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  • 1
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Nostoc punctiforme is an example of a filamentous cyanobacterium that is capable of differentiating non-growing cells that constitute gliding filaments termed hormogonia. These gliding filaments serve in short distance dispersal and as infective units in establishing a symbiosis with plants, such as the bryophyte Anthoceros punctatus . Mutants of N . punctiforme exist which show elevated levels of initial infection of A . punctatus as a consequence of repeated cycles of hormogonium differentiation. Such mutations occur within the hrmA and hrmU genes. Further characterization of the hrm locus revealed several genes with an organizational and predicted protein sequence similarity to genes of heterotrophic bacteria that are involved in hexuronic acid metabolism. Genes in the N. punctiforme locus are transcribed in response to the presence of a plant extract containing hormogonium-repressing factors. A predicted transcriptional repressor encoded in the locus, HrmR , was shown herein to be a specific DNA binding protein that regulates the transcription of its own gene and that of hrmE , a nearby gene. The ability of HrmR to bind DNA was abolished upon addition of either galacturonate or lysate from specifically induced N . punctiforme cells, implying that the in vivo HrmR binding activity is modulated via an internal compound, most likely a sugar molecule.
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  • 2
    Electronic Resource
    Electronic Resource
    Oxford BSL : Blackwell Science Ltd
    Molecular microbiology 22 (1996), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The gene encoding glucose-6-phosphate dehydrogenase (G6PD), zwf, in Nostocpunctiforme strain ATCC 29133 is part of a four-gene operon that also encodes fructose bisphosphatase (fbp), transaldolase (tal ) and a gene product termed OpcA, which is cotranscribed with zwf and essential for G6PD activity. The effect of exogenous nitrogen and carbon sources on transcription of these genes was investigated. Growth in the presence of ammonium yielded low levels of transcripts encoding all genes of the operon, while growth under nitrogen-fixing conditions resulted in a large increase of transcripts encoding for fbp and zwf–opcA. When cells are grown in the presence of fructose, levels of transcripts encoding tal and zwf–opcA were increased, relative to levels in ammonium-grown cells. These results indicate that this facultatively heterotrophic cyanobacterium can respond to changes in its environment by altering transcription of genes involved in carbon catabolism. Primer extension identified five 5′ ends corresponding to the major regulated transcripts which we conclude arise from independent transcriptional start points.
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Planta 158 (1983), S. 157-165 
    ISSN: 1432-2048
    Keywords: Anthoceros ; Bryophyte, symbiotic ; Cyanobacteria, symbiotic ; Nostoc ; Symbiosis, reconstitution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The partners of the symbiotic association between Anthoceros punctatus L. and Nostoc spp. have been cultured separately in a pure state. The symbiotic association was reconstituted following dual culture in liquid Anthoceros growth medium with a variety of axenic Nostoc isolates and mutant strains. The heterocyst frequency of competent Nostoc strains increased four- to fivefold when in symbiotic association relative to free-living N2-grown cultures. Dinitrogen fixation by symbiotic Nostoc supported the growth of Anthoceros tissue, although this growth was nitrogen-limited relative to that supported by exogenous ammonium. When the association was reconstituted in the presence of two or three wild-type and mutant Nostoc strains some of these strains were found to compete in infection of Anthoceros tissue and a fraction of the symbiotic Nostoc colonies contained more than one strain. Exogenous ammonium did not affect infection, but repressed development of the symbiotic Nostoc colonies in Anthoceros tissue, and symbiotic Nostoc in N2-grown Anthoceros tissue appeared to regress from the symbiotic state in the presence of exogenous ammonium. The results show that the Anthoceros-Nostoc symbiotic association is amenable to specific experimental manipulations; their implications are discussed with respect to infection of Anthoceros tissue and control of the development of symbiotic Nostoc.
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  • 4
    ISSN: 1432-2048
    Keywords: Anabaena ; Cell uptake ; Cyanobacterium ; Nicotiana ; Polyethylene glycol ; Protoplast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Auxotrophic cells of the filamentous cyanobacterium Anabaena variabilis Kütz. were introduced into protoplasts of Nicotiana tabacum L. in an attempt to engineer a nitrogen-fixing endosymbiosis. Conditions were established to maximize uptake of the cyanobacteria by use of polyethylene glycol. Culture of the novel association was not successful: tobacco protoplasts with Anabaena inside did not appear to divide. Most of the protoplasts expelled the cyanobacteria and simultaneously disintegrated. The reasons for the incompatibility are not known.
