Abstract
The assembly of the bacterial flagellum is exquisitely controlled. Flagellar biosynthesis is underpinned by a specialized type III secretion system that allows export of proteins from the cytoplasm to the nascent structure. Bacillus subtilis regulates flagellar assembly using both conserved and species-specific mechanisms. Here, we show that YvyG is essential for flagellar filament assembly. We define YvyG as an orthologue of the Salmonella enterica serovar Typhimurium type III secretion system chaperone, FlgN, which is required for the export of the hook-filament junction proteins, FlgK and FlgL. Deletion of flgN (yvyG) results in a nonmotile phenotype that is attributable to a decrease in hag translation and a complete lack of filament polymerization. Analyses indicate that a flgK-flgL double mutant strain phenocopies deletion of flgN and that overexpression of flgK-flgL cannot complement the motility defect of a ΔflgN strain. Furthermore, in contrast to previous work suggesting that phosphorylation of FlgN alters its subcellular localization, we show that mutation of the identified tyrosine and arginine FlgN phosphorylation sites has no effect on motility. These data emphasize that flagellar biosynthesis is differentially regulated in B. subtilis from classically studied Gram-negative flagellar systems and questions the biological relevance of some posttranslational modifications identified by global proteomic approaches.
Highlights
The assembly of the bacterial flagellum is exquisitely controlled
The basal body consists of the flagellar motor, which is required to power rotation of the flagellum, and a type III secretion (T3S) system that permits the export of proteins required for the biosynthesis of the hook and filament
Through the use of site-directed mutagenesis, we demonstrate that mutation of the tyrosine and arginine phosphorylation sites of FlgN has no effect on the ability of B. subtilis to become motile
Summary
The assembly of the bacterial flagellum is exquisitely controlled. Flagellar biosynthesis is underpinned by a specialized type III secretion system that allows export of proteins from the cytoplasm to the nascent structure. We define YvyG as an orthologue of the Salmonella enterica serovar Typhimurium type III secretion system chaperone, FlgN, which is required for the export of the hook-filament junction proteins, FlgK and FlgL. The basal body consists of the flagellar motor, which is required to power rotation of the flagellum, and a type III secretion (T3S) system that permits the export of proteins required for the biosynthesis of the hook and filament. Given the potential for YvyG to play a crucial role in the tightly controlled process of flagellar biosynthesis [31, 36], we hypothesized that protein phosphorylation might present an additional route for B. subtilis to regulate flagellar assembly and, motility. We aimed to define the function of YvyG during motility by B. subtilis and to ascertain the in vivo role of YvyG tyrosine and arginine phosphorylation
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