found that conduit diameter scaled with relative, not absolute, distance from the apex and absolute under-bark shoot diameter independently of species within the first-year shoots. Shoot hydraulic conductivity scaled with shoot diameter and hydraulic diameter. Larger shoots had higher hydraulic conductance. We further demonstrate by novel model calculations that ignoring foliage distribution along the hydraulic pathway overestimates water potential loss in shoots and branches and therefore overestimates related water stress effects. Scaling of hydraulic properties with shoot size enhances apical dominance and may contribute to the decline of whole-tree conductance in large trees.

Description: Vulnerability of xylem to embolism is an important trait related to drought resistance of plants. Methods continue to be developed and debated for measuring embolism. We tested three methods (benchtop dehydration/hydraulic, micro-computed tomography (microCT) and optical) for assessing the vulnerability to embolism of a native California oak species (Quercus douglasii Hook. & Arn.), including an analysis of three different stem ages. All three methods were found to significantly differ in their estimates, with a greater resistance to embolism as follows: microCT 〉 optical 〉 hydraulic. Careful testing was conducted for the hydraulic method to evaluate multiple known potential artifacts, and none was found. One-year-old stems were more resistant than older stems using microCT and optical methods, but not hydraulic methods. Divergence between the microCT and optical methods from the standard hydraulic method was consistent with predictions based on known errors when estimating theoretical losses in hydraulic function in both microCT and optical methods. When the goal of a study is to describe or predict losses in hydraulic conductivity, neither the microCT nor optical methods are reliable for accurately constructing vulnerability curves of stems; nevertheless, these methods may be useful if the goal of a study is to identify embolism events irrespective of hydraulic conductivity or hydraulic function.

Description: Nitrogen (N) is a key nutrient impacting plant growth and physiological processes. However, the supply of N is often not sufficient to meet the requirements of trees in many terrestrial ecosystems. Because of differences in production costs, male and female plants have evolved different stress resistance strategies for N limitation. However, little is known about differential gene expression according to sex in poplars responding to N limitation. To explore sex-related constitutive defenses, Populus cathayana Rehder transcriptomic, proteomic and metabolic analyses were performed on the roots of male and female Populus cathayana. We detected 16,816 proteins and 37,286 transcripts, with 2797 overlapping proteins and mRNAs in the roots. In combination with the identification of 90 metabolites, we found that N deficiency greatly altered gene expression related to N metabolism as well as carbohydrate metabolism, secondary metabolism and stress-related processes in both sexes. Nitrogen-deficient P. cathayana females exhibited greater root biomass and less inhibition of citric acid production and glycolysis as well as higher secondary metabolic activity and abscisic acid contents than N-deficient P. cathayana males. Interestingly, males presented a better osmotic adjustment ability and higher expression of resistance genes, suggesting that P. cathayana males exhibit a better stress tolerance ability and can invest fewer resources in defense compared with females. Therefore, our study provides new molecular evidence that P. cathayana males and females adopt different resistance strategies to cope with N deficiency in their roots.

Description: Working in tandem with root exclusion, stems may provide salt-tolerant woody perennials with some additional capacity to restrict sodium (Na) and chloride (Cl) accumulation in leaves. The Pistacia genus, falling at the nexus of salt tolerance and human intervention, provided an ideal set of organisms for studying the influences of both variable root exclusion and potentially variable discontinuities at the bud union on stem processes. In three experiments covering a wide range of salt concentrations (0 to 150 mM NaCl) and tree ages (1, 2 and 10 years) as well as nine rootstock-scion combinations we show that proportional exclusion of both Na and Cl reached up to ~85% efficacy, but efficacy varied by both rootstock and budding treatment. Effective Na exclusion was augmented by significant retrieval of Na from the xylem sap, as evidenced by declines in the Na concentrations of both sap and wood tissue along the transpiration stream. However, while we observed little to no differences between the concentrations of the two ions in leaves, analogous declines in sap concentrations of Cl were not observed. We conclude that some parallel but separate mechanism must be acting on Cl to provide leaf protection from toxicity specific to this ion and suggest that this mechanism is recirculation of Cl in the phloem. The presented findings underline the importance of holistic assessments of salt tolerance in woody perennials. In particular, greater emphasis might be placed on the dynamics of salt sequestration in the significant storage volumes offered by the stems of woody perennials and on the potential for phloem discontinuity introduced with a bud/graft union.

Description: Aluminum (Al) toxicity is the main constraint of root growth and productivity on arable acidic soil. Although boron (B) is used to ameliorate Al stress, the exact mechanisms underlying the effects of B on Al-induced alteration on root metabolites are poorly understood, especially in the trifoliate orange, which is an important rootstock in China. Therefore, a hydroponics experiment was conducted to explore the mechanisms of B mitigates Al toxicity in roots of citrus by metabolomics. A total of 60 metabolites were identified and analyzed in the present study. The 17 amino acids and 8 sugars were up-regulated in Al-treated roots, mainly histidine, cycloleucine, asparagine, citrulline, raffinose and trehalose, and increased by 38.5-, 8.7-, 6.0-, 6.0-, 7.5- and 6.6-fold, respectively. Meanwhile, significant down-regulation of aspartic acid, isoleucine, glutamic acid and six sugars were indicated under Al stress. Aluminum induced a decrease of nine organic acids, especially l-malic acid, citric acid and threonic acid, by 98.2, 93.6 and 95.1%, respectively. Interestingly, in the presence of Al, B application decreased the contents of asparagine, cycloleucine, citrulline and histidine as well as myo-inositol, raffinose, galactinol and 3,6-anhydro-d-galactose by 52.2, 57.4, 46.7, 63.0, 65.4, 74.3, 62.5 and 55.0%, respectively. However, there was no obvious difference in the organic acid contents in Al-stressed roots treated with B. Conclusively, our results show that B regulates the metabolic patterns of amino acids and carbohydrates and reduces Al toxicity. Nevertheless, B addition did not affect the Al-induced changes in the metabolic modes of organic acids.

