Flagellar brake protein YcgR interacts with motor proteins MotA and FliG to regulate the flagellar rotation speed and direction

Qun Han; Shao-Feng Wang; Xin-Xin Qian; Lu Guo; Yi-Feng Shi; Rui He; Jun-Hua Yuan; Yan-Jie Hou; De-Feng Li

doi:10.3389/fmicb.2023.1159974

Flagellar brake protein YcgR interacts with motor proteins MotA and FliG to regulate the flagellar rotation speed and direction

Front Microbiol. 2023 Apr 14:14:1159974. doi: 10.3389/fmicb.2023.1159974. eCollection 2023.

Authors

Qun Han^{1

2}, Shao-Feng Wang^{1

2}, Xin-Xin Qian¹, Lu Guo¹, Yi-Feng Shi¹, Rui He³, Jun-Hua Yuan³, Yan-Jie Hou⁴, De-Feng Li^{1

2}

Affiliations

¹ State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
² College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China.
³ Hefei National Laboratory for Physical Sciences at the Microscale and Department of Physics, University of Science and Technology of China, Hefei, Anhui, China.
⁴ National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Abstract

In E. coli and related species, flagellar brake protein YcgR responds to the elevated intracellular c-di-GMP, decreases the flagellar rotation speed, causes a CCW rotation bias, and regulates bacterial swimming. Boehm et al. suggested that c-di-GMP-activated YcgR directly interacted with the motor protein MotA to curb flagellar motor output. Paul et al. proposed that YcgR disrupted the organization of the FliG C-terminal domain to bias the flagellar rotation. The target proteins are controversial, and the role of motor proteins remains unclear in flagellar rotation speed and direction regulation by YcgR. Here we assayed the motor proteins' affinity via a modified FRET biosensor and accessed the role of those key residue via bead assays. We found that YcgR could interact with both MotA and FliG, and the affinities could be enhanced upon c-di-GMP binding. Furthermore, residue D54 of YcgR-N was needed for FliG binding. The mutation of the FliG binding residue D54 or the MotA binding ones, F117 and E232, restored flagellar rotation speed in wild-type cells and cells lacking chemotaxis response regulator CheY that switched the flagellar rotation direction and decreased the CCW ratio in wild-type cells. We propose that c-di-GMP-activated YcgR regulated the flagellar rotation speed and direction via its interaction with motor proteins MotA and FliG. Our work suggest the role of YcgR-motor proteins interaction in bacterial swimming regulation.

Keywords: Escherichia coli; YcgR; c-di-GMP; flagellar brake protein; flagellar motility.