CwlQ is required for swarming motility but not flagellar assembly in<i>Bacillus subtilis</i>

Kearns, Daniel B.; Sánchez, Sandra; Dunn, Caroline

doi:10.1101/2021.01.13.426625

Cited by 2 publications

(1 citation statement)

References 99 publications

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“…Seemingly missing from the genome are homologs for the rod-cap protein that catalyzes rod polymerization and bushing proteins that surround the rod to stabilize flagellar rotation in the context of the envelope. Finally, while B. subtilis encodes many hydrolase enzymes that remodel peptidoglycan, there is currently no known hydrolase required to create space in the wall for rod assembly (23,24). Thus in B. subtilis , the “missing parts” are either unnecessary or await discovery.…”

Section: Introductionmentioning

confidence: 99%

Identification of genes required for swarming motility in Bacillus subtilis using transposon mutagenesis and high-throughput sequencing (TnSeq)

Sánchez

Snider

Wang

et al. 2022

Preprint

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Bacillus subtilis exhibits swarming motility, a flagellar-mediated form of surface motility. Here we use high-throughput transposon mutagenesis and sequencing (TnSeq) to identify candidate genes required for swarming. The TnSeq approach identified all of the known genes required for flagellar biosynthesis as well as nearly all of the previously reported regulators that promote swarming. Moreover, we identified an additional 36 genes that improve swarming and validated them individually. Among these, two mutants with severe defects were recovered including fliT, required for flagellar biosynthesis, and a gene of unknown function yolB, whose defect could not be attributed to a lack of flagella. Our work indicates that TnSeq is a powerful approach for the identification of genes required for motility that drives outward expansion of a colony on plates.IMPORTANCEIn TnSeq, transposons are randomly inserted throughout the chromosome at a population level, but insertions that disrupt genes of essential function cause strains that carry them to fall out of the population and appear under-represented at the sequence level. Here we apply TnSeq to the non-essential phenotype of motility in B. subtilis and spatially select for cells proficient in swarming. We find that insertions in nearly all genes previously identified as required for swarming are under-represented in TnSeq analysis, and we identify 36 additional genes that enhance swarming. We suggest that TnSeq is a powerful tool for the genetic analysis of motility and likely other non-lethal screens for which enrichment is strong.

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Section: Introductionmentioning

confidence: 99%

Identification of genes required for swarming motility in Bacillus subtilis using transposon mutagenesis and high-throughput sequencing (TnSeq)

Sánchez

Snider

Wang

et al. 2022

Preprint

Self Cite

View full text Add to dashboard Cite

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An Exhaustive Multiple Knockout Approach to Understanding Cell Wall Hydrolase Function inBacillus subtilis

Wilson

Garner

2021

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Most bacteria are surrounded by their cell wall, a highly crosslinked protective envelope of peptidoglycan. To grow, bacteria must continuously remodel their wall, inserting new material and breaking old bonds. Bond cleavage is performed by cell wall hydrolases, allowing the wall to expand. Understanding the functions of individual hydrolases has been impeded by their redundancy: single knockouts usually present no phenotype. We used an exhaustive multiple-knockout approach to determine the minimal set of hydrolases required for growth in Bacillus subtilis. We identified 42 candidate cell wall hydrolases. Strikingly, we were able to remove all but two of these genes in a single strain; this 'Δ40' strain shows a normal growth rate, indicating that none of the 40 hydrolases are necessary for cell growth. The Δ40 strain does not shed old cell wall, demonstrating that turnover is not essential for growth. The remaining two hydrolases in the Δ40 strain are LytE and CwlO, previously shown to be synthetically lethal. Either can be knocked out in Δ40, indicating that either hydrolase alone is sufficient for cell growth. Environmental screening and zymography revealed that LytE activity is inhibited by Mg2+ and that RlpA-like proteins may stimulate LytE activity. Together, these results demonstrate that the only essential function of cell wall hydrolases in B. subtilis is to enable cell growth by expanding the wall and that LytE or CwlO alone is sufficient for this function. These experiments introduce the Δ40 strain as a tool to study hydrolase activity and regulation in B. subtilis.

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CwlQ is required for swarming motility but not flagellar assembly inBacillus subtilis

Cited by 2 publications

References 99 publications

Identification of genes required for swarming motility in Bacillus subtilis using transposon mutagenesis and high-throughput sequencing (TnSeq)

Identification of genes required for swarming motility in Bacillus subtilis using transposon mutagenesis and high-throughput sequencing (TnSeq)

An Exhaustive Multiple Knockout Approach to Understanding Cell Wall Hydrolase Function inBacillus subtilis

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