Description: Development of lateral buds on the underground rhizome in moso bamboo is essentially the early stage of the development of aboveground branching, which is regulated by Phosphatidyl-Ethanolamine Binding Protein (PEBP) family genes, but it is unknown whether the PEBP family genes are involved in the activation and development of lateral buds underground. By scanning the whole-genome sequence of moso bamboo, we identified 25 PhePEBP family genes and amplified their full-length open reading frames (ORFs). A sequence analysis revealed that they are composed of four exons and three introns, except for PheFT10, which contains six exons and five introns. PheFT10 underwent alternative splicing, resulting in at least four transcripts (PheFT10α, PheFT10β, PheFT10γ and PheFT10δ). Although PhePEBP genes are generally expressed at low levels and show dramatically organ-specific expressions, the transcription levels of most PhePEBP genes, including the transcripts of PheFT10, change with plant age. Together with the observation that the expression of PhePEBP family genes can be regulated by plant hormones and drought, our data suggest that PhePEBP family genes might be involved in the activation of lateral buds and seedling growth. Particularly, PheFT9, PheTFL2 and PheTFL8 may play vital roles during the activation of dormant buds based on the analysis of amino acid substitution and expression profile. These findings provide insights for in-depth exploration of the biological functions of the PhePEBP family genes in regulating the activation of dormant bud and the development of seedling in moso bamboo.

Description: On-going climate change is increasing the risk of drought stress across large areas worldwide. Such drought events decrease ecosystem productivity and have been increasingly linked to tree mortality. Understanding how trees respond to water shortage is key to predicting the future of ecosystem functions. Phloem is at the core of the tree functions, moving resources such as non-structural carbohydrates, nutrients, and defence and information molecules across the whole plant. Phloem function and ability to transport resources is tightly controlled by the balance of carbon and water fluxes within the tree. As such, drought is expected to impact phloem function by decreasing the amount of available water and new photoassimilates. Yet, the effect of drought on the phloem has received surprisingly little attention in the last decades. Here we review existing knowledge on drought impacts on phloem transport from loading and unloading processes at cellular level to possible effects on long-distance transport and consequences to ecosystems via ecophysiological feedbacks. We also point to new research frontiers that need to be explored to improve our understanding of phloem function under drought. In particular, we show how phloem transport is affected differently by increasing drought intensity, from no response to a slowdown, and explore how severe drought might actually disrupt the phloem transport enough to threaten tree survival. Because transport of resources affects other organisms interacting with the tree, we also review the ecological consequences of phloem response to drought and especially predatory, mutualistic and competitive relations. Finally, as phloem is the main path for carbon from sources to sink, we show how drought can affect biogeochemical cycles through changes in phloem transport. Overall, existing knowledge is consistent with the hypotheses that phloem response to drought matters for understanding tree and ecosystem function. However, future research on a large range of species and ecosystems is urgently needed to gain a comprehensive understanding of the question.

Description: Climate change will increase the risk of flooding in several areas of the world where Populus deltoides Marshall (eastern cottonwood) is planted, so it would be desirable for this species to select for flooding tolerance. The aims of this work were to explore the variability in growth, leaf traits and flooding tolerance in an F1 full-sib intraspecific progeny of P. deltoides, to analyze the correlations of leaf and growth traits with flooding tolerance and to assess their suitability for use in breeding programs. Two-month-old parental clones and their progeny of 30 full-sib F1 genotypes were grown in pots and subjected to two treatments: (i) plants watered to field capacity (control) and (ii) plants flooded up to 10 cm above soil level for 35 days. Growth (height, diameter and biomass partition) and leaf traits (leaf size and number, specific leaf area, leaf senescence, abscission, stomatal conductance, carbon isotope discrimination, stomatal index) were measured. Flooding tolerance for each genotype was estimated as the ratio of the biomass of stressed plants to the biomass of control plants. Results showed segregation in terms of flooding tolerance in the F1 progeny. A significant genotype effect was found for leaf size and number, carbon isotopic discrimination and stomatal conductance, but it did not correlate with flooding tolerance. Height, diameter and root-to-shoot ratio had a positive phenotypic correlation with flooding tolerance, and there was a positive genetic correlation of height and diameter with biomass on both treatments. The narrow sense heritability values for the traits analyzed ranged from 0 to 0.56. We conclude that growth traits are more adequate than leaf traits for selection to increase flooding tolerance. A vigorous initial growth would increase flooding tolerance in young poplar plants.

Description: Vascular plants have two types of water-conducting cells, xylem vessel cells (in angiosperms) and tracheid cells (in ferns and gymnosperms). These cells are commonly characterized by secondary cell wall (SCW) formation and programmed cell death (PCD), which increase the efficiency of water conduction. The differentiation of xylem vessel cells is regulated by a set of NAC (NAM, ATAF1/2 and CUC2) transcription factors, called the VASCULAR-RELATED NAC-DOMAIN (VND) family, in Arabidopsis thaliana Linne. The VNDs regulate the transcriptional induction of genes required for SCW formation and PCD. However, information on the transcriptional regulation of tracheid cell differentiation is still limited. Here, we performed functional analysis of loblolly pine (Pinus taeda Linne) VND homologs (PtaVNS, for VND, NST/SND, SMB-related protein). We identified five PtaVNS genes in the loblolly pine genome, and four of these PtaVNS genes were highly expressed in tissues with tracheid cells, such as shoot apices and developing xylem. Transient overexpression of PtaVNS genes induced xylem vessel cell-like patterning of SCW deposition in tobacco (Nicotiana benthamiana Domin) leaves, and up-regulated the promoter activities of loblolly pine genes homologous to SCW-related MYB transcription factor genes and cellulose synthase genes, as well as to cysteine protease genes for PCD. Collectively, our data indicated that PtaVNS proteins possess transcriptional activity to induce the molecular programs required for tracheid formation, i.e., SCW formation and PCD. Moreover, these findings suggest that the VNS–MYB-based transcriptional network regulating water-conducting cell differentiation in angiosperm and moss plants is conserved in gymnosperms.

