A genetic screen unveils key molecular steps in the polymerization cycle of the bacterial actin-like MreB
Abstract
MreB, a bacterial actin homolog and polymerizing ATPase, is central to cell-shape maintenance and cell-wall integrity. Its functions rely on its ability to assemble into dynamic, membrane-associated polymers. However, how nucleotide binding and hydrolysis, MreB-MreB contacts, and membrane association are coordinated to enable polymer assembly and disassembly remains unclear. Here, we combined genetics and live-cell microscopy with biochemical approaches to dissect these processes. Using a highly sensitive reporter of MreB activity, we identified, through a genetic screen, residues critical for MreB function in Bacillus subtilis . Subsequent extensive characterization of corresponding stable variants of the homologous Geobacillus stearothermophilus MreB revealed that ATP binding, but not ATP hydrolysis, is required for polymerization. Productive longitudinal intraprotofilament contacts are required for efficient ATP hydrolysis and enhance membrane association. Perturbations predicted to weaken lateral interprotofilament contacts altered membrane association and modulated ATPase activity. Together, these effects provide experimental evidence consistent with long-range functional coupling among the longitudinal and lateral protofilament interfaces, the distant nucleotide-binding site, and membrane association dynamics. Moreover, impaired ATP hydrolysis delays disassembly of lipid-associated polymers, indicating that hydrolysis promotes polymer turnover. These results establish key mechanistic steps coordinating ATP-driven MreB polymerization and turnover and provide a basis for a complete MreB assembly–disassembly cycle and for further elucidating how MreB dynamics contribute to cell-wall organization.
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BibTeX
@misc{eustaquiocampillo2026genetic,
title = {{A genetic screen unveils key molecular steps in the polymerization cycle of the bacterial actin-like MreB}},
author = {Alba de San Eustaquio-Campillo and Charlene Cornilleau and Sana Afensiss and Catalina Marchioni and Hind Oulkfif and Lilia Huynh and Davy Martin and Lars Renner and Rut Carballido-López and Louis Renault and Arnaud Chastanet},
howpublished = {HAL},
year = {2026},
month = jul,
doi = {10.64898/2026.07.23.740262},
url = {https://hal.science/hal-05771616v1},
note = {CC BY-NC},
}
RIS
TY - UNPB
TI - A genetic screen unveils key molecular steps in the polymerization cycle of the bacterial actin-like MreB
AU - Alba de San Eustaquio-Campillo
AU - Charlene Cornilleau
AU - Sana Afensiss
AU - Catalina Marchioni
AU - Hind Oulkfif
AU - Lilia Huynh
AU - Davy Martin
AU - Lars Renner
AU - Rut Carballido-López
AU - Louis Renault
AU - Arnaud Chastanet
PY - 2026
DA - 2026/07/23
PB - HAL
DO - 10.64898/2026.07.23.740262
UR - https://hal.science/hal-05771616v1
AB - MreB, a bacterial actin homolog and polymerizing ATPase, is central to cell-shape maintenance and cell-wall integrity. Its functions rely on its ability to assemble into dynamic, membrane-associated polymers. However, how nucleotide binding and hydrolysis, MreB-MreB contacts, and membrane association are coordinated to enable polymer assembly and disassembly remains unclear. Here, we combined genetics and live-cell microscopy with biochemical approaches to dissect these processes. Using a highly sensitive reporter of MreB activity, we identified, through a genetic screen, residues critical for MreB function in Bacillus subtilis . Subsequent extensive characterization of corresponding stable variants of the homologous Geobacillus stearothermophilus MreB revealed that ATP binding, but not ATP hydrolysis, is required for polymerization. Productive longitudinal intraprotofilament contacts are required for efficient ATP hydrolysis and enhance membrane association. Perturbations predicted to weaken lateral interprotofilament contacts altered membrane association and modulated ATPase activity. Together, these effects provide experimental evidence consistent with long-range functional coupling among the longitudinal and lateral protofilament interfaces, the distant nucleotide-binding site, and membrane association dynamics. Moreover, impaired ATP hydrolysis delays disassembly of lipid-associated polymers, indicating that hydrolysis promotes polymer turnover. These results establish key mechanistic steps coordinating ATP-driven MreB polymerization and turnover and provide a basis for a complete MreB assembly–disassembly cycle and for further elucidating how MreB dynamics contribute to cell-wall organization.
ER -