On the role of nucleotides and lipids in the polymerization of the actin homolog MreB from a Gram-positive bacterium
- Micalis Institute, France
- Leibniz Institute of Polymer Research, Germany
- CNRS Université Paris-Saclay, France
- Institut Curie, France
Abstract
In vivo, bacterial actin MreB assembles into dynamic membrane-associated filamentous structures that exhibit circumferential motion around the cell. Current knowledge of MreB biochemical and polymerization properties in vitro remains limited and is mostly based on MreB proteins from Gram-negative species. In this study, we report the first observation of organized protofilaments by electron microscopy and the first 3D-structure of MreB from a Gram-positive bacterium. We show that Geobacillus stearothermophilus MreB forms straight pairs of protofilaments on lipid surfaces in the presence of ATP or GTP, but not in the presence of ADP, GDP or non-hydrolysable ATP analogs. We demonstrate that membrane anchoring is mediated by two spatially close short hydrophobic sequences while electrostatic interactions also contribute to lipid binding, and show that the population of membrane-bound protofilament doublets is in steady-state. In solution, protofilament doublets were not detected in any condition tested. Instead, MreB formed large sheets regardless of the bound nucleotide, albeit at a higher critical concentration. Altogether, our results indicate that both lipids and ATP are facilitators of MreB polymerization, and are consistent with a dual effect of ATP hydrolysis, in promoting both membrane binding and filaments assembly/disassembly.
Data availability
Protein structures data have been deposited in PDB under the accession codes 7ZPT and 8AZG.
Article and author information
Author details
Ines Li de la Sierra-Gallay
Rut Carballido-Lopez
Funding
European Research Council (ERC-SG,311231)
- Rut Carballido-Lopez
European Research Council (ERC-CG,772178)
- Rut Carballido-Lopez
Agence Nationale de la Recherche (ANR-11-LABX0038)
- Aurélie Bertin
Agence Nationale de la Recherche (ANR-10-IDEX-0001-02)
- Aurélie Bertin
VolkswagenStiftung
- Lars D Renner
Agence Nationale de la Recherche (ANR-10-INSB-05-05)
- Sylvie Nessler
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2023, Mao et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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On the role of nucleotides and lipids in the polymerization of the actin homolog MreB from a Gram-positive bacterium
eLife 12:e84505.
https://doi.org/10.7554/eLife.84505
Further reading
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- Biochemistry and Chemical Biology
In healthy cells, cyclin D1 is expressed during the G1 phase of the cell cycle, where it activates CDK4 and CDK6. Its dysregulation is a well-established oncogenic driver in numerous human cancers. The cancer-related function of cyclin D1 has been primarily studied by focusing on the phosphorylation of the retinoblastoma (RB) gene product. Here, using an integrative approach combining bioinformatic analyses and biochemical experiments, we show that GTSE1 (G-Two and S phases expressed protein 1), a protein positively regulating cell cycle progression, is a previously unrecognized substrate of cyclin D1–CDK4/6 in tumor cells overexpressing cyclin D1 during G1 and subsequent phases. The phosphorylation of GTSE1 mediated by cyclin D1–CDK4/6 inhibits GTSE1 degradation, leading to high levels of GTSE1 across all cell cycle phases. Functionally, the phosphorylation of GTSE1 promotes cellular proliferation and is associated with poor prognosis within a pan-cancer cohort. Our findings provide insights into cyclin D1’s role in cell cycle control and oncogenesis beyond RB phosphorylation.
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- Biochemistry and Chemical Biology
- Microbiology and Infectious Disease
Teichoic acids (TA) are linear phospho-saccharidic polymers and important constituents of the cell envelope of Gram-positive bacteria, either bound to the peptidoglycan as wall teichoic acids (WTA) or to the membrane as lipoteichoic acids (