Description: Most bamboo species including Moso bamboo (Phyllostachys edulis) are tropical or subtropical plants that greatly contribute to human well-being. Low temperature is one of the main environmental factors restricting bamboo growth and geographic distribution. Our knowledge of the molecular changes during bamboo adaption to cold stress remains limited. Here, we provided a general overview of the cold-responsive transcriptional profiles in Moso bamboo by systematically analyzing its transcriptomic response under cold stress. Our results showed that low temperature induced strong morphological and biochemical alternations in Moso bamboo. To examine the global gene expression changes in response to cold, 12 libraries (non-treated, cold-treated 0.5, 1 and 24 h at −2 °C) were sequenced using an Illumina sequencing platform. Only a few differentially expressed genes (DEGs) were identified at early stage, while a large number of DEGs were identified at late stage in this study, suggesting that the majority of cold response genes in bamboo are late-responsive genes. A total of 222 transcription factors from 24 different families were differentially expressed during 24-h cold treatment, and the expressions of several well-known C-repeat/dehydration responsive element-binding factor negative regulators were significantly upregulated in response to cold, indicating the existence of special cold response networks. Our data also revealed that the expression of genes related to cell wall and the biosynthesis of fatty acids were altered in response to cold stress, indicating their potential roles in the acquisition of bamboo cold tolerance. In summary, our studies showed that both plant kingdom-conserved and species-specific cold response pathways exist in Moso bamboo, which lays the foundation for studying the regulatory mechanisms underlying bamboo cold stress response and provides useful gene resources for the construction of cold-tolerant bamboo through genetic engineering in the future.

Description: To increase yield and/or enhance resistance to diseases, grafting is often applied in agriculture and horticulture. Interspecific grafting could possibly be used in forestry as well to improve drought resistance, but our understanding of how the rootstock of a more drought-resistant species can affect the grafted plant is very limited. Reciprocal grafts of two poplar species, Populus cathayana Rehder (less drought-resistant, C) and Populus deltoides Bart. ex Marsh (more drought-resistant, D) were generated. Four grafting combinations (scion/rootstock: C/C, C/D, D/D and D/C) were subjected to well-watered and drought stress treatments. C/D and D/C had a higher diameter growth rate, leaf biomass, intrinsic water-use efficiency (WUEi) and total non-structural carbohydrate (NSC) content than C/C and D/D in well-watered condition. However, drought caused greater differences between P. deltoides-rooted and P. cathayana-rooted grafting combinations, especially between C/D and D/C. The C/D grafting combination showed higher resistance to drought, as indicated by a higher stem growth rate, net photosynthetic rate, WUEi, leaf water potential, proline concentration and NSC concentration and maintenance of integrity of the leaf cellular ultrastructure under drought when compared with D/C. D/C exhibited severely damaged cell membranes, mitochondria and chloroplasts under drought. The scion genotype caused a strong effect on the root proline concentration: the P. cathayana scion increased the root proline concentration more than the P. deltoides scion (C/C vs D/C and C/D vs D/D) under water deficit. Our results demonstrated that mainly the rootstock was responsible for the drought resistance of grafting combinations. Grafting of the P. cathayana scion onto P. deltoides rootstock resulted in superior growth and biomass when compared with the other three combinations both in well-watered and drought stress conditions.

Description: Trees of the genus Prunus produce some of the most widely consumed fruits globally. The combination of climate change-related warming and increased drought stress, scarcity of freshwater resources for irrigation, and increasing demands due to population growth creates a need for increased drought tolerance in these tree species. Recently, we have shown in the field that a native wild pear species performs better under drought than two cultivated pear species. Here, a comparative field study was conducted in Israel to investigate traits associated with drought tolerance in almond (cultivated Prunus dulcis (Mill.) D. A. Webb vs wild Prunus ramonensis Danin) and plum (cultivated Prunus domestica L. vs wild Prunus ursina Kotschy). Measurements of xylem embolism and shoot and root carbon reserves were done along a year, including seasonal drought in the wild and a 35-day drought experiment in the orchards. Synchronous measurements of native xylem embolism and shoot water potential showed that cultivated and wild almond trees lost ~50% of hydraulic conductivity at −2.3 and −3.2 MPa, respectively. Micro-CT images confirmed the higher embolism ratio in cultivated versus wild almond, whereas the two plum species were similar. Dynamics of tissue concentrations of nonstructural carbohydrates were mostly similar across species, with higher levels in cultivated versus wild plum. Our results indicate an advantage for the wild almond over its cultivated relative in terms of xylem resistance to embolism, a major risk factor for trees under drought stress. This result is in line with our previous experiment on pear species. However, the opposite trends observed among the studied plum species mean that these trends cannot be generalized. It is possible that the potential for superior drought tolerance in wild tree species, relative to their cultivated relatives, is limited to wild species from dry and hot habitats.

