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8 Publications visible to you, out of a total of 8
Diel metabolomics analysis of a hot spring chlorophototrophic microbial mat leads to new hypotheses of community member metabolisms
Energy and Material Processing, Microbial Community Dynamics, Adaptive Responses

Abstract (Expand)

Dynamic environmental factors such as light, nutrients, salt, and temperature continuously affect chlorophototrophic microbial mats, requiring adaptive and acclimative responses to stabilize composition … and function. Quantitative metabolomics analysis can provide insights into metabolite dynamics for understanding community response to such changing environmental conditions. In this study, we quantified volatile organic acids, polar metabolites (amino acids, glycolytic and citric acid cycle intermediates, nucleobases, nucleosides, and sugars), wax esters, and polyhydroxyalkanoates, resulting in the identification of 104 metabolites and related molecules in thermal chlorophototrophic microbial mat cores collected over a diel cycle in Mushroom Spring, Yellowstone National Park. A limited number of predominant taxa inhabit this community and their functional potentials have been previously identified through metagenomic and metatranscriptomic analyses and in situ metabolisms, and metabolic interactions among these taxa have been hypothesized. Our metabolomics results confirmed the diel cycling of photorespiration (e.g., glycolate) and fermentation (e.g., acetate, propionate, and lactate) products, the carbon storage polymers polyhydroxyalkanoates, and dissolved gasses (e.g., H2 and CO2) in the waters overlying the mat, which were hypothesized to occur in major mat chlorophototrophic community members. In addition, we have formulated the following new hypotheses: (1) the morning hours are a time of biosynthesis of amino acids, DNA, and RNA; (2) photo-inhibited cells may also produce lactate via fermentation as an alternate metabolism; (3) glycolate and lactate are exchanged among Synechococcus and Roseiflexus spp.; and (4) fluctuations in many metabolite pools (e.g., wax esters) at different times of day result from species found at different depths within the mat responding to temporal differences in their niches.

Authors: Young-Mo Kim, Shane Nowack, Millie T. Olsen, Eric D. Becraft, Jason M. Wood, Vera Thiel, Isaac Klapper, Michael Kühl, James K. Fredrickson, Donald A. Bryant, David M. Ward, Thomas O. Metz

Date Published: 17th Apr 2015

Publication Type: Not specified

Genomic distribution of B-vitamin auxotrophy and uptake transporters in environmental bacteria from the Chloroflexi phylum
Energy and Material Processing, Microbial Community Dynamics, Adaptive Responses

Abstract (Expand)

Bacteria from the Chloroflexi phylum are dominant members of phototrophic microbial mat communities in terrestrial thermal environments. Vitamins of B group are key intermediates (precursors) in the … biosynthesis of indispensable enzyme cofactors driving numerous metabolic processes in all forms of life. A genomics-based reconstruction and comparative analysis of respective biosynthetic and salvage pathways and riboswitch regulons in over 20 representative Chloroflexi genomes revealed a widespread auxotrophy for some of the vitamins. The most prominent predicted phenotypic signature, auxotrophy for vitamins B1 and B7 was experimentally confirmed for the best studied model organism Chloroflexus aurantiacus. These observations along with identified candidate genes for the respective uptake transporters pointed to B vitamin cross-feeding as an important aspect of syntrophic metabolism in microbial communities. Inferred specificities of homologous substrate-binding components of ABC transporters for vitamins B1 (ThiY) and B2 (RibY) were verified by thermofluorescent shift approach. A functional activity of the thiamine-specific transporter ThiXYZ from C. aurantiacus was experimentally verified by genetic complementation in E. coli. Expanding the integrative approach, which was applied here for a comprehensive analysis of B-vitamin metabolism in Chloroflexi would allow reconstruction of metabolic interdependencies in microbial communities.

