Flagellar assembly factor FliW2 de-represses Helicobacter pylori FlaA-mediated motility by allosteric obstruction of global regulator CsrA

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Su, MS, Dickins, B ORCID: https://orcid.org/0000-0002-0866-6232, Kiang, FY, Tsai, W, Chen, Y, Chen, J, Wang, S, Tsai, P and Wu, J, 2025. Flagellar assembly factor FliW2 de-represses Helicobacter pylori FlaA-mediated motility by allosteric obstruction of global regulator CsrA. Helicobacter, 30 (2): e70019. ISSN 1083-4389

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Official URL: https://doi.org/10.1111/hel.70019

Abstract

Background

Helicobacter pylori colonizes the human stomach as a dominant member of the gastric microbiota and constitutively expresses flagellar motility for survival. Carbon storage regulator A (CsrA) is a posttranscriptional global regulator and a critical determinant of H. pylori's motility and pathogenicity. The regulation of H. pylori CsrA is still uncertain although in other species CsrA is reported to be antagonized by small RNAs and proteins. In this study, we attempted to unveil how CsrA is regulated and hypothesized that H. pylori CsrA activity is antagonized by a flagellar assembly factor, FliW2, via protein allosteric obstruction.

Materials and Methods

Multiple sequence comparisons indicated that, along its length and in contrast to fliW1, the fliW2 of H. pylori J99 is conserved. We then generated an isogenic ΔfliW2 strain whose function was characterized using phenotypic and biochemical approaches. We also applied a machine learning approach (AlphaFold2) to predict FliW2-CsrA binding domains and investigated the FliW2-CsrA interaction using pull-down assays and in vivo bacterial two-hybrid systems.

Results

We observed the reduced expression of major flagellin FlaA and impaired flagellar filaments that attenuated the motility of the ΔfliW2 strain. Furthermore, a direct interaction between FliW2 and CsrA was demonstrated, and a novel region of the C-terminal extension of CsrA was suggested to be crucial for CsrA interacting with FliW2. Based on our AlphaFold2 prediction, this C-terminal region of FliW2-CsrA interaction does not overlap with CsrA's N-terminal RNA binding domain, implying that FliW2 allosterically antagonizes CsrA activity and restricts CsrA's binding to flaA mRNAs.

Conclusions

Our data points to novel regulatory roles that the H. pylori flagellar assembly factor FliW2 has in obstructing CsrA activity, and thus FliW2 may indirectly antagonize CsrA's regulation of flaA mRNA processing and translation. Our findings reveal a new regulatory mechanism of flagellar motility in H. pylori.

Item Type:	Journal article
Publication Title:	Helicobacter
Creators:	Su, M.S., Dickins, B., Kiang, F.Y., Tsai, W., Chen, Y., Chen, J., Wang, S., Tsai, P. and Wu, J.
Publisher:	Wiley
Date:	2025
Volume:	30
Number:	2
ISSN:	1083-4389
Identifiers:	Number Type 10.1111/hel.70019 DOI 2419868 Other
Rights:	© 2025 The Author(s). Helicobacter published by John Wiley & Sons Ltd. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.
Divisions:	Schools > School of Science and Technology
Record created by:	Melissa Cornwell
Date Added:	01 Apr 2025 14:49
Last Modified:	01 Apr 2025 14:49
URI:	https://irep.ntu.ac.uk/id/eprint/53343