Description: Due to its ubiquity across northern latitudes, silver birch (Betula pendula Roth) is an attractive model species for studying geographical trait variation and acclimation capacity. Six birch provenances from 60 to 67°N across Finland were grown in a common garden and studied for provenance and genotype variation. We looked for differences in height growth, photosynthetic gas exchange and chlorophyll content index (CCI) and compared the gas exchange of early and late leaves on short and long shoots, respectively. The provenances stratified into southern and northern groups. Northern provenances attained less height growth increment and had higher stomatal conductance (gs) and lower intrinsic water-use efficiency (WUE, Anet/gs) than southern provenances, whereas net photosynthesis (Anet) or CCI did not show clear grouping. Short shoot leaves had lower gs and higher WUE than long shoot leaves in all provenances, but there was no difference in Anet between shoot types. The separation of the provenances into two groups according to their physiological responses might reflect the evolutionary history of B. pendula. Latitudinal differences in gas exchange and water use traits can have plausible consequences for global carbon and water fluxes in a warming climate.

Description: Maximum freezing resistance is a component of winter survival and is associated with the eco-dormant state. Differential thermal analysis (DTA) has shown that changes of the freezing response of the dormant buds depend not only on species and bud type, but also on cooling rates. In order to clarify the freezing adaptation at the cellular level of eco-dormant buds in Japanese white birch, birch buds cooled at a rate of 0.2 °C min−1 and 5 °C day−1 were precisely examined by cryo-scanning electron microscopy (cryo-SEM). Freezing responses of floral dormant buds having female inflorescent primordia and leaf primordia with high-cold hardiness were assessed for extracellular freezing patterns by DTA. Cryo-SEM observation showed freezing of viscous solution filling intercellular spaces within buds and formation of extracellular ice in a random distribution within certain tissues, including green scales, leaf primordia and peduncles. The tissues producing extracellular ice had the common property that distinct intercellular spaces were present among cells having comparatively thick primary walls. In contrast, extracellular ice was not formed within flower primordium and parts of leaf primordium. These tissues had also the common property that no detectable intercellular spaces existed around the cells having thin primary walls. Cryo-SEM observation confirmed that all cells in tissues, regardless of whether extracellular ice was formed within tissues, and also regardless of differences in cooling rates, showed distinct cellular shrinkage by freezing. Recrystallization experiments by cryo-SEM confirmed that all freezable water in cells was eliminated by cooling at 0.2 °C min−1 at least to −30 °C. These results confirmed that all cells in birch buds responded to subzero temperatures through rapid equilibrium dehydration. In contrast to deep supercooling associated with extraorgan freezing of other freezing resistant buds of trees in an eco-dormant state, the mechanism of freezing resistance in eco-dormant birch buds is freezing adaptations by extracellular freezing.

Description: Drought frequency and intensity are projected to increase throughout the southeastern USA, the natural range of loblolly pine (Pinus taeda L.), and are expected to have major ecological and economic implications. We analyzed the carbon and oxygen isotopic compositions in tree ring cellulose of loblolly pine in a factorial drought (~30% throughfall reduction) and fertilization experiment, supplemented with trunk sap flow, allometry and microclimate data. We then simulated leaf temperature and applied a multi-dimensional sensitivity analysis to interpret the changes in the oxygen isotope data. This analysis found that the observed changes in tree ring cellulose could only be accounted for by inferring a change in the isotopic composition of the source water, indicating that the drought treatment increased the uptake of stored moisture from earlier precipitation events. The drought treatment also increased intrinsic water-use efficiency, but had no effect on growth, indicating that photosynthesis remained relatively unaffected despite 19% decrease in canopy conductance. In contrast, fertilization increased growth, but had no effect on the isotopic composition of tree ring cellulose, indicating that the fertilizer gains in biomass were attributable to greater leaf area and not to changes in leaf-level gas exchange. The multi-dimensional sensitivity analysis explored model behavior under different scenarios, highlighting the importance of explicit consideration of leaf temperature in the oxygen isotope discrimination (Δ18Oc) simulation and is expected to expand the inference space of the Δ18Oc models for plant ecophysiological studies.

Description: Plants prevent uncontrolled water loss by synthesizing, depositing and maintaining a hydrophobic layer over their primary aerial organs—the plant cuticle. Quercus coccifera L. can plastically respond to environmental conditions at the cuticular level. When exposed to hot summer conditions with high vapour-pressure deficit (VPD) and intense solar radiation (Mediterranean atmospheric conditions; MED), this plant species accumulates leaf cuticular waxes even over the stomata, thereby decreasing transpirational water loss. However, under mild summer conditions with moderate VPD and regular solar radiation (temperate atmospheric conditions; TEM), this effect is sharply reduced. Despite the ecophysiological importance of the cuticular waxes of Q. coccifera, the wax composition and its contribution to avoiding uncontrolled dehydration remain unknown. Thus, we determined several leaf traits for plants exposed to both MED and TEM conditions. Further, we qualitatively and quantitatively investigated the cuticular lipid composition by gas chromatography. Finally, we measured the minimum leaf conductance (gmin) as an indicator of the efficacy of the cuticular transpiration barrier. The MED leaves were smaller, stiffer and contained a higher load of cuticular lipids than TEM leaves. The amounts of leaf cutin and cuticular waxes of MED plants were 1.4 times and 2.6 times higher than that found for TEM plants, respectively. In detail, MED plants produced higher amounts of all compound classes of cuticular waxes, except for the equivalence of alkanoic acids. Although MED leaves contained higher cutin and cuticular wax loads, the gmin was not different between the two habitats. Our findings suggest that the qualitative accumulation of equivalent cuticular waxes might compensate for the higher wax amount of MED plants, thereby contributing equally to the efficacy of the cuticular transpirational barrier of Q. coccifera. In conclusion, we showed that atmospheric conditions profoundly affect the cuticular lipid composition of Q. coccifera leaves, but do not alter its transpiration barrier properties.

