An integrative genomics approach to explore diversity of light harvesting antenna complexes from environmental isolates of the purple non-sulfur bacterium Rhodopseudomonas palustris
ABSTRACT: Peripheral light harvesting (LH) antenna complexes have been studied extensively in the purple nonsulfur bacterium Rhodopseudomonas palustris because it produces different types of LH complexes under high light intensities (LH2 complex) and low light intensities (LH3 and LH4 complex). The ability of R. palustris to alter its peripheral LH complexes in response to changes in light intensity is attributed to the multiple operons that encode the a and b peptides that make up these complexes, whose expression is affected by light intensity, light quality, and oxygen tension. However, low resolution structures, amino acid similarities between the complexes, and a lack of transcriptional analysis made it difficult to determine the LH complexes composition and functions under different light intensities. It was also unclear how much diversity of the R. palustris LH complexes exists in nature.Results: To gain insight into the composition of the LH complexes, their function under high light intensities and low light intensities, and their prevalence in the environment we undertook an integrative genomics approach using 15 closely related R. palustris strains isolated from the environment and 5 R. palustris ecotypes whose genomes have been sequenced. We sequenced the genomes for the 15 closely related strains and using RNA-seq carried out transcriptomic analysis on all 20 strains grown under high light intensity and low light intensity. We were able to determine that even closely related R. palustris strains had differences in their pucBA gene content and expression, even under the same growth conditions. We also found that the LH2 complex could compensate for the lack of an LH4 complex under LL intensities but not under extremely LL intensities. Conclusions: This is the first time an integrative genomics approach has been used to study light harvesting in the environment. The variation observed in LH gene composition and expression in environmental isolates of R. palustris likely reflects how these strains have adapted to specific light conditions in the environment. We have also shown that there is redundancy between some of the LH complexes under certain light intensities, which may partially explain why multiple operons encoding LH complexes have evolved and been maintained in R. palustris. Examing the variation observed in LH gene composition and expression in various environmental isolates
Project description:Transcriptome analysis was performed in order to better understand the metabolic activity of non-growing cells of Rhodopseudomonas palustris for improve biofuel production. Gene expression profilings of cells from various time points during non-growing phase were compared using RNA-seq.
Project description:In order to define the AadR regulon we did transcriptome analysis of an aadR in frame-deletion mutant and compared to that of wild type. We found that AadR positively modulated expression of 43 genes, most of which are involved in degradation of aromatic acids under anaerobic conditions. It also activated expression of four genes encoding for unknown proteins and a possible DNA-binding stress protein. Furthermore, AadR repressed the expression of 100 genes, including fixK and many genes controlled by FixK, mainly those involved in a microaerobic lifestyle R. palustris strains were grown in defined medium anaerobically in light (photosynthetically) in sealed tubes with nitrogen gas in the headspace. Succinate and p-coumarate were provided as carbon sources.
Project description:To address the question of how photosynthetic bacterium Rhodopseudomonas palustris metabolize lignin derived compound p-coumarate, transcriptomics and quantitative proteomics were combined to characterize gene expression profiles at both the mRNA level and protein level in Rhodopseudomonas palustris grown with succinate, benzoate, and p-coumarate as the carbon source. Transcriptome profiles among Rhodopseudomonas palustris cells grown with succinate, benzoate, and p-coumarate as the carbon source were compared.
Project description:Photosynthetic microbes can produce the clean-burning fuel hydrogen using one of nature’s most plentiful resources, sunlight 1,2. Anoxygenic photosynthetic bacteria generate hydrogen and ammonia during a process known as biological nitrogen fixation. This reaction is catalyzed by the enzyme nitrogenase and consumes nitrogen gas, ATP and electrons 3. One bacterium, Rhodopseudomonas palustris, has a remarkable ability to obtain electrons from green plant-derived material 4,5 and to efficiently absorb both high and low intensity light energy to form ATP 6. Manipulating R. palustris or a similar organism to produce hydrogen commercially will require us to identify all its genes that contribute to hydrogen production and to understand how this process is regulated in cells. Here we describe mutant strains in which metabolism is redirected such that hydrogen production is uncoupled from nitrogen fixation. Our data indicate that three different single amino acid changes in the transcriptional regulator NifA each yielded strains that produced hydrogen even in the presence of the repressing nitrogen source ammonium and in the absence of specific inducing metabolic signals. We used the mutants to show that, in addition to nitrogenase genes, 18 genes outside of the nitrogenase gene cluster may contribute to hydrogen production. Some of these genes are likely involved in efficient ATP acquisition and in channeling electrons to nitrogenase for reduction of protons to molecular hydrogen. Our results demonstrate that photosynthetic bacteria can be genetically manipulated for sustained production of pure hydrogen in a variety of cultivation conditions in the absence of oxygen, nitrogen or other gases as long as light and an electron donor are supplied. Transcriptome profile of wild type (CGA009) growing photosynthetically in the presence of amonium an acetate was compare with that of 4 different mutants (CGA570, CGA571, CGA572 and CGA574). We did 2 biological replicates per strain.
