Feedback control of Campylobacter jejuni flagellin levels through reciprocal binding of FliW to flagellin and the global regulator CsrA

Abstract

Summary Bacterial flagella assembly is tightly regulated to ensure a timely and sequential production of the various flagellum constituents. In the pathogen Campylobacter jejuni the hierarchy in flagella biosynthesis is largely determined at the transcriptional level through the activity of the alternative sigma factors sigma54 and sigma28. Here, we report that C. jejuni flagellin levels are also controlled at the post‐transcriptional level via the thus far poorly‐characterized flagellar assembly factor FliW. Analysis of flagellin synthesis in C. jejuni 81116 and a ΔfliW knock‐out mutant showed reduced flagellin protein levels in the mutant strain while ectopic expression of FliW resulted in enhanced levels. Real‐time RT‐PCR revealed relatively minor changes in flaA and flaB mRNA levels for the recombinant and parent strain consistent with post‐transcriptional regulation. Purified FliW was found to bind to FlaA and FlaB flagellin as well as to the global post‐transcriptional regulator CsrA. Inactivation of CsrA resulted in increased levels of flagellin translation. An in vitro translation assay confirmed the regulatory role of CsrA in flagellin biosynthesis. We propose that competitive reciprocal binding of FliW to flagellins and the RNA binding protein CsrA serves as a feedback mechanism to control the number of cytosolic flagellin copies at the protein level.

Microbiota-Derived Short-Chain Fatty Acids Modulate Expression of Campylobacter jejuni Determinants Required for Commensalism and Virulence

“…These studies also showed that while acetate activates expression of invasion genes, butyrate and propionate function to repress expression of invasion genes, demonstrating that different SCFAs have opposite effects on invasion gene expression (40, 65). Although C. jejuni produces CsrA, which affects expression and translation of some virulence and colonization genes, it lacks homologs of the BarA/UvrY system, CsrB, CsrC, and HilD (66–69). Thus, C. jejuni employs a different system than E. coli and Salmonella species to sense and respond to these metabolites in the chick intestinal tract.…”

Genome-wide fitness analyses of the foodborne pathogen Campylobacter jejuni in in vitro and in vivo models

“…8). This included the filament chaperone gene fliW 13, 62 and the flagellar cap fliD . A FliD subunit vaccine induced a transient but significant reduction (~2 log) of C .…”

Importance of Campylobacter jejuni FliS and FliW in Flagella Biogenesis and Flagellin Secretion

“…The resulting PCR fragment was cloned directly into pSCODON1.2 with the use of NdeI and XhoI restriction enzymes, yielding pSCODON1.2-FliS. The cloning of C. jejuni racR gene to pSCODON1.2 vector has been described elsewhere (Radomska et al, 2016). …”

“…EcoRI and XhoI sites present in the MCS of the vector were used for cloning. The cloning of fliW gene to pGEX4T-2 expression vector has been described elsewhere (Radomska et al, 2016). …”

“…In C. jejuni and B. subtilis CsrA interacts with 5′-UTR of flagellin transcripts and inhibits the translation initiation. Binding of FliW to CsrA resolves this inhibition (Mukherjee et al, 2011; Dugar et al, 2016; Radomska et al, 2016). In B. subtilis the interaction of FliS with the flagellin indirectly antagonizes CsrA activity by enhancing flagellin secretion.…”