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  • 5
    ISSN: 1432-2048
    Keywords: Ammonium assimilation ; Anthoceros ; Bryophyta ; Cyanobacteria ; Glutamine and glutamate formation ; Nostoc ; Symbiosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The pathways of assimilation of ammonium by pure cultures of symbiont-free Anthoceros punctatus L. and the reconstituted Anthoceros-Nostoc symbiotic association were determined from time-course (5–300 s) and inhibitor experiments using 13NH 4 + . The major product of assimilation after all incubation times was glutamine, whether the tissues were cultured with excess ammonium or no combined nitrogen. The 13N in glutamine was predominantly in the amide-nitrogen position. Formation of glutamine and glutamate by Anthoceros-Nostoc was strongly inhibited by either 1mM methionine sulfoximine (MSX) or 1 mM exogenous ammonium. These data are consistent with the assimilation of 13NH 4 + and formation of glutamate by the glutamine synthetase (EC 6.3.1.2)-glutamate synthase (EC 1.4.7.1) pathway in dinitrogen-grown Anthoceros-Nostoc. However, in symbiont-free Anthoceros, grown with 2.5 mM ammonium, formation of glutamine, but not glutamate, was decreased by either MSX or exogenous ammonium. These results indicate that during short incubation times ammonium is assimilated in nitrogenreplete Anthoceros by the activities of both glutamine synthetase and glutamate dehydrogenase (EC 1.4.1.2). In-vitro activities of glutamine synthetase were similar in nitrogen-replete Anthoceros and Anthoceros-Nostoc, indicating that the differences in the routes of glutamate formation were not based upon regulation of synthesis of the initial enzyme of the glutamine synthetase-glutamate synthase pathway. When symbiont-free Anthoceros was cultured for 2 d in the absence of combined nitrogen, total 13NH 4 + assimilation, and glutamine and glutamate formation in the presence of inhibitors, were similar to dinitrogen-grown Anthoceros-Nostoc. The routes of immediate (within 2 min) glutamate formation and ammonium assimilation in Anthoceros were apparently determined by the intracellular levels of ammonium; at low levels the glutamine synthetase-glutamate synthase pathway was predominant, while at high levels independent activities of both glutamine synthetase and glutamate dehydrogenase were expressed.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 78 (1971), S. 25-41 
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A high temperature strain of the blue-green alga, Synechococcus lividus has been cultured and cloned in defined medium. 1. S. lividus (Culture OH-68-s, Clone H-Xf) is an obligate thermophile with a temperature range of growth from 54 to 72°C. The optimal conditions for growth were at 63 to 67°C and a light intensity greater than 700 ft-c, resulting in a reproducible minimum generation time of 11 hours. Growth was depressed in the supra-optimal range from 68 to 72°C. The temperature characteristic or coefficient (μ) of growth was calculated as 13,750 cal mole-1. This value would not distinguish this organism from mesophilic and psychrophilic yeasts and bacteria. 2. Clone H-Xf photosynthesized from as low as 33 to 75°C during short exposure times (20 min) without prior acclimation to the incubation temperatures. Longer exposures to the higher temperatures indicated that the “stable” upper limit for photosynthesis was 73°C when cells were grown above 60°C, but was only 70°C for cultures grown at 55 and 57°C. 3. Abrupt shifts of exponential cultures between optimal (65°C) and near minimal (55°C) growth temperatures showed that lag periods occurred before normal growth commenced at the new temperatures. However, these lag periods on downward temperature shifts followed only after a period of residual growth. Similar shifts from optimal to subminimal (45°C) temperatures indicated that growth continued for a period of time before entering an extended stationary phase prior to senescence. Both of the latter types of experiments may indicate that products synthesized at 65°C are consumed by residual growth after the shifts to lower temperatures, but that these are replaced after a long delay (acclimation) at 55°C and not at all at 45°C. 4. Photoincorporation of 14C−NaHCO3 was highly sensitive to subminimal temperatures after the first hour of exposure. The data suggest that the photosynthetic system could be involved in determining both the upper and lower limits of growth in this organism.
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  • 7
    ISSN: 1432-072X
    Keywords: Ammonium assimilation ; Excretion ; Anabaena azollae ; Azolla caroliniana ; Cyanobacteria ; Glutamine ; Glutamate formation ; Nitrogen fixation ; Symbiosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Anabaena azollae was isolated fromAzolla caroliniana by the “gentle roller” method and differential centrifugation. Incubation of suchAnabaena preparations for 10 min with [13N]N2 resulted in the formation of four radioactive compounds; ammonium, glutamine, glutamate and alanine. Ammonium accounted for 66% of the total radioactivity recovered and 58% of the ammonium was in an extracellular fraction. Since essentially no extracellular13N-labeled organic compounds were found, it appears that ammonium is the compound most probably made available toAzolla during dinitrogen-dependent growth of the association. The kinetics of incorporation of exogenous13NH 4 + into glutamine and glutamate were characteristic of a precursor (glutamine)-product (glutamate) relationship and consistent with assimilation by the glutamine synthetase-glutamate synthase pathway. The results of experiments using the glutamine synthetase inhibitor, methionine sulfoximine, the glutamate synthase inhibitor, diazo-oxonorleucine, and increasing the ammonium concentration to greater than 1 mM, provided evidence for assimilation primarily by the glutamine synthetase-glutamate synthase pathway with little or no contribution from biosynthetic glutamate dehydrogenase. While showing that N2 fixation and NH 4 + assimilation were not tightly coupled metabolic processes in symbioticAnabaena, these results reflect a composite picture and do not indicate the extent to which ammonium assimilatory enzymes might be regulated in filaments associated with specific stages in theAzolla-Anabaena developmental profile.
