Dataset Information

Proteomic and Isotopic Response of Desulfovibrio vulgaris to DsrC Perturbation.

ABSTRACT: Dissimilatory sulfate reduction is a microbial energy metabolism that can produce sulfur isotopic fractionations over a large range in magnitude. Calibrating sulfur isotopic fractionation in laboratory experiments allows for better interpretations of sulfur isotopes in modern sediments and ancient sedimentary rocks. The proteins involved in sulfate reduction are expressed in response to environmental conditions, and are collectively responsible for the net isotopic fractionation between sulfate and sulfide. We examined the role of DsrC, a key component of the sulfate reduction pathway, by comparing wildtype Desulfovibrio vulgaris DSM 644^T to strain IPFG07, a mutant deficient in DsrC production. Both strains were cultivated in parallel chemostat reactors at identical turnover times and cell specific sulfate reduction rates. Under these conditions, sulfur isotopic fractionations between sulfate and sulfide of 17.3 ± 0.5‰ or 12.6 ± 0.5‰ were recorded for the wildtype or mutant, respectively. The enzymatic machinery that produced these different fractionations was revealed by quantitative proteomics. Results are consistent with a cellular-level response that throttled the supply of electrons and sulfur supply through the sulfate reduction pathway more in the mutant relative to the wildtype, independent of rate. We conclude that the smaller fractionation observed in the mutant strain is a consequence of sulfate reduction that proceeded at a rate that consumed a greater proportion of the strains overall capacity for sulfate reduction. These observations have consequences for models of sulfate reducer metabolism and how it yields different isotopic fractionations, notably, the role of DsrC in central energy metabolism.

SUBMITTER: Leavitt WD

PROVIDER: S-EPMC6470260 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Proteomic and Isotopic Response of <i>Desulfovibrio vulgaris</i> to DsrC Perturbation.

Leavitt William D WD Venceslau Sofia S SS Waldbauer Jacob J Smith Derek A DA Pereira Inês A Cardoso IAC Bradley Alexander S AS

Frontiers in microbiology 20190411

Dissimilatory sulfate reduction is a microbial energy metabolism that can produce sulfur isotopic fractionations over a large range in magnitude. Calibrating sulfur isotopic fractionation in laboratory experiments allows for better interpretations of sulfur isotopes in modern sediments and ancient sedimentary rocks. The proteins involved in sulfate reduction are expressed in response to environmental conditions, and are collectively responsible for the net isotopic fractionation between sulfate ...[more]

PMID: 31031715

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Project description:Desulfovibrio vulgaris has been studied extensively for its potential in the bioremediation of heavy metals and radionuclides. Hydrocarbons and solvents, as frequent environmental co-contaminants, have been reported to inhibit microbial activities and thereby pose a limitation on bioremediation efficiency. As a part of the Genomes: GTL project to deduce the stress response pathways in metal/radionuclide reducing bacteria, we studied the responses of D. vulgaris to ethanol, which is a solvent and co-contaminant frequently encountered at contaminated DOE sites. Growth experiments in closed vessels at 37 °C indicated that D. vulgaris could maintain normal growth with 1%(v/v) ethanol. The growth rates were reduced with increasing ethanol concentrations at 2% and 5%. Growth ceased when ethanol concentration was raised to 5%(v/v). Cell lysis was apparent with decreasing optical density following 10% ethanol addition. To assess the mechanism of ethanol inhibition, genome-wide transcriptional profiles were analyzed from D. vulgaris cultures following ethanol (5% v/v) treatment using whole-genome microarrays. The ethanol treatment altered the expression of a large number of genes in the D. vulgaris genome, of which 354 were up-regulated greater than 2 fold and 217 were down-regulated by over 2 fold. As expected, changes in the transcriptional profile were similar to those of the stress response to acetone, which is also a solvent. Transcripts highly up-regulated included genes encoding the flagella structural subunits, suggesting motility as a mechanism of solvent stress response. Another group of genes highly induced were chaperones, such as dnaJ, groES, and hsp20, indicating the importance of maintaining proper protein folding under ethanol stress. Down-regulated genes included two groups of genes, ribosomal proteins and amino acid transporters, consistent with the growth inhibition by ethanol observed in growth studies. These results were interpreted that D. vulgaris responds to elevated solvent levels by increased motility and maintenance of proper protein functions. Current work is focused on the analysis of regulatory pathways based on temporal transcriptional dynamics. Keywords: Stress response

Project description:Desulfovibrio vulgaris Hildenborough belongs to a class of sulfate-reducing bacteria (SRB) and is found ubiquitously in nature. Given the importance of SRB-mediated reduction for bioremediation of metal ion contaminants, ongoing research on D. vulgaris has been in the direction of elucidating regulatory mechanisms for this organism under a variety of stress conditions. This work presents a global view of this organism's response to elevated growth temperature using whole-cell transcriptomics and proteomics tools. Transcriptional response (1.7-fold change or greater; Z >/= 1.5) ranged from 1,135 genes at 15 min to 1,463 genes at 120 min for a temperature up-shift of 13 degrees C from a growth temperature of 37 degrees C for this organism and suggested both direct and indirect modes of heat sensing. Clusters of orthologous group categories that were significantly affected included posttranslational modifications; protein turnover and chaperones (up-regulated); energy production and conversion (down-regulated), nucleotide transport, metabolism (down-regulated), and translation; ribosomal structure; and biogenesis (down-regulated). Analysis of the genome sequence revealed the presence of features of both negative and positive regulation which included the CIRCE element and promoter sequences corresponding to the alternate sigma factors sigma(32) and sigma(54). While mechanisms of heat shock control for some genes appeared to coincide with those established for Escherichia coli and Bacillus subtilis, the presence of unique control schemes for several other genes was also evident. Analysis of protein expression levels using differential in-gel electrophoresis suggested good agreement with transcriptional profiles of several heat shock proteins, including DnaK (DVU0811), HtpG (DVU2643), HtrA (DVU1468), and AhpC (DVU2247). The proteomics study also suggested the possibility of posttranslational modifications in the chaperones DnaK, AhpC, GroES (DVU1977), and GroEL (DVU1976) and also several periplasmic ABC transporters.

