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N-Deacetylases required for muramic-?-lactam production are involved in Clostridium difficile sporulation, germination, and heat resistance.

ABSTRACT: Spores are produced by many organisms as a survival mechanism activated in response to several environmental stresses. Bacterial spores are multilayered structures, one of which is a peptidoglycan layer called the cortex, containing muramic-?-lactams that are synthesized by at least two bacterial enzymes, the muramoyl-l-alanine amidase CwlD and the N-deacetylase PdaA. This study focused on the spore cortex of Clostridium difficile, a Gram-positive, toxin-producing anaerobic bacterial pathogen that can colonize the human intestinal tract and is a leading cause of antibiotic-associated diarrhea. Using ultra-HPLC coupled with high-resolution MS, here we found that the spore cortex of the C. difficile 630?erm strain differs from that of Bacillus subtilis Among these differences, the muramic-?-lactams represented only 24% in C. difficile, compared with 50% in B. subtilis CD630_14300 and CD630_27190 were identified as genes encoding the C. difficile N-deacetylases PdaA1 and PdaA2, required for muramic-?-lactam synthesis. In a pdaA1 mutant, only 0.4% of all muropeptides carried a muramic-?-lactam modification, and muramic-?-lactams were absent in the cortex of a pdaA1-pdaA2 double mutant. Of note, the pdaA1 mutant exhibited decreased sporulation, altered germination, decreased heat resistance, and delayed virulence in a hamster infection model. These results suggest a much greater role for muramic-?-lactams in C. difficile than in other bacteria, including B. subtilis In summary, the spore cortex of C. difficile contains lower levels of muramic-?-lactams than that of B. subtilis, and PdaA1 is the major N-deacetylase for muramic-?-lactam biosynthesis in C. difficile, contributing to sporulation, heat resistance, and virulence.

SUBMITTER: Coullon H

PROVIDER: S-EPMC6254358 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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N-Deacetylases required for muramic-δ-lactam production are involved in Clostridium difficile sporulation, germination, and heat resistance.

Coullon Héloise H Rifflet Aline A Wheeler Richard R Janoir Claire C Boneca Ivo Gomperts IG Candela Thomas T

The Journal of biological chemistry 20180928 47

Spores are produced by many organisms as a survival mechanism activated in response to several environmental stresses. Bacterial spores are multilayered structures, one of which is a peptidoglycan layer called the cortex, containing muramic-δ-lactams that are synthesized by at least two bacterial enzymes, the muramoyl-l-alanine amidase CwlD and the N-deacetylase PdaA. This study focused on the spore cortex of Clostridium difficile, a Gram-positive, toxin-producing anaerobic bacteri ...[more]

PMID: 30266804

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Project description:UnlabelledClostridium difficile must form a spore to survive outside the gastrointestinal tract. The factors that trigger sporulation in C. difficile remain poorly understood. Previous studies have suggested that a link exists between nutritional status and sporulation initiation in C. difficile In this study, we investigated the impact of the global nutritional regulator CodY on sporulation in C. difficile strains from the historical 012 ribotype and the current epidemic 027 ribotype. Sporulation frequencies were increased in both backgrounds, demonstrating that CodY represses sporulation in C. difficile The 027 codY mutant exhibited a greater increase in spore formation than the 012 codY mutant. To determine the role of CodY in the observed sporulation phenotypes, we examined several factors that are known to influence sporulation in C. difficile Using transcriptional reporter fusions and quantitative reverse transcription-PCR (qRT-PCR) analysis, we found that two loci associated with the initiation of sporulation, opp and sinR, are regulated by CodY. The data demonstrate that CodY is a repressor of sporulation in C. difficile and that the impact of CodY on sporulation and expression of specific genes is significantly influenced by the strain background. These results suggest that the variability of CodY-dependent regulation is an important contributor to virulence and sporulation in current epidemic isolates. This report provides further evidence that nutritional state, virulence, and sporulation are linked in C. difficileImportanceThis study sought to examine the relationship between nutrition and sporulation in C. difficile by examining the global nutritional regulator CodY. CodY is a known virulence and nutritional regulator of C. difficile, but its role in sporulation was unknown. Here, we demonstrate that CodY is a negative regulator of sporulation in two different ribotypes of C. difficile We also demonstrate that CodY regulates known effectors of sporulation, Opp and SinR. These results support the idea that nutrient limitation is a trigger for sporulation in C. difficile and that the response to nutrient limitation is coordinated by CodY. Additionally, we demonstrate that CodY has an altered role in sporulation regulation for some strains.

Dataset Information

N-Deacetylases required for muramic-?-lactam production are involved in Clostridium difficile sporulation, germination, and heat resistance.

Publications

<i>N</i>-Deacetylases required for muramic-δ-lactam production are involved in <i>Clostridium difficile</i> sporulation, germination, and heat resistance.

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