Description: The receptor for activated C kinase 1 (RACK1) protein acts as a central hub for the integration of many physiological processes in eukaryotic organisms. Plant RACK1 is implicated in abiotic stress responses, but the underlying molecular mechanisms of stress adaptation remain largely unknown. Here, the overexpression of the mulberry (Morus alba L.) RACK1 gene in Arabidopsis decreased tolerance to drought and salt stresses and MaRACK1 overexpression changed expression levels of genes in response to stress and stimuli. We developed a simple and efficient transient transformation system in mulberry, and the mulberry seedlings transiently expressing MaRACK1 were hypersensitive to drought and salt stresses. The expression levels of guanine nucleotide-binding protein (G-protein) encoding genes in mulberry and Arabidopsis were not affected by MaRACK1 overexpression. The interactions between RACK1 and G-proteins were confirmed, and the RACK1 proteins from mulberry and Arabidopsis could not interact with their respective G-proteins, which indicated that RACK1 may regulate stress responses independently of G-proteins. Additionally, MaRACK1 may regulate drought and salt stress tolerances by interacting with a fructose 1, 6-bisphosphate aldolase. Our findings provide new insights into the mechanisms underlying RACK1 functions in abiotic stress responses and important information for their further characterization.

Description: We previously identified the FLOWERING LOCUS C (FLC)-like gene, a MADS-box transcription factor gene that belongs to Arabidopsis thaliana FLC clade, in apple (Malus x domestica), and its expression in dormant flower buds is positively correlated with cumulative cold exposure. To elucidate the role of the MdFLC-like in the dormancy process and flower development, we first characterized the phenotypes of MdFLC-like overexpressing lines with the Arabidopsis Columbia-0 background. The overexpression of MdFLC-like significantly delayed the bolting date and reduced the plant size, but it did not significantly affect the number of rosette leaves or flower organ formation. Thus, MdFLC-like may affect vegetative growth and development rather than flowering when expressed in Arabidopsis, which is not like Arabidopsis FLC that affects development of flowering. We compared seasonal expression patterns of MdFLC-like in low-chill ‘Anna’ and high-chill ‘Fuji’ and ‘Tsugaru’ apples collected from trees grown in a cold winter region in temperate zone, and found an earlier up-regulation in ‘Anna’ compared with ‘Fuji’ and ‘Tsugaru’. Expression patterns were also compared in relation to developmental changes in the flower primordia during the chilling accumulation period. Overall, MdFLC-like was progressively up-regulated during flower primordia differentiation and development in autumn to early winter, and reached a maximum expression level at around the same time as the genotype-dependent chilling requirements were fulfilled in high-chill cultivars. Thus, we hypothesize MdFLC-like may be up-regulated in response to cold exposure and flower primordia development during the progress of endodormancy. Our study also suggests MdFLC-like may have a growth inhibiting function during the end of endodormancy and ecodormancy, when the temperature is low and unfavorable for rapid bud outgrowth.

Description: Accumulation of phosphate in leaves as external environmental phosphate concentrations increase has been observed across the plant kingdom. The excess storage of anions, such as phosphate, has various metabolic trade-offs, including a corresponding influx of counter-ions to maintain charge balance and/or the reduction in organic acid content to maintain internal pH. The leaves and roots of four hybrid poplar genotypes were tested for differences in metabolic response to increasing external phosphate and further effects on patterns of anion resorption among hybrid poplar and willow were explored. Organic acid concentrations increased or remained constant across treatments, suggesting that metabolic adjustments were made in response to greater influxes of inorganic cations rather than a response to increasing phosphate. During senescence, the hybrid poplar Tristis had higher sulfate and organic acid resorption, while hybrid willow, AAFC-5, had higher phosphate resorption proficiencies, suggesting differing anion remobilization mechanisms. Furthermore, phosphate accumulation was shown to continue well after bud-set in poplar hybrids, which may contribute to the low phosphorus resorption efficiency. This indicates that closely related species, with similar growth strategies, show preferential resorption toward different nutrients.

Description: Climate change scenarios predict increasing atmospheric CO2 concentrations ([CO2]), temperatures and droughts in tropical regions. Individually, the effects of these climate factors on plants are well established, whereas experiments on the interactive effects of a combination of factors are rare. Moreover, how these environmental factors will affect tree species along a wet to dry gradient (e.g., along tropical forest–savanna transitions) remains to be investigated. We hypothesized that under the simulated environmental conditions, plant growth, physiological performance and survivorship would vary in a manner consistent with the species’ positions of origin along this gradient. In a glasshouse experiment, we raised seedlings of three Eucalyptus species, each occurring naturally in a wet forest, savanna and forest–savanna ecotone, respectively. We evaluated the effect of drought, elevated temperature (4 °C above ambient glasshouse temperature of 22 °C) and elevated temperature in combination with elevated [CO2] (400 ppm [CO2] above ambient of 400 ppm), on seedling growth, survivorship and physiological responses (photosynthesis, stomatal conductance and water-use efficiency). Elevated temperature under ambient [CO2] had little effect on growth, biomass and plant performance of well-watered seedlings, but hastened mortality in drought-affected seedlings, affecting the forest and ecotone more strongly than the savanna species. In contrast, elevated [CO2] in combination with elevated temperatures delayed the appearance of drought stress symptoms and enhanced survivorship in drought-affected seedlings, with the savanna species surviving the longest, followed by the ecotone and forest species. Elevated [CO2] in combination with elevated temperatures also enhanced growth and biomass and photosynthesis in well-watered seedlings of all species, but modified shoot:root biomass partitioning and stomatal conductance differentially across species. Our study highlights the need for a better understand of the interactive effects of elevated [CO2], temperature and drought on plants and the potential to upscale these insights for understanding biome changes.