Project description:Desulfovibrio vulgaris has been studied extensively for its potential in the bioremediation of heavy metals and radionuclides. Hydrocarbons and solvents, as frequent environmental co-contaminants, have been reported to inhibit microbial activities and thereby pose a limitation on bioremediation efficiency. As a part of the Genomes: GTL project to deduce the stress response pathways in metal/radionuclide reducing bacteria, we studied the responses of D. vulgaris to acetone, which is a ketone solvent frequently observed at contaminated DOE sites. Growth experiments in closed vessels at 37 Â°C indicated that D. vulgaris could maintain normal growth with 3%(v/v) acetone following a 1-h lag phase. When the acetone concentration was raised to 5%(v/v), we observed a 2-h lag phase followed by a growth rate which was only 15% that of normal. At an acetone concentration of 8%(v/v), no active growth was observed following 10 hours of incubation. To assess the mechanism of acetone inhibition, genome-wide transcriptional profiles were analyzed from D. vulgaris cultures following acetone (5% v/v) treatment using whole-genome microarrays. This acetone shock altered the expression of a large number of genes in the D. vulgaris genome, of which 309 were up-regulated greater than 2 fold and 199 were down-regulated by over 2 fold. Transcripts highly up-regulated included genes encoding the flagella structural subunits, flgB (15 fold), fliE (11 fold), and flgH (10 fold). Another group of genes highly induced were chaperones, such as dnaJ (11 fold), groES (8 fold), and hsp20 (8 fold). Down-regulated genes included two groups of genes, ribosomal proteins and amino acid transporters, suggesting a state of growth arrest upon acetone addition. These results were interpreted to mean that D. vulgaris responds to elevated solvent levels by increased motility and maintenance of proper protein functions. Current work is focused on the analysis of regulatory pathways based on temporal transcriptional dynamics. Keywords: Stress response Desulfovibrio vulgaris from our laboratory culture collection was cultured at 37Â°C in Lactate-Sulfate medium to mid-log phase. Subsequently, acetone (5% v/v) was added into triplicate cultures. Gene expression profiles 30 and 100 minutes following acetone exposure were compared with those of the control cultures without acetone treatment. Total RNA was harvested from three replicate control and treated cultures at two time points following acetone addition. RNA extraction, purification, and labeling were performed independently on each cell sample.Two samples of each total RNA preparation were labeled, one with Cy3-dUTP and another with Cy5-dUTP for microarray hybridization (Dye swap).

Project description:Desulfovibrio vulgaris has been studied extensively for its potential in the bioremediation of heavy metals and radionuclides. Hydrocarbons and solvents, as frequent environmental co-contaminants, have been reported to inhibit microbial activities and thereby pose a limitation on bioremediation efficiency. As a part of the Genomes: GTL project to deduce the stress response pathways in metal/radionuclide reducing bacteria, we studied the responses of D. vulgaris to ethanol, which is a solvent and co-contaminant frequently encountered at contaminated DOE sites. Growth experiments in closed vessels at 37 Â°C indicated that D. vulgaris could maintain normal growth with 1%(v/v) ethanol. The growth rates were reduced with increasing ethanol concentrations at 2% and 5%. Growth ceased when ethanol concentration was raised to 5%(v/v). Cell lysis was apparent with decreasing optical density following 10% ethanol addition. To assess the mechanism of ethanol inhibition, genome-wide transcriptional profiles were analyzed from D. vulgaris cultures following ethanol (5% v/v) treatment using whole-genome microarrays. The ethanol treatment altered the expression of a large number of genes in the D. vulgaris genome, of which 354 were up-regulated greater than 2 fold and 217 were down-regulated by over 2 fold. As expected, changes in the transcriptional profile were similar to those of the stress response to acetone, which is also a solvent. Transcripts highly up-regulated included genes encoding the flagella structural subunits, suggesting motility as a mechanism of solvent stress response. Another group of genes highly induced were chaperones, such as dnaJ, groES, and hsp20, indicating the importance of maintaining proper protein folding under ethanol stress. Down-regulated genes included two groups of genes, ribosomal proteins and amino acid transporters, consistent with the growth inhibition by ethanol observed in growth studies. These results were interpreted that D. vulgaris responds to elevated solvent levels by increased motility and maintenance of proper protein functions. Current work is focused on the analysis of regulatory pathways based on temporal transcriptional dynamics. Keywords: Stress response Desulfovibrio vulgaris from our laboratory culture collection was cultured at 37Â°C in Lactate-Sulfate medium to mid-log phase. Subsequently, ethanol (5% v/v) was added into triplicate cultures. Gene expression profiles 30 and 100 minutes following ethanol exposure were compared with those of the control cultures without ethanol treatment. Total RNA was harvested from three replicate control and treated cultures at two time points following ethanol addition. RNA extraction, purification, and labeling were performed independently on each cell sample.Two samples of each total RNA preparation were labeled, one with Cy3-dUTP and another with Cy5-dUTP for microarray hybridization (Dye swap).

Dataset Information

Proteomic and Isotopic Response of Desulfovibrio vulgaris to DsrC Perturbation.

Publications

Proteomic and Isotopic Response of <i>Desulfovibrio vulgaris</i> to DsrC Perturbation.

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