Authors: I. A. Rodionova, X. Li, A. E. Plymale, K. Motamedchaboki, A. E. Konopka, , , A. L. Osterman, D. A. Rodionov

Date Published: 28th Oct 2014

Publication Type: Not specified

Inference of interactions in cyanobacterial-heterotrophic co-cultures via transcriptome sequencing
Energy and Material Processing, Microbial Community Dynamics, Adaptive Responses

Abstract (Expand)

We used deep sequencing technology to identify transcriptional adaptation of the euryhaline unicellular cyanobacterium Synechococcus sp. PCC 7002 and the marine facultative aerobe Shewanella putrefaciens … W3-18-1 to growth in a co-culture and infer the effect of carbon flux distributions on photoautotroph-heterotroph interactions. The overall transcriptome response of both organisms to co-cultivation was shaped by their respective physiologies and growth constraints. Carbon limitation resulted in the expansion of metabolic capacities, which was manifested through the transcriptional upregulation of transport and catabolic pathways. Although growth coupling occurred via lactate oxidation or secretion of photosynthetically fixed carbon, there was evidence of specific metabolic interactions between the two organisms. These hypothesized interactions were inferred from the excretion of specific amino acids (for example, alanine and methionine) by the cyanobacterium, which correlated with the downregulation of the corresponding biosynthetic machinery in Shewanella W3-18-1. In addition, the broad and consistent decrease of mRNA levels for many Fe-regulated Synechococcus 7002 genes during co-cultivation may indicate increased Fe availability as well as more facile and energy-efficient mechanisms for Fe acquisition by the cyanobacterium. Furthermore, evidence pointed at potentially novel interactions between oxygenic photoautotrophs and heterotrophs related to the oxidative stress response as transcriptional patterns suggested that Synechococcus 7002 rather than Shewanella W3-18-1 provided scavenging functions for reactive oxygen species under co-culture conditions. This study provides an initial insight into the complexity of photoautotrophic-heterotrophic interactions and brings new perspectives of their role in the robustness and stability of the association.

Authors: A. S. Beliaev, , M. Serres, Hans Bernstein, B. E. Linggi, , N. G. Isern, , L. A. Kucek, E. A. Hill, G. E. Pinchuk, D. A. Bryant, , , A. Konopka

Date Published: 29th Apr 2014

Publication Type: Not specified

Genome Sequence of the Thermophilic Cyanobacterium Thermosynechococcus sp. Strain NK55a
Energy and Material Processing, Microbial Community Dynamics, Adaptive Responses

Abstract (Expand)

The genome of the unicellular cyanobacterium Thermosynechococcus sp. strain NK55a, isolated from the Nakabusa hot spring, Nagano Prefecture, Japan, comprises a single, circular, 2.5-Mb chromosome. The … genome is predicted to contain 2,358 protein-encoding genes, including genes for all typical cyanobacterial photosynthetic and metabolic functions. No genes encoding hydrogenases or nitrogenase were identified.

Authors: S. Stolyar, , V. Thiel, L. P. Tomsho, N. Pinel, , , , S. Haruta, S. C. Schuster, D. A. Bryant,

Date Published: 1st Feb 2014

Publication Type: Not specified

Control of proteobacterial central carbon metabolism by the HexR transcriptional regulator: a case study in Shewanella oneidensis
Energy and Material Processing, Microbial Community Dynamics, Adaptive Responses

Abstract (Expand)