Description: Understanding which hydraulic traits are under genetic control and/or are phenotypically plastic is essential in understanding how tree species will respond to rapid shifts in climate. We quantified hydraulic traits in Eucalyptus obliqua L'Her. across a precipitation gradient in the field to describe (i) trait variation in relation to long-term climate and (ii) the short-term (seasonal) ability of traits to adjust (i.e., phenotypic plasticity). Seedlings from each field population were raised under controlled conditions to assess (iii) which traits are under strong genetic control. In the field, drier populations had smaller leaves with anatomically thicker xylem vessel walls, a lower leaf hydraulic vulnerability and a lower water potential at turgor loss point, which likely confers higher hydraulic safety. Traits such as the water potential at turgor loss point and ratio of sapwood to leaf area (Huber value) showed significant adjustment from wet to dry conditions in the field, indicating phenotypic plasticity and importantly, the ability to increase hydraulic safety in the short term. In the nursery, seedlings from drier populations had smaller leaves and a lower leaf hydraulic vulnerability, suggesting that key traits associated with hydraulic safety are under strong genetic control. Overall, our study suggests a strong genetic control over traits associated with hydraulic safety, which may compromise the survival of wet-origin populations in drier future climates. However, phenotypic plasticity in physiological and morphological traits may confer sufficient hydraulic safety to facilitate genetic adaptation.

Description: Cork cambium (or phellogen) is a secondary meristem responsible for the formation of phelloderm and phellem/cork, which together compose the periderm. In Quercus suber L., the phellogen is active throughout the entire life of the tree, producing a continuous and renewable outer bark of cork. To identify specific candidate genes associated with cork cambium activity and phellem differentiation, we performed a comparative transcriptomic study of Q. suber secondary growth tissues (xylem and phellogen/phellem) using RNA-seq. The present work provides a high-resolution map of all the transcripts identified in the phellogen/phellem tissues. A total of 6013 differentially expressed genes were identified, with 2875 of the transcripts being specifically enriched during the cork formation process versus secondary xylem formation. Furthermore, cork samples originating from the original phellogen (`virgin’ cork) and from a traumatic phellogen (`amadia’ cork) were also compared. Our results point to a shortlist of potentially relevant candidate genes regulating phellogen activity and phellem differentiation, including novel genes involved in the suberization process, as well as genes associated to ethylene and jasmonate signaling and to meristem function. The future functional characterization of some of the identified candidate genes will help to elucidate the molecular mechanisms underlying cork cambium activity and phellem differentiation.

Description: Vapour pressure deficit (D) is projected to increase in the future as temperatures rise. In response to increased D, stomatal conductance (gs) and photosynthesis (A) are reduced, which may result in significant reductions in terrestrial carbon, water, and energy fluxes. It is thus important for gas exchange models to capture the observed responses of gs and A with increasing D. We tested a series of coupled A-gs models against leaf gas exchange measurements from the Cumberland Plain Woodland (Australia), where D regularly exceeds 2 kPa and can reach 8 kPa in summer. Two commonly used A-gs models (Leuning 1995 and Medlyn et al. 2011) were not able to capture the observed decrease in A and gs with increasing D at the leaf scale. To explain this decrease in A and gs, two alternative hypotheses were tested: hydraulic limitation (i.e., plants reduce gs and/or A due to insufficient water supply) and non-stomatal limitation (i.e., downregulation of photosynthetic capacity). We found that the model that incorporated a non-stomatal limitation captured the observations with high fidelity and required the fewest number of parameters. While the model incorporating hydraulic limitation captured the observed A and gs, it did so via a physical mechanism that is incorrect. We then incorporated a non-stomatal limitation into the stand model, MAESPA, to examine its impact on canopy transpiration and gross primary production. Accounting for a non-stomatal limitation reduced the predicted transpiration by ~19%, improving the correspondence with sap flow measurements, and gross primary production by ~14%. Given the projected global increases in D associated with future warming, these findings suggest that models may need to incorporate non-stomatal limitation to accurately simulate A and gs in the future with high D. Further data on non-stomatal limitation at high D should be a priority, in order to determine the generality of our results and develop a widely applicable model.

Description: Seasonality in tree cambial activity and xylem formation encompass large variation in environmental conditions. Abiotic stressors such as warming or drought also modulate plant behavior at species and individual level. Despite xylem formation susceptibility to carbon (C) and water availability, it is still unknown which are the key physiological variables that regulate xylogenesis, and to what extent plant performance contributes to further explain the number of cells in the different phases of xylem development. Xylogenesis and physiological behavior was monitored in saplings of Pinus pinaster Aiton, a bimodal growth pattern species, distributed in different irrigation regimes. Xylogenesis and plant physiological behavior were compared between treatments and the relationship between climate, physiology and the number of cells in the cambium, enlargement and cell-wall thickening phases was evaluated. Xylogenesis regulation shifted from physiological to climatic control as cell differentiation advanced to mature tracheids. The number of cells in the cambium increased with assimilation rates and decreased with the water potential gradient through the plant. Enlargement was the most susceptible phase to plant relative water content, whereas no physiological variable contributed to explain the number of cells in the wall thickening phase, which declined as temperatures increased. All treatments showed a bimodal growth pattern with a second growth period starting when primary growth was completed and after plants had experienced the highest summer hydraulic losses. Our study demonstrates the importance of including physiological responses and not only climate to fully understand xylogenesis, with special attention to the enlargement phase. This is critical when studying species with a bimodal growth pattern because the second growth peak responds to internal shifts of C allocation and may strongly depend on plant hydraulic responses and not on a fine tuning of cambial activity with soil water availability.

