ALBERT

Description: Although nitric oxide (NO) is an important signaling molecule in bacteria and higher organisms, excessive intracellular NO is highly reactive and dangerous. Therefore, living cells need strict regulation systems for cellular NO homeostasis. Recently, we discovered that Streptomyces coelicolor A3(2) retains the nitrogen oxide cycle (NO 3 – -〉NO 2 – -〉NO-〉NO 3 – ) and nitrite removal system. The nitrogen oxide cycle regulates cellular NO levels, thereby controlling secondary metabolism initiation (red-pigmented antibiotic, RED production) and morphological differentiation. Nitrite induces gene expression in neighboring cells, suggesting another role for this cycle as a producer of transmittable intercellular communication molecules. Here, we demonstrated that ammonium-producing nitrite reductase (NirBD) is involved in regulating NO homeostasis in S. coelicolor A3(2). NirBD was constitutively produced in culture independently of GlnR, a known transcriptional factor. NirBD cleared the accumulated nitrite from the medium. Nir deletion mutants showed increased NO-dependent gene expression at later culture stages, whereas the wild-type M145 showed decreased expression, suggesting that high NO concentration was maintained in the mutant. Moreover, the nir deletion mutant produced more RED than that produced by the wild-type M145. These results suggest that NO 2 – removal by NirBD is important to regulate NO homeostasis and to complete NO signaling in S. coelicolor .

Description: Earlier, vitamin C was demonstrated to sterilize Mycobacterium tuberculosis culture via Fenton's reaction at high concentration. It alters the regulatory pathways associated with stress response and dormancy. Since (p)ppGpp is considered to be the master regulator of stress response and is responsible for bacterial survival under stress, we tested the effect of vitamin C on the formation of (p)ppGpp. In vivo estimation of (p)ppGpp showed a decrease in (p)ppGpp levels in vitamin C-treated M. smegmatis cells in comparison to the untreated cells. Furthermore, in vitro (p)ppGpp synthesis using Rel MSM enzyme was conducted in order to confirm the specificity of the inhibition in the presence of variable concentrations of vitamin C. We observed that vitamin C at high concentration can inhibit the synthesis of (p)ppGpp. We illustrated binding of vitamin C to Rel MSM by isothermal titration calorimetry. Enzyme kinetics was followed where K 0.5 was found to be increased with the concomitant reduction of V max value suggesting mixed inhibition. Both long-term survival and biofilm formation were inhibited by vitamin C. The experiments suggest that vitamin C has the potential to be developed as the inhibitor of (p)ppGpp synthesis and stress response, at least in the concentration range used here.

Description: RNA molecules are attractive therapeutic targets because non-coding RNA molecules have increasingly been found to play key regulatory roles in the cell. Comparing and classifying RNA 3D structures yields unique insights into RNA evolution and function. With the rapid increase in the number of atomic-resolution RNA structures, it is crucial to have effective tools to classify RNA structures and to investigate them for structural similarities at different resolutions. We previously developed the algorithm CLICK to superimpose a pair of protein 3D structures by clique matching and 3D least squares fitting. In this study, we extend and optimize the CLICK algorithm to superimpose pairs of RNA 3D structures and RNA–protein complexes, independent of the associated topologies. Benchmarking Rclick on four different datasets showed that it is either comparable to or better than other structural alignment methods in terms of the extent of structural overlaps. Rclick also recognizes conformational changes between RNA structures and produces complementary alignments to maximize the extent of detectable similarity. Applying Rclick to study Ribonuclease III protein correctly aligned the RNA binding sites of RNAse III with its substrate. Rclick can be further extended to identify ligand-binding pockets in RNA. A web server is developed at http://mspc.bii.a-star.edu.sg/minhn/rclick.html .