Bacteria exploit multiple mechanisms for controlling central carbon metabolism (CCM). Thus, a bioinformatic analysis together with some experimental data implicated the HexR transcriptional factor as … a global CCM regulator in some lineages of Gammaproteobacteria operating as a functional replacement of the Cra regulator characteristic of Enterobacteriales. In this study, we combined a large scale comparative genomic reconstruction of HexR-controlled regulons in 87 species of Proteobacteria with the detailed experimental analysis of the HexR regulatory network in the Shewanella oneidensis model system. Although nearly all of the HexR-controlled genes are associated with CCM, remarkable variations were revealed in the scale (from 1 to 2 target operons in Enterobacteriales up to 20 operons in Aeromonadales) and gene content of HexR regulons between 11 compared lineages. A predicted 17-bp pseudo-palindrome with a consensus tTGTAATwwwATTACa was confirmed as a HexR-binding motif for 15 target operons (comprising 30 genes) by in vitro binding assays. The negative effect of the key CCM intermediate, 2-keto-3-deoxy-6-phosphogluconate, on the DNA-regulator complex formation was verified. A dual mode of HexR action on various target promoters, repression of genes involved in catabolic pathways and activation of gluconeogenic genes, was for the first time predicted by the bioinformatic analysis and experimentally verified by changed gene expression pattern in S. oneidensis DeltahexR mutant. Phenotypic profiling revealed the inability of this mutant to grow on lactate or pyruvate as a single carbon source. A comparative metabolic flux analysis of wild-type and mutant strains of S. oneidensis using [(13)C]lactate labeling and GC-MS analysis confirmed the hypothesized HexR role as a master regulator of gluconeogenic flux from pyruvate via the transcriptional activation of phosphoenolpyruvate synthase (PpsA).

Authors: S. A. Leyn, X. Li, Q. Zheng, P. S. Novichkov, S. Reed, , , C. Yang, A. L. Osterman, D. A. Rodionov

Date Published: 17th Aug 2011

Publication Type: Not specified

Comparative genomic reconstruction of transcriptional networks controlling central metabolism in the Shewanella genus
Energy and Material Processing, Microbial Community Dynamics, Adaptive Responses

Abstract (Expand)

BACKGROUND: Genome-scale prediction of gene regulation and reconstruction of transcriptional regulatory networks in bacteria is one of the critical tasks of modern genomics. The Shewanella genus is … comprised of metabolically versatile gamma-proteobacteria, whose lifestyles and natural environments are substantially different from Escherichia coli and other model bacterial species. The comparative genomics approaches and computational identification of regulatory sites are useful for the in silico reconstruction of transcriptional regulatory networks in bacteria. RESULTS: To explore conservation and variations in the Shewanella transcriptional networks we analyzed the repertoire of transcription factors and performed genomics-based reconstruction and comparative analysis of regulons in 16 Shewanella genomes. The inferred regulatory network includes 82 transcription factors and their DNA binding sites, 8 riboswitches and 6 translational attenuators. Forty five regulons were newly inferred from the genome context analysis, whereas others were propagated from previously characterized regulons in the Enterobacteria and Pseudomonas spp.. Multiple variations in regulatory strategies between the Shewanella spp. and E. coli include regulon contraction and expansion (as in the case of PdhR, HexR, FadR), numerous cases of recruiting non-orthologous regulators to control equivalent pathways (e.g. PsrA for fatty acid degradation) and, conversely, orthologous regulators to control distinct pathways (e.g. TyrR, ArgR, Crp). CONCLUSIONS: We tentatively defined the first reference collection of ~100 transcriptional regulons in 16 Shewanella genomes. The resulting regulatory network contains ~600 regulated genes per genome that are mostly involved in metabolism of carbohydrates, amino acids, fatty acids, vitamins, metals, and stress responses. Several reconstructed regulons including NagR for N-acetylglucosamine catabolism were experimentally validated in S. oneidensis MR-1. Analysis of correlations in gene expression patterns helps to interpret the reconstructed regulatory network. The inferred regulatory interactions will provide an additional regulatory constrains for an integrated model of metabolism and regulation in S. oneidensis MR-1.

Authors: D. A. Rodionov, P. S. Novichkov, E. D. Stavrovskaya, I. A. Rodionova, X. Li, M. D. Kazanov, D. A. Ravcheev, A. V. Gerasimova, A. E. Kazakov, G. Y. Kovaleva, E. A. Permina, O. N. Laikova, R. Overbeek, , , A. P. Arkin, I. Dubchak, A. L. Osterman, M. S. Gelfand

Date Published: 15th Jun 2011

Publication Type: Not specified

Genomic encyclopedia of sugar utilization pathways in the Shewanella genus
Energy and Material Processing, Microbial Community Dynamics, Adaptive Responses

