Global analysis of genes induced during sporulation of Clostridium difficile
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ABSTRACT:
ORGANISM(S): Clostridioides difficile 630
PROVIDER: GSE45977 | GEO | 2013/08/10
SECONDARY ACCESSION(S): PRJNA196825
REPOSITORIES: GEO
Similar Datasets
Project description:Purpose: A goal of this study was to identify genes induced in the mother cell during Clostridium difficile sporulation (specifically in a σE-, σK-, and SpoIIID-dependent manner). Methods: Whole genome RNA sequencing was performed on wildtype, sigE-, sigK- and spoIIID- C. difficile strains (strain 630 background; JIR8094 = parent strain), and the transcriptional profiles of the different mutants during growth on 70:30 agar plates were determined using an Illumina MiSeq1000. Results: This analysis identified 200 genes whose expression is collectively activated by sporulation sigma factors: 159 were σF-dependent, 162 were σE-dependent, 28 were σG-dependent, and 36 were σK-dependent. A total of 254 genes were identified as requiring Spo0A for their expression when sporulation was induced on the 70:30 plates. Conclusions: These results provide the first genome-wide transcriptional analysis of genes whose expression is induced by specific sporulation sigma factors in the Clostridia and highlight that diverse mechanisms regulate sporulation sigma factor activity in the Firmicutes. For example, in contrast with the B. subtilis sporulation pathway, C. difficile σE was not required to fully activate σG, and σG was not required to activate σK.
2014-12-02 | GSE63777 | GEO
Project description:Purpose: The goal of this study was to identify genes whose expression is induced during sporulation in a Spo0A-, M-OM-^CF-, M-OM-^CE-, M-OM-^CG-, and M-OM-^CK-dependent manner. Methods: Whole genome RNA sequencing was performed on wildtype, spo0A-, sigF-, sigE-, sigG-, and sigK- C. difficile strains (strain 630 background; JIR8094 = parent strain), and the transcriptional profiles of the different mutants during growth on 70:30 agar plates were determined using an Illumina HiSeq1000. Results: This analysis identified 185 genes whose expression is collectively activated by sporulation sigma factors: 150 were M-OM-^CF-dependent, 150 were M-OM-^CE-dependent, 30 were M-OM-^CG-dependent, and 31 were M-OM-^CK-dependent. A total of 237 genes were identified as requiring Spo0A for their expression when sporulation was induced on the 70:30 plates. Conclusions: These results provide the first genome-wide transcriptional analysis of genes whose expression is induced by specific sporulation sigma factors in the Clostridia and highlight that diverse mechanisms regulate sporulation sigma factor activity in the Firmicutes. For example, in contrast with the B. subtilis sporulation pathway, C. difficile M-OM-^CE was not required to fully activate M-OM-^CG, and M-OM-^CG was not required to activate M-OM-^CK. Wildtype, spo0A-, sigF-, sigE-, sigG-, and sigK- were streaked onto 70:30 plates in triplicate, and sporulation was induced. RNA from the three biological replicate samples was harvested, DNAse-I treated, and ribosomally depleted using Ribo-Zero kits. This RNA was transformed into cDNA, and the resulting libraries were subjected to paired end sequencing using an Illumina HiSeq 1000.
2013-08-10 | E-GEOD-45977 | biostudies-arrayexpress
Project description:Clostridium acetobutylicum, the endospore-forming anaerobe best known for its ABE (acetone-butanol-ethanol) fermentation, has received renewed attention recently for the biological production of butanol, both for bulk chemical production and as a potential biofuel. With butanol production in mind, most of the recent research on C. acetobutylicum has focused on increasing butanol production, tolerance to butanol, and optimizing it for various substrates. However, an equally important trait, though less understood, is its sporulation program, which it coupled to solvent formation. The model organism for endospore formation is Bacillus subtilis, but significant physiological, metabolic, and genomic differences exist between the two organisms. Despite these differences, the major sporulation-related transcription/sigma factors are conserved between the two species. These transcription/sigma factors are activated in a cascade manner such that Spo0A becomes active first, followed by σF, then σE, then σG, and finally σK. The goal of this study is to determine the regulons of 4 of these transcription/sigma factors (Spo0A, σF, σE, and σG) and compare them to those in B. subtilis. To accomplish this goal, individual mutant strains were created for Spo0A, σF, σE, and σG, in which the transcription/sigma factor is silenced. These mutants were then compared transcriptionally using microarrays to determine the regulon of each transcription/sigma factor. To help avoid false positives, comparisons were made between strains in which the downstream transcription/sigma factor is silenced (e.g., for the Spo0A regulon, the Spo0A mutant was compared against the σF mutant and the σE mutant, since they are both upregulated by Spo0A) rather than just the WT. For each regulon, 4 timepoints were taken, since it is very difficult to synchronize sporulation in C. acetobutylicum cultures, and dye-swaps were prepared for each timepoint.
2011-02-28 | GSE24103 | GEO
Project description:The obligate anaerobic, enteric pathogen Clostridioides difficile persists in the intestinal tract by forming antibiotic resistant endospores that contribute to relapsing and recurrent infections. Despite the importance of sporulation for C. difficile pathogenesis, environmental cues, and molecular mechanisms regulating sporulation initiation remain ill defined. Here, using RIL-seq to capture the Hfq-dependent RNA-RNA interactome, we discovered a network of small RNAs that bind to mRNAs encoding sporulation-related genes. We show that two of these small RNAs, SpoX and SpoY, regulate translation of the master regulator of sporulation, Spo0A, in an opposing manner, which ultimately leads to altered sporulation rates. Infection of antibiotic-treated mice with SpoX and SpoY deletion mutants revealed a global effect on gut colonization and intestinal sporulation. Our work uncovers an elaborate RNA-RNA interactome controlling the physiology and virulence of C. difficile and identifies a complex post-transcriptional layer in the regulation of spore formation in this important human pathogen.
2022-12-31 | GSE213005 | GEO
Project description:Clostridium acetobutylicum, the endospore-forming anaerobe best known for its ABE (acetone-butanol-ethanol) fermentation, has received renewed attention recently for the biological production of butanol, both for bulk chemical production and as a potential biofuel. With butanol production in mind, most of the recent research on C. acetobutylicum has focused on increasing butanol production, tolerance to butanol, and optimizing it for various substrates. However, an equally important trait, though less understood, is its sporulation program, which it coupled to solvent formation. The model organism for endospore formation is Bacillus subtilis, but significant physiological, metabolic, and genomic differences exist between the two organisms. Despite these differences, the major sporulation-related transcription/sigma factors are conserved between the two species. These transcription/sigma factors are activated in a cascade manner such that Spo0A becomes active first, followed by M-OM-^CF, then M-OM-^CE, then M-OM-^CG, and finally M-OM-^CK. The goal of this study is to determine the regulons of 4 of these transcription/sigma factors (Spo0A, M-OM-^CF, M-OM-^CE, and M-OM-^CG) and compare them to those in B. subtilis. To accomplish this goal, individual mutant strains were created for Spo0A, M-OM-^CF, M-OM-^CE, and M-OM-^CG, in which the transcription/sigma factor is silenced. These mutants were then compared transcriptionally using microarrays to determine the regulon of each transcription/sigma factor. To help avoid false positives, comparisons were made between strains in which the downstream transcription/sigma factor is silenced (e.g., for the Spo0A regulon, the Spo0A mutant was compared against the M-OM-^CF mutant and the M-OM-^CE mutant, since they are both upregulated by Spo0A) rather than just the WT. For each regulon, 4 timepoints were taken, since it is very difficult to synchronize sporulation in C. acetobutylicum cultures, and dye-swaps were prepared for each timepoint. Four transcription/sigma factor regulons were investigated: Spo0A, M-OM-^CF, M-OM-^CE, and M-OM-^CG. For Spo0A, two comparison were made: Spo0A mutant vs M-OM-^CF mutant and Spo0A mutant vs M-OM-^CE mutant. For M-OM-^CF, two comparison were also made: M-OM-^CF mutant vs M-OM-^CE mutant and M-OM-^CF mutant vs M-OM-^CG mutant. Finally for M-OM-^CE and M-OM-^CG, only one comparison for each was made: M-OM-^CE mutant vs M-OM-^CG mutant and M-OM-^CG mutant vs wild-type, respectively. For each comparison, 4 timepoints were analyzed, and dye-swaps were prepared for each timepoint comparison. Timepoints were chosen based on when each transcription/sigma factor was expected to be active, based on a previous study [Jones, SW, et al. Genome Biol. 2008;9(7):R114.].
2011-02-28 | E-GEOD-24103 | biostudies-arrayexpress
Project description:The universally conserved protein Elongation Factor P facilitates translation at amino acids that form peptide bonds with low efficiency, particularly poly-proline tracts. Despite its wide conservation, it is not essential in most bacteria and its physiological role remains unclear. Here, we show that EF-P affects the process of sporulation initiation in the bacterium Bacillus subtilis. We observe that lack of EF-P delays expression of sporulation-specific genes. Using ribosome profiling, we observe that expression of spo0A, encoding a transcription factor that functions as the master regulator of sporulation, is lower in a ∆efp strain as compared to the wildtype. Ectopic expression of Spo0A rescues the sporulation initiation phenotype, indicating that reduced spo0A expression explains the sporulation defect in ∆efp cells. Since Spo0A is the earliest sporulation transcription factor, these data suggest that sporulation initiation can be delayed when protein synthesis is impaired.
2023-01-04 | GSE222121 | GEO
Project description:Clostridium difficile is an emergent human pathogen and the most common cause of nosocomial diarrhea. Our recent deep-sequencing data strongly suggest the importance of RNA-based mechanisms for the control of gene expression in C. difficile. The RNA chaperon protein Hfq is a key component of post-transcriptional regulatory network in various bacterial systems. In an effort to understand the function of Hfq in C. difficile, we have constructed and characterized an Hfq-depleted strain in this bacterium. We present evidence that Hfq is necessary for the normal cell growth, for the establishment of bacillary form and might be involved in metabolic adaptations, stress response and sporulation, important during C. difficile infection cycle. In accordance to these phenotypic changes, our transcriptome analysis revealed pleiotropic effects of Hfq on gene expression in C. difficile. This microarray analysis showed altered expression of genes encoding cell wall and membrane transport components, sporulation proteins, as well as stress response, transcriptional regulator genes and genes of unknown function. Remarkably, a great number of genes belonging to the regulon dependent on sporulation-specific sigma K factor were generally up-regulated in the strain depleted for Hfq. Many important metabolic pathways were also affected by Hfq depletion. Finally, the altered accumulation of several previously identified sRNAs suggests potential involvement of Hfq in these regulatory RNA function. Altogether, these results provide evidence for the pleiotropic role of Hfq protein in C. difficile physiology expanding our knowledge of Hfq-dependent regulation in Gram-positive bacteria.
2014-07-15 | GSE49168 | GEO
Project description:Clostridium difficile is an important nosocomial pathogen and the leading cause of hospital-acquired diarrhea. Antibiotic use is the primary risk factor for the development of C. difficile-associated disease because it disrupts normal protective gut flora and enables C. difficile to colonize the colon. C. difficile damages host tissue by secreting toxins and disseminates by forming spores. The toxin-encoding genes, tcdA and tcdB are part of a pathogenicity locus, which also encodes the gene tcdR that codes for the toxin genes positive regulator. TcdR is an alternate sigma factor that initiates transcription of tcdA and tcdB at their promoters. Alternative sigma factors are known to regulate virulence and virulence associated genes in many pathogenic bacteria. We created a tcdR mutant in the epidemic-type C. difficile R20291 strain in an attempt to identify the global role of tcdR. A site-directed mutation in tcdR affected both toxin production and sporulation in C. difficile R20291. Spores derived from the tcdR mutant were found to be mildly temperature sensitive. Moreover, nearly two fold more taurocholate was needed to germinate spores from the tcdR mutant than the spores prepared from the wild-type parent strain. Comparison of the tcdR mutant transcriptome with the parent strain revealed many differentially expressed late sporulation genes in the tcdR mutant. These data suggests that gene regulatory networks of toxin production and sporulation in Clostridium difficile are linked with each other.
2017-03-07 | GSE85395 | GEO
Project description:Clostridium difficile is an emergent human pathogen and the most common cause of nosocomial diarrhea. Our recent deep-sequencing data strongly suggest the importance of RNA-based mechanisms for the control of gene expression in C. difficile. The RNA chaperon protein Hfq is a key component of post-transcriptional regulatory network in various bacterial systems. In an effort to understand the function of Hfq in C. difficile, we have constructed and characterized an Hfq-depleted strain in this bacterium. We present evidence that Hfq is necessary for the normal cell growth, for the establishment of bacillary form and might be involved in metabolic adaptations, stress response and sporulation, important during C. difficile infection cycle. In accordance to these phenotypic changes, our transcriptome analysis revealed pleiotropic effects of Hfq on gene expression in C. difficile. This microarray analysis showed altered expression of genes encoding cell wall and membrane transport components, sporulation proteins, as well as stress response, transcriptional regulator genes and genes of unknown function. Remarkably, a great number of genes belonging to the regulon dependent on sporulation-specific sigma K factor were generally up-regulated in the strain depleted for Hfq. Many important metabolic pathways were also affected by Hfq depletion. Finally, the altered accumulation of several previously identified sRNAs suggests potential involvement of Hfq in these regulatory RNA function. Altogether, these results provide evidence for the pleiotropic role of Hfq protein in C. difficile physiology expanding our knowledge of Hfq-dependent regulation in Gram-positive bacteria. Two-conditions experiments, 630E strain vs. hfQ mutant grown in TYt, 4 biological replicates for each condition.
2014-07-15 | E-GEOD-49168 | biostudies-arrayexpress
Project description:Genes encoding the alternative sigma factor SigmaB, involved in the general stress response in firmicutes, as well as its regulators are present in the genome of the enteropathogen Clostridium difficile. We inactivated the sigB gene using the ClosTron technology in order to identify the role of this sigma factor in the physiology of this bacterium. Transcriptomic experiments showed that SigB positively and negatively controlls several genes involved in different cellular processes such as metabolism, sporulation and stress response.
2017-07-07 | GSE85981 | GEO