Description: By introducing two types of zenith troposphere delay (ZTD) products in precise point positioning (PPP), we developed the ZTD-corrected PPP and the ZTD-constrained PPP, both of them reduced the PPP convergence time. Both enhanced PPP methods are examined by global empirical ZTD models and regional ZTD corrections. For global ZTD models, we verified that ZTD-corrected PPP will deviate the positioning results, while ZTD-constrained PPP could produce unbiased estimations. Therefore, the latter is utilized to study the performance of global ZTD models (ITG, GPT2w, GZTD and UNB3m). After numerous experiments, we found that the performance of ZTD models was positively related to the real ZTD accuracy, and we proposed a universal tropospheric stochastic model 2SQR(9rms) which denotes double the square of nine times ZTD rms, to constrain ZTD in PPP. The proposed model subsequently was validated by real-time static and kinematic ZTD-constrained PPP on the premise that the ZTD rms on every station was known. Compared with traditional PPP, in static PPP, the number of improved stations is increased by 15.5 per cent (ITG), 14.4 per cent (GPT2w), 11.1 per cent (GZTD) and 8.3 per cent (UNB3m). For kinematic PPP, PPP constrained by ITG model still had the best performance, the number of improved stations is increased by 14.4 per cent, after 30 min of initialization time, 13.4 cm east, 13.4 cm north and 11.7 cm up positioning accuracy was obtained, compared with 15.3 cm east, 15.3 cm north and 14.3 cm up accuracy by traditional PPP. In addition, experiments using regional ZTD corrections to enhance real-time PPP showed that both ZTD-corrected PPP and ZTD-constrained PPP can notably reduce the convergence time on the vertical component (within 15 cm).

Description: We study fluctuations in the degree-2 zonal spherical harmonic coefficient of the Earth's gravity potential, C 20 , over the period 2003–2015. This coefficient is related to the Earth's oblateness and studying its temporal variations, C 20 , can be used to monitor large-scale mass movements between high and low latitude regions. We examine C 20 inferred from six different sources, including satellite laser ranging (SLR), GRACE and global geophysical fluids models. We further include estimates that we derive from measured variations in the length-of-day (LOD), from the inversion of global crustal displacements as measured by GPS, as well as from the combination of GRACE and the output of an ocean model as described by Sun et al. We apply a sequence of trend and seasonal moving average filters to the different time-series in order to decompose them into an interannual, a seasonal and an intraseasonal component. We then perform a comparison analysis for each component, and we further estimate the noise level contained in the different series using an extended version of the three-cornered-hat method. For the seasonal component, we generally obtain a very good agreement between the different sources, and except for the LOD-derived series, we find that over 90 per cent of the variance in the seasonal components can be explained by the sum of an annual and semiannual oscillation of constant amplitudes and phases, indicating that the seasonal pattern is stable over the considered time period. High consistency between the different estimates is also observed for the intraseasonal component, except for the solution from GRACE, which is known to be affected by a strong tide-like alias with a period of about 161 d. Estimated interannual components from the different sources are generally in agreement with each other, although estimates from GRACE and LOD present some discrepancies. Slight deviations are further observed for the estimate from the geophysical models, likely to be related to the omission of polar ice and groundwater changes in the model combination we use. On the other hand, these processes do not seem to play an important role at seasonal and shorter timescales, as the sum of modelled atmospheric, oceanic and hydrological effects effectively explains the observed C 20 variations at those scales. We generally obtain very good results for the solution from SLR, and we confirm that this well-established technique accurately tracks changes in C 20 . Good agreement is further observed for the estimate from the GPS inversion, showing that this indirect method is successful in capturing fluctuations in C 20 on scales ranging from intra- to interannual. Obtaining accurate estimates from LOD, however, remains a challenging task and more reliable models of atmospheric wind fields are needed in order to obtain high-quality C 20 , in particular at the seasonal scale. The combination of GRACE data and the output of an ocean model appears to be a promising approach, particularly since corresponding C 20 is not affected by tide-like aliases, and generally gives better results than the solution from GRACE, which still seems to be of rather poor quality.

Description: The culturability of Escherichia coli , Ralstonia eutropha and Bacillus subtilis after incubation in phosphate-buffered saline at either 5°C or 30°C was determined. The culturability of B. subtilis showed little dependence on temperature. The culturability of E. coli rapidly decreased at 30°C but remained almost constant at 5°C. In contrast, the culturability of R. eutropha decreased by three orders of magnitude at 5°C within 24 h but only moderately decreased (one order of magnitude) at 30°C. Remarkably, prolonged incubation of R. eutropha at 30°C resulted in a full recovery of colony forming units in contrast to only a partial recovery at 5°C. Ralstonia eutropha cells at 30°C remained culturable for 3 weeks while culturability at 5°C constantly decreased. The effect of temperature was significantly stronger in a polyhydroxybutyrate-negative mutant. Our data show that accumulated polyhydroxybutyrate has a cold-protective function and can prevent R. eutropha entering the viable but not culturable state.

