Abstract
In Bacillus subtilis, motility genes are expressed in a hierarchical pattern – governed by the σD transcription factor and other proteins such as the EpsE molecular clutch and SlrA/SlrR regulator proteins. In contrast, motile species in the Bacillus cereus group seem to express their motility genes in a non-hierarchical pattern, and less is known about their regulation, also given that no orthologs to σD, EpsE, SlrA or SlrR are found in B. cereus group genomes. Here we show that deletion of cdgL (BTB_RS26690/BTB_c54300) in Bacillus thuringiensis 407 (cry-) resulted in a six-to ten-fold downregulation of the entire motility locus, and loss of flagellar structures and swimming motility. cdgL is unique to the B. cereus group and is found in all phylogenetic clusters in the population except for group I, which comprises isolates of non-motile Bacillus pseudomycoides. Analysis of RNA-Seq data revealed cdgL to be expressed in a three-gene operon with a NupC like nucleoside transporter, and a putative glycosyl transferase for which transposon-based gene inactivation was previously shown to produce a similar phenotype to cdgL deletion. Interestingly, all three proteins were predicted to be membrane-bound and may provide a concerted function in the regulation of B. cereus group motility.
Highlights
The ability to receive and respond to environmental signals is imperative for the survival of bacteria
Low copy number expression/shuttle vector; xylA promoter (Ampr, Eryr) Wild type cdgL gene from B. thuringiensis 407 cloned in vector pHT304-Pxyl Integrative plasmid vector used for allelic exchange and construction of gene deletion strains for Bacillus cereus group bacteria (Ampr) Helper plasmid for gene deletion mutant construction, expressing the I-SceI restriction endonuclease (Kmr) GGDEF domain from the wild type cdgL gene in B. thuringiensis 407 cloned in expression vector pET-11 (Kmr)
CdgL is essential for flagellar motility in B. thuringiensis
Summary
The ability to receive and respond to environmental signals is imperative for the survival of bacteria. The key transcriptional regulator of virulence genes, PlcR, affects motility and biofilm formation [17,18]. B. cereus group bacteria were recently shown to carry a range of c-di-GMP metabolism genes, affecting phenotypes such as motility, biofilm and virulence [19]. The effector molecules can bind c-di-GMP through a variety of domains, including GEMM-I riboswitches and PilZ protein domains [22,23] Proteins such as PelD from Pseudomonas aeruginosa, which regulates the biosynthesis of exopolysaccharides [24], and PopA which promotes cell cycle progression in Caulobacter crescentus [25], have been shown to bind c-di-GMP through degenerate and thereby enzymatically inactive GGDEF domains, thereby acting as downstream effectors responding to changes in cellular c-di-GMP levels and mediating c-di-GMP-dependent phenotypes [21]. B. thuringiensis 407, a model strain for functional studies of B. cereus and B. thuringiensis
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