Project description:Characterization of post-translational modification of nitrogenase in Rhodopseudomonas palustris strains that produce hydrogen gas constitutively. To characterize the effect of yeast extract as nitrogen source on gene expression under nitrogen fixing conditions, expression profiles of wild-type cells grown on either NF plus yeast extract and succinate and PM plus ammonium sulfate and succinate were compared using Affymetrix GeneChip. Using NF plus yeast extract and succinate media, transcriptome profiles of various mutants and wild-type cells were compared using glass spotted microarrays.
Project description:Quorum sensing is a term used to describe cell-to-cell communication that allows cell density-dependent gene expression. Many Gram-negative bacteria use acyl-homoserine lactone (acyl-HSL) synthases to generate fatty acyl-HSL quorum sensing signals, which function with signal receptors to control expression of specific genes. The fatty acyl group is derived from fatty acid biosynthesis and provides signal specificity, but the variety of signals is limited. We have discovered that the photosynthetic bacterium Rhodopseudomonas palustris uses an acyl-HSL synthase to produce p-coumaroyl-HSL by using environmental p-coumaric acid rather than fatty acids from cellular pools. The bacterium has a signal receptor with homology to fatty acyl-HSL receptors that responds to p-coumaroyl-HSL to regulate global gene expression. We also found that p-coumaroyl-HSL is made by other bacteria including Bradyrhizobium BTAi1 and Silicibacter pomeroyi DSS-3. This discovery extends the range of possibilities for acyl-HSL quorum sensing and raises fundamental questions about quorum sensing within the context of environmental signaling. Keywords: Comparison of transcriptome profiles Transcriptome profiles between Rhodopseudomonas palustris cells grown in the in the presence or absence of pC-HSL were compared.
Project description:In order to define the AadR regulon we did transcriptome analysis of an aadR in frame-deletion mutant and compared to that of wild type. We found that AadR positively modulated expression of 43 genes, most of which are involved in degradation of aromatic acids under anaerobic conditions. It also activated expression of four genes encoding for unknown proteins and a possible DNA-binding stress protein. Furthermore, AadR repressed the expression of 100 genes, including fixK and many genes controlled by FixK, mainly those involved in a microaerobic lifestyle Overall design: R. palustris strains were grown in defined medium anaerobically in light (photosynthetically) in sealed tubes with nitrogen gas in the headspace. Succinate and p-coumarate were provided as carbon sources.
Project description:Plants are subjected to perpetual fluctuations of light intensity and spectral composition in their natural growth environment, particularly due to movement of clouds and upper canopy leaves. Sudden exposure to intense light is accompanied by absorption of excess light energy, which results in an overload of photosynthetic electron transport chain and generation of reactive oxygen species in and around thylakoid membranes. To cope with this photooxidative stress and to prevent chronic photoinhibition under dynamically changing light intensities, plants have evolved various short- and long-term photoprotective mechanisms. We used quantitative mass spectrometry to investigate long-term acclimation of Arabidopsis thaliana leaf proteome to fluctuating light (FL) which induces photooxidative stress. After three days of FL exposure the proteomes of young and mature leaves were analyzed separately in the morning and at the end of day to examine possible interaction between FL acclimation and leaf development or time of day.
Project description:In this paper, we present the first comparative transcriptome profiles with ARR treated and control of R. palustris. Moreover, putative two ARR biotransformation mechanisms in R. palustris were first given. All of these provided a valuable genomic resource for further studying molecular mechanism of biotransformation and genetic modification of R. palustris. Overall design: mRNA profiles of the control of R. palustris (RPS) and alpha-rhamnetin-3-rhamnose (ARR) treated of R. palustris (RPB) were generated by deep sequencing using Illumina Hiseq2000.