Description: We consider a new approach to both the forward and inverse problems in post-seismic deformation. We present a method for forward modelling post-seismic deformation in a self-gravitating, heterogeneous and compressible earth with a variety of linear and nonlinear rheologies. We further demonstrate how the adjoint method can be applied to the inverse problem both to invert for rheological structure and to calculate the sensitivity of a given surface measurement to changes in rheology or time-dependence of the source. Both the forward and inverse aspects are illustrated with several numerical examples implemented in a spherically symmetric earth model.

Description: A complex disease generally results not from malfunction of individual molecules but from dysfunction of the relevant system or network, which dynamically changes with time and conditions. Thus, estimating a condition-specific network from a single sample is crucial to elucidating the molecular mechanisms of complex diseases at the system level. However, there is currently no effective way to construct such an individual-specific network by expression profiling of a single sample because of the requirement of multiple samples for computing correlations. We developed here with a statistical method, i.e. a sample-specific network (SSN) method, which allows us to construct individual-specific networks based on molecular expressions of a single sample. Using this method, we can characterize various human diseases at a network level. In particular, such SSNs can lead to the identification of individual-specific disease modules as well as driver genes, even without gene sequencing information. Extensive analysis by using the Cancer Genome Atlas data not only demonstrated the effectiveness of the method, but also found new individual-specific driver genes and network patterns for various types of cancer. Biological experiments on drug resistance further validated one important advantage of our method over the traditional methods, i.e. we can even identify such drug resistance genes that actually have no clear differential expression between samples with and without the resistance, due to the additional network information.

Description: Motivation: Many biological processes, such as cell cycle, circadian clock, menstrual cycles, are governed by oscillatory systems consisting of numerous components that exhibit rhythmic patterns over time. It is not always easy to identify such rhythmic components. For example, it is a challenging problem to identify circadian genes in a given tissue using time-course gene expression data. There is a great potential for misclassifying non-rhythmic as rhythmic genes and vice versa. This has been a problem of considerable interest in recent years. In this article we develop a constrained inference based methodology called Order Restricted Inference for Oscillatory Systems (ORIOS) to detect rhythmic signals. Instead of using mathematical functions (e.g. sinusoidal) to describe shape of rhythmic signals, ORIOS uses mathematical inequalities. Consequently, it is robust and not limited by the biologist's choice of the mathematical model. We studied the performance of ORIOS using simulated as well as real data obtained from mouse liver, pituitary gland and data from NIH3T3, U2OS cell lines. Our results suggest that, for a broad collection of patterns of gene expression, ORIOS has substantially higher power to detect true rhythmic genes in comparison to some popular methods, while also declaring substantially fewer non-rhythmic genes as rhythmic. Availability and Implementation: A user friendly code implemented in R language can be downloaded from http://www.niehs.nih.gov/research/atniehs/labs/bb/staff/peddada/index.cfm . Contact: peddada@niehs.nih.gov

Description: Pseudomonas plecoglossicida is a facultative fish pathogen. Recent studies showed that P. plecoglossicida infection in fish was associated with temperature. The aim of this study was to compare the secretomes of P. plecoglossicida cultured in vitro at representative temperatures for pathogenic (20°C) and less pathogenic (30°C) phenotypes. Thirteen proteins in the culture supernatants of P. plecoglossicida showed significant difference in abundance at 20 vs. 30°C. Four proteins were strongly increased at 20°C, including two hemolysin co-regulated proteins (Hcp) that are part of the bacterial type VI secretion system (T6SS), flagellin and an unknown protein. Immunoblot analysis verified the induced secretion of Hcps at 20°C. Furthermore, the upregulation of Hcps at 20°C was confirmed at transcriptional level by RT-qPCR analysis, which also demonstrated the induction of expression of other T6SS-related genes at 20°C. Taken together, we demonstrate the presence of two functionally active T6SS proteins in fish pathogenic P. plecoglossicida strains, as evidenced by the secretion of the T6SS substrate Hcp, the production of which were found to be controlled by temperature. Our findings also support efforts to develop vaccines targeting secreted virulence factors as prophylactic strategies for diseases in fish caused by P. plecoglossicida .