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  • 8
    ISSN: 1432-072X
    Keywords: Cyanobacteria ; DNA restriction and cloning ; Gene fusions ; Leucine biosynthesis ; Mutant complementation ; Nostoc ; Transcription
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Genomic DNA extracted from the symbiotically-competent, heterocyst-forming cyanobacterium Nostoc sp. strain 7801 was resistant to cleavage by a number of restriction endonucleases. A cosmid library of Nostoc DNA was prepared and maintained in the modification-limited Escherichia coli strain HB101. Analysis of cloned Nostoc DNA fragments indicated infrequent occurrence of restriction endonuclease recognition sites in the Nostoc genome. The Nostoc genomic library was screened for sequences complementing mutations in the E. coli leucine and proline biosynthetic operons. Two cosmids complementing leuB were isolated but none for leuA, leuC, leuD, or proA were detected in 1000 cosmids. A 3.0 kb fragment subcloned from one of the cosmids complemented mutations in leuB when inserted into the HindIII site of pBR322 in either orientation, demonstrating that transcription of leuB originated within the cloned fragment. The cloned fragment also carries a second site capable of initiating transcription of fused antibiotic resistance genes. While transcription of Nostoc DNA sequences did occur in E. coli, unknown barriers must also exist that prevented additional biological complementation of specific E. coli mutations.
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  • 9
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 150 (1988), S. 61-71 
    ISSN: 1432-072X
    Keywords: Anthoceros ; Azolla ; DNA isolation and restriction ; nif gene organization ; Nostoc ; Symbiotic cyanobacteria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The sizes of endonuclease digestion fragments of DNA from cyanobacteria in symbiotic association with Azolla caroliniana or Anthoceros punctatus, or in free-living culture, were compared by Southern hybridization using cloned nitrogenase (nif) genes from Anabaena sp. PCC 7120 as probes. The restriction fragment pattern produced by cyanobacteria isolated from A. caroliniana by culture through symbiotic association with Anthoceros differed from that of the major symbiotic cyanobacterium freshly separated from A. caroliniana. The results indicate that minor cyanobacterial symbionts occur in association with Azolla and that the dominant symbiont was not cultured in the free-living state. Both the absence of hybridization to an xisA gene probe and the mapping of restriction fragments indicated a contiguous nifHDK organization in all cells of the symbiont in association with Azolla. On the other hand, in the cultured isolate from Azolla and in Nostoc sp. 7801, the nifD and nifK genes are nominally separated by an interval of unknown length, compatible with the interruption of the nifHDK operon by a DNA element as observed in Anabaena sp. PCC 7120. In the above cultured strains, restriction fragments consistent with a contiguous nifHDK operon were also present at varying hybridization intensities, especially in Nostoc sp. 7801 grown in association with Anthoceros, presumably due to gene rearrangement in a fraction of the cells.
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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    Molecular genetics and genomics 197 (1984), S. 309-317 
    ISSN: 1617-4623
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Stadtman, Holzer and their colleagues (reviewed in Stadtman and Ginsburg 1974) demonstrated that the enzyme glutamine synthetase (GS) [L-glutamate: ammonia ligase (ADP-forming), EC 6.3.1.2] is covalently modified by adenylylation in a variety of bacterial genera and that the modification is reversible. These studies further indicated that adenylylated GS is the less active form in vitro. To assess the physiological significance of adenylylation of GS we have determined the growth defects of mutant strains (glnE) of S. typhimurium that are unable to modify GS and we have determined the basis for these growth defects. The glnE strains, which lack GS adenylyl transferase activity (ATP: [L-glutamate: ammonia ligase (ADP-forming)] adenylyltransferase, EC 2.7.7.42), show a large growth defect specifically upon shift from a nitrogen-limited growth medium to medium containing excess ammonium (NH4 +). The growth defect appears to be due to very high catalytic activity of GS after shift, which lowers the intracellular glutamate pool to ∼10% that under preshift conditions. Consistent with this view, recovery of a rapid growth rate on NH4 + is accompanied by an increase in the glutamate pool. The glnE strains have normal ATP pools after shift. They synthesize very large amounts of glutamine and excrete glutamine into the medium, but excess glutamine does not seem to inhibit growth. We hypothesize that a major function for adenylylation of bacterial GS is to protect the cellular glutamate pool upon shift to NH4 +-excess conditions and thereby to allow rapid growth.
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