Carbon storage regulator A (CsrABb) is a repressor of Borrelia burgdorferi flagellin protein FlaB

Abstract

SUMMARY The Lyme disease spirochete Borrelia burgdorferi lacks the transcriptional cascade control of flagellar protein synthesis common to other bacteria. Instead, it relies on a post-transcriptional mechanism to control its flagellar synthesis. The underlying mechanism of this control remains elusive. A recent study reported that the increased level of BB0184 (CsrABb; a homolog of carbon storage regulator A) substantially inhibited the accumulation of FlaB, the major flagellin protein of B. burgdorferi. In this report, we deciphered the regulatory role of CsrABb on FlaB synthesis and the mechanism involved by analyzing two mutants, csrABb− (a deletion mutant of csrABb) and csrABb+ (a mutant conditionally over-expressing csrABb). We found that FlaB accumulation was significantly inhibited in csrABb+ but was substantially increased in csrABb−. In contrast, the levels of other flagellar proteins remained unchanged. Cryo-electron tomography and immuno-fluorescence microscopic analyses revealed that the altered synthesis of CsrABb in these two mutants specifically affected flagellar filament length. The leader sequence of flaB transcript contains two conserved CsrA-binding sites, with one of these sites overlapping the Shine-Dalgarno sequence. We found that CsrABb bound to the flaB transcripts via these two binding sites, and this binding inhibited the synthesis of FlaB at the translational level. Taken together, our results indicate that CsrABb specifically regulates the periplasmic flagellar synthesis by inhibiting translation initiation of the flaB transcript.

FliW antagonizes CsrA RNA binding by a noncompetitive allosteric mechanism

“…Recent work in the distantly related C. jejuni is consistent with the B. subtilis model, in which FliW antagonizes CsrA and CsrA inhibits flagellin translation (39). Thus, the noncompetitive mechanism of inhibition that governs the CsrA-FliW-flagellin module is likely to be directly applicable to all FliW-encoding organisms, including members of the firmicutes, the spirochetes, and the deep branches of the proteobacteria (34,39,(45)(46)(47)(48). In contrast, competitive inhibition of CsrA by sRNA has been extensively studied, but only in bacteria belonging to the family of γ-proteobacteria (29).…”

“…The phylogenetic distribution of sRNA competitors is poorly understood because of difficulties in predicting sRNA presence, expression, and function, but the conservation of the BarA-UvrY two component system that regulates csrB and csrC expression is largely restricted to the γ-proteobacteria, suggesting the sRNA mechanism of CsrA antagonism is narrowly distributed (34,38,49). Finally we note that, whether an organism uses sRNAs or FliW for regulation, CsrA regulates virulence factors in each pathogen in which it has been studied (1,(45)(46)(47)(48). We suggest that CsrA is a candidate therapeutic target and that the precedent for noncompetitive inhibition can be exploited to isolate noncompetitive inhibitors, which are typically effective at lower doses than competitive counterparts.…”

Motility Is Crucial for the Infectious Life Cycle of Borrelia burgdorferi

“…Furthermore, antibodies against FlaB are detected in all stages (early or late) of Lyme disease (93). Although flaB is expressed throughout the spirochete's life cycle, recent reports indicate that the translation of flaB is regulated by carbon storage regulator A (CsrA) (94,95), suggesting that motility is likely to be necessary for a specific stage of the organism's infectious cycle (see below).…”

FliW and FliS Function Independently To Control Cytoplasmic Flagellin Levels in Bacillus subtilis

“…This mechanism suggests that the Hag protein is not only structural but also regulatory and that Hag expression is coupled to secretion. The homeostatic restriction of flagellin translation may also be functioning by a ho-mologous system in Borrelia burgdorferi and perhaps other organisms (23,25,26).…”