The Flagellin-Specific Chaperone FliS of Borrelia burgdorferi Controls the Cytoplasmic Pool of Flagellins at the Level of Translation Initiation, Secretion, and Proteolysis.

Abstract

The flagellin-specific chaperone FliS has been studied in externally flagellated bacteria; however, its role in spirochetes, a group of bacteria that possess unique internalized flagella (termed endo- or periplasmic flagella), remains unexplored. Here, we investigate the function of FliS in the Lyme disease spirochete Borrelia burgdorferi. Using loss-of-function studies, combined with biochemical assays and cryo-electron tomography, we demonstrate that FliS deletion selectively reduces FlaB expression, the major flagellin protein, resulting in non-motile mutants with defective flagellar filaments. Mechanistically, we show that FlaB interacts with both FliS and FliW, the latter being an allosteric repressor of the RNA-binding protein CsrA, which inhibits FlaB translation. These four components form a regulatory circuit that fine-tunes FlaB levels and flagellar assembly via a partner-switching mechanism. Deletion of fliS disrupts FlaB secretion, leading to its cytoplasmic accumulation, sequestration of FliW, and subsequent release of CsrA to suppress FlaB synthesis. Accumulation of cytoplasmic FlaB also triggers its degradation to prevent toxicity. Our findings reveal a post-transcriptional regulatory mechanism governing flagellar assembly in B. burgdorferi, an evolutionary outlier that lacks the canonical transcriptional cascade controlling flagellar biosynthesis in most bacteria.