Abstract (Expand)

BACKGROUND: Carbohydrates are a primary source of carbon and energy for many bacteria. Accurate projection of known carbohydrate catabolic pathways across diverse bacteria with complete genomes constitutes … a substantial challenge due to frequent variations in components of these pathways. To address a practically and fundamentally important challenge of reconstruction of carbohydrate utilization machinery in any microorganism directly from its genomic sequence, we combined a subsystems-based comparative genomic approach with experimental validation of selected bioinformatic predictions by a combination of biochemical, genetic and physiological experiments. RESULTS: We applied this integrated approach to systematically map carbohydrate utilization pathways in 19 genomes from the Shewanella genus. The obtained genomic encyclopedia of sugar utilization includes ~170 protein families (mostly metabolic enzymes, transporters and transcriptional regulators) spanning 17 distinct pathways with a mosaic distribution across Shewanella species providing insights into their ecophysiology and adaptive evolution. Phenotypic assays revealed a remarkable consistency between predicted and observed phenotype, an ability to utilize an individual sugar as a sole source of carbon and energy, over the entire matrix of tested strains and sugars.Comparison of the reconstructed catabolic pathways with E. coli identified multiple differences that are manifested at various levels, from the presence or absence of certain sugar catabolic pathways, nonorthologous gene replacements and alternative biochemical routes to a different organization of transcription regulatory networks. CONCLUSIONS: The reconstructed sugar catabolome in Shewanella spp includes 62 novel isofunctional families of enzymes, transporters, and regulators. In addition to improving our knowledge of genomics and functional organization of carbohydrate utilization in Shewanella, this study led to a substantial expansion of our current version of the Genomic Encyclopedia of Carbohydrate Utilization. A systematic and iterative application of this approach to multiple taxonomic groups of bacteria will further enhance it, creating a knowledge base adequate for the efficient analysis of any newly sequenced genome as well as of the emerging metagenomic data.

Authors: D. A. Rodionov, C. Yang, X. Li, I. A. Rodionova, Y. Wang, A. Y. Obraztsova, O. P. Zagnitko, R. Overbeek, M. F. Romine, S. Reed, J. K. Fredrickson, K. H. Nealson, A. L. Osterman

Date Published: 13th Sep 2010

Publication Type: Not specified

Predicting Species-Resolved Macronutrient Acquisition during Succession in a Model Phototrophic Biofilm Using an Integrated 'Omics Approach.
Microbial Community Dynamics

Abstract (Expand)

The principles governing acquisition and interspecies exchange of nutrients in microbial communities and how those exchanges impact community productivity are poorly understood. Here, we examine energy … and macronutrient acquisition in unicyanobacterial consortia for which species-resolved genome information exists for all members, allowing us to use multi-omic approaches to predict species' abilities to acquire resources and examine expression of resource-acquisition genes during succession. Metabolic reconstruction indicated that a majority of heterotrophic community members lacked the genes required to directly acquire the inorganic nutrients provided in culture medium, suggesting high metabolic interdependency. The sole primary producer in consortium UCC-O, cyanobacterium Phormidium sp. OSCR, displayed declining expression of energy harvest, carbon fixation, and nitrate and sulfate reduction proteins but sharply increasing phosphate transporter expression over 28 days. Most heterotrophic members likewise exhibited signs of phosphorus starvation during succession. Though similar in their responses to phosphorus limitation, heterotrophs displayed species-specific expression of nitrogen acquisition genes. These results suggest niche partitioning around nitrogen sources may structure the community when organisms directly compete for limited phosphate. Such niche complementarity around nitrogen sources may increase community diversity and productivity in phosphate-limited phototrophic communities.

Authors: S. R. Lindemann, J. M. Mobberley, J. K. Cole, L. M. Markillie, R. C. Taylor, E. Huang, W. B. Chrisler, H. S. Wiley, M. S. Lipton, W. C. Nelson, J. K. Fredrickson, M. F. Romine

Date Published: No date defined

Publication Type: Not specified

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