Description: Understanding which structural and functional traits are linked to species’ vulnerability to embolism formation (P50) may provide fundamental knowledge on plant strategies to maintain an efficient water transport. We measured P50, wood density (WD), mean conduit area, conduit density, percentage areas occupied by vessels, parenchyma cells (PATOT) and fibers (FA) on branches of angiosperm and gymnosperm species. Moreover, we compiled a dataset of published hydraulic and anatomical data to be compared with our results. Species more vulnerable to embolism had lower WD. In angiosperms, the variability in WD was better explained by PATOT and FA, which were highly correlated. Angiosperms with a higher P50 (less negative) had a higher amount of PATOT and total amount of nonstructural carbohydrates. Instead, in gymnosperms, P50 vs PATOT was not significant. The correlation between PATOT and P50 might have a biological meaning and also suggests that the causality of the commonly observed relationship of WD vs P50 is indirect and dependent on the parenchyma fraction. Our study suggests that angiosperms have a potential active embolism reversal capacity in which parenchyma has an important role, while in gymnosperms this might not be the case.

Description: Absorptive fine roots are an important driver of soil biogeochemical cycles. Yet, the spatio-temporal dynamics of those roots in the presence of neighboring species remain poorly understood. The aim of this study was to analyze shifts in absorptive fine-root traits in monoculture or mixtures of Fagus sylvatica [L.] and Picea abies [L.] Karst. We hypothesized that root competition would be higher under single-species than mixed-species interactions, leading to changes in (i) root survivorship, diameter and respiration and (ii) spatio-temporal patterns of root growth and death. Using minirhizotron methods, we monitored the timing and location of absorptive fine-root growth and death at an experimental forest in southern Germany from 2011 to 2013. We also measured root respiration in the spring and fall seasons of 2012 and 2013. Our findings show that the absorptive fine roots of F. sylvatica had a 50% higher risk of root mortality and higher respiration rates in the single-species compared to mixed-species zones. These results support our hypothesis that root competition is less intense for F. sylvatica in mixture versus monoculture. We were unable to find confirmation for the same hypothesis for P. abies. To analyze spatio-temporal patterns of absorptive fine-root production and mortality, we used a mixed-effects model considering root depth (space) and seasons (time) simultaneously. This analysis showed that F. sylvatica shifts root production towards shallower soil layers in mixed-species stands, besides significant seasonal fluctuations in root production depths for both species. Ultimately, the impact of neighbor species identity on root traits observed in this study has important implications for where, when and how fast root-facilitated carbon cycling takes place in single-species versus mixed-species forests. In addition, our study highlights the need for inclusion of absorptive fine-root spatio-temporal dynamics when examining belowground plant interactions and biogeochemical cycles.

Description: There is a limited understanding of the impacts of global warming on intra- and interspecific plant competition. Resolving this knowledge gap is important for predicting the potential influence of global warming on forests, particularly on high-altitude trees, which are more sensitive to warming. In the present study, effects of intra- and interspecific competition on plant growth and associated physiological, structural and chemical traits were investigated in Abies faxoniana and Picea purpurea seedlings under control (ambient temperature) and elevated temperature (ET, 2 °C above ambient temperature) conditions for 2 years. We found that A. faxoniana and P. purpurea grown under intra- and interspecific competition showed significant differences in dry matter accumulation (DMA), photosynthetic capacity, nutrient absorption, non-structural carbohydrate (NSC) contents and leaf ultrastructure under ET conditions. ET increased leaf, stem and root DMA of both conifers under both competition patterns. Moreover, under ET and interspecific competition, P. purpurea had overall superior competitive capacity characterized by higher organ (leaf, stem and root) and total DMA, height growth rate, net photosynthetic rate, specific leaf area, water use efficiency (δ13C), leaf and root N and NSC concentrations and greater plasticity for absorption of different soil N forms. Thus, the growth of P. purpurea benefitted from the presence of A. faxoniana under ET. Our results demonstrated that ET significantly affects the asymmetric competition patterns in subalpine conifer species. Potential alteration of plant competitive interactions by global warming can influence the composition, structure and functioning of subalpine coniferous forests.

Description: Limited knowledge about vertical variation in wood CO2 efflux (Rwood) is still a cause of uncertainty in Rwood estimates at individual and ecosystem scales. Although previous studies found higher Rwood in the canopy, they examined several tree species of similar size. In contrast, in the present study, we measured vertical variation in Rwood for 18 trees including 13 species, using a canopy crane for a more precise determination of the vertical variation in Rwood, for various species and sizes of trees in order to examine the factors affecting vertical variation in Rwood and thus, to better understand the effect of taking into account the vertical and inter-individual variation on estimates of Rwood at the individual scale. We did not find any clear pattern of vertical variation; Rwood increased significantly with measurement height for only one tree, while it decreased for two more trees, and was not significantly related with measurement height in 15 other trees. Canopy to breast height Rwood ratio was not related to diameter at breast height or crown ratio, which supposedly are factors affecting vertical variation in Rwood. On average, Rwood estimates at individual scale, considering inter-individual variation but ignoring vertical variation, were only 6% higher than estimates considering both forms of variation. However, estimates considering vertical variation, while ignoring inter-individual variation, were 13% higher than estimates considering both forms of variation. These results suggest that individual measurements at breast height are more important for estimating Rwood at the individual scale, and that any error in Rwood estimation at this scale, due to the absence of any more measurements along tree height, is really quite negligible. This study measured various species and sizes of trees, which may be attributed to no clear vertical variation because factors causing vertical variation can differ among species and sizes.

Description: Few studies have quantified intraspecific variation of hydraulic functional traits in conifers across elevation gradients that include range boundaries. In the Intermountain West, USA, the lower elevational limit of forests (lower treeline) is generally assumed to be caused by water limitations to growth and water relations, yet few studies directly show this. To test this assumption, we measured changes in a suite of traits that characterize drought tolerance such as drought-induced hydraulic vulnerability, hydraulic transport capacity and morphological traits in branch tips and branches of Douglas-fir (Pseudotsuga menziesii var. glauca (Mirb.) Franco) along a 400-m elevation gradient in southeastern Idaho that included lower treeline. As elevation decreased, vulnerability to hydraulic dysfunction and maximum conductivity both decreased in branches; some hydraulic safety–efficiency trade-offs were evident. In branch tips, the water potential at the turgor loss point decreased, while maximum conductance increased with decreasing elevation, highlighting that branch-tip-level responses to less moisture availability accompanied by warmer temperatures might not be coordinated with branch responses. As the range boundary was approached, we did not observe non-linear changes in parameters among sites or increased variance within sites, which current ecological hypotheses on range limits suggest. Our results indicate that there is substantial plasticity in hydraulic functional traits in branch tips and branches of Douglas-fir, although the direction of the trends along the elevation gradient sometimes differed between organs. Such plasticity may mitigate the negative impacts of future drought on Douglas-fir productivity, slowing shifts in its range that are expected to occur with climate change.

Description: Previous studies on the fast growth of bamboo shoots mainly focused on the entire culm. No work about the fast elongation of a single internode, which is the basic unit for the fast growth of bamboo shoots, has been reported so far according to our knowledge. In this study, we have systematically investigated the regulating mechanisms underlying the fast growth of a single bamboo internode of Bambusa multiplex (Lour.) Raeusch. ex Schult. We discovered that the growth of the internode displays a logistic pattern, and the two sections located in the bottom of the internode, one for cell division and, another for cell elongation, each with an ~1-cm length, comprise the effective zones for the internode growth. RNA-Seq analysis identified a number of genes potentially involved in regulating the fast growth of bamboo internode such as those that have positive roles in promoting cell growth or division, which were dramatically down-regulated in the internode at fast growth decreasing stage. Further analysis revealed that sugar plays an important role in promoting the fast growth of bamboo internodes through inhibition of BmSnf1. Mechanical stress is found to be involved in the triggering of the internode growth decrease through activation of the generation of reactive oxygen species by upregulating Calmodulins. These results provide systematic insight into the biological mechanisms underlying the fast growth of bamboo shoots based on the behavior of a single internode.

Description: Several recent reviews highlight the molecular mechanisms that underpin phenological responses to temperature and photoperiod; however, these have mostly overlooked the influence of solar radiation and its spectral composition on these processes. For instance, solar radiation in the blue and ultraviolet (UV) regions of the spectrum, as well as the red/far-red (R:FR) ratio, can influence spring and autumn phenology. Solar radiation reaching the Earth changes diurnally and seasonally; however, rising global temperatures, latitudinal range shifts and light pollution are likely to produce novel combinations of phenological cues for tree species. Here, we review the literature on phenological responses to spectral composition. Our objective was to explore the natural variation in spectral composition using radiative transfer models and to reveal any species-specific or ecotype-specific responses relating to latitudinal origin. These responses are likely to be most pronounced at high latitudes where spectral composition varies most throughout the year. For instance, trees from high latitudes tend to be more sensitive to changes in R:FR than those from low latitudes. The effects of blue light and UV radiation on phenology have not been studied as much as those of R:FR, but the limited results available suggest both could be candidate cues affecting autumn leaf colouration and senescence. Failure of more–southern species and ecotypes to adapt and use spectral cues during northwards range shifts could result in mistimed phenology, potentially resulting in frost damage, reduced fitness and limited range expansion. Future areas for research should look to establish how consistently different functional types of tree respond to spectral cues and identify photoreceptor-mediated mechanisms that allow plants to combine information from multiple light cues to coordinate the timing of phenological events. It should then be feasible to consider the synchronous or sequential action of light cues within a hierarchy of environmental factors regulating phenology.

Description: Insect defoliation contributes to tree mortality under drought conditions. Defoliation-induced alterations to the vascular transport structure may increase tree vulnerability to drought; however, this has been rarely studied. To evaluate the response of tree vascular function following defoliation, 2-year-old balsam poplar were manually defoliated, and both physiological and anatomical measurements were made after allowing for re-foliation. Hydraulic conductivity measurements showed that defoliated trees had both increased vulnerability to embolism and decreased water transport efficiency, likely due to misshapen xylem vessels. Anatomical measurements revealed novel insights into defoliation-induced alterations to the phloem. Phloem sieve tube diameter was reduced in the stems of defoliated trees, suggesting reduced transport capability. In addition, phloem fibers were absent, or reduced in number, in stems, shoot tips and petioles of new leaves, potentially reducing the stability of the vascular tissue. Results from this study suggest that the defoliation leads to trees with increased risk for vascular dysfunction and drought-induced mortality through alterations in the vascular structure, and highlights a route through which carbon limitation can influence hydraulic dysfunction.

Description: It has long been known that the pollen shedding time in pine trees is correlated with temperature, but the molecular basis for this has remained largely unknown. To better understand the mechanisms driving temperature response and to identify the hub regulators of pollen shedding time regulation in Pinus tabuliformis Carr., we identified a set of temperature-sensitive genes by carrying out a comparative transcriptome analysis using six early pollen shedding trees (EPs) and six late pollen shedding trees (LPs) during mid-winter and at three consecutive time points in early spring. We carried out a weighted gene co-expression network analysis and constructed a transcription factor (TF) collaborative network, merging the common but differentially expressed TFs of the EPs and LPs into a joint network. We found five hub genes in the core TF module whose expression was rapidly induced by low temperatures. The transcriptional activity of this TF module was strongly associated with pollen shedding time, and likely to produce the fine balance between cold hardiness and growth activity in early spring. We confirmed the key role of temperature in regulating flowering time and identified a transcription factor module associated with pollen shedding time in P. tabuliformis. This suggests that repression of growth activity by repressors is the main mechanism balancing growth and cold hardiness in pine trees in early spring. Our results provide new insights into the molecular mechanisms regulating seasonal flowering time in pines.