The Phosphotransfer Protein CD1492 Represses Sporulation Initiation in Clostridium difficile

Abstract

The formation of spores is critical for the survival of Clostridium difficile outside the host gastrointestinal tract. Persistence of C. difficile spores greatly contributes to the spread of C. difficile infection (CDI), and the resistance of spores to antimicrobials facilitates the relapse of infection. Despite the importance of sporulation to C. difficile pathogenesis, the molecular mechanisms controlling spore formation are not well understood. The initiation of sporulation is known to be regulated through activation of the conserved transcription factor Spo0A. Multiple regulators influence Spo0A activation in other species; however, many of these factors are not conserved in C. difficile and few novel factors have been identified. Here, we investigated the function of a protein, CD1492, that is annotated as a kinase and was originally proposed to promote sporulation by directly phosphorylating Spo0A. We found that deletion of CD1492 resulted in increased sporulation, indicating that CD1492 is a negative regulator of sporulation. Accordingly, we observed increased transcription of Spo0A-dependent genes in the CD1492 mutant. Deletion of CD1492 also resulted in decreased toxin production in vitro and in decreased virulence in the hamster model of CDI. Further, the CD1492 mutant demonstrated effects on gene expression that are not associated with Spo0A activation, including lower sigD and rstA transcription, suggesting that this protein interacts with factors other than Spo0A. Altogether, the data indicate that CD1492 negatively affects sporulation and positively influences motility and virulence. These results provide further evidence that C. difficile sporulation is regulated differently from that of other endospore-forming species.

The alternative sigma factor σB plays a crucial role in adaptive strategies of Clostridium difficile during gut infection

“…Interestingly, CD1492 , encoding a kinase sharing 30% identity with Cac0437, was down‐regulated in the sigB mutant. Furthermore, it was recently reported that CD1492 negatively controls sporulation (Childress et al ., ). In conclusion, we propose that σ B controls sporulation initiation by modulating the level of phosphorylation of Spo0A as suggested in B. subtilis .…”

Ethanolamine is a valuable nutrient source that impacts Clostridium difficile pathogenesis

“…AM180355). CLC Genomics Workbench v9.0 was used for all variant analysis, as previously described (Childress et al, 2016). No nucleotide changes were observed between the mutant and parent strain genomes.…”

Phase variation of a signal transduction system controlsClostridioides difficilecolony morphology, motility, and virulence

“…C. difficile cultures were grown in BHIS medium supplemented with 0.1% taurocholate and 0.2% fructose until mid-exponential phase (i.e., OD 600 of 0.5), and 0.25 ml aliquots were plated onto 70:30 agar supplemented with 2 µg/ml thiamphenicol (79). After 24 h growth, ethanol resistance assays were performed as previously described (84, 85). Briefly, the cells were removed from plates after 24 h (H 24 ) and suspended in BHIS medium to an OD 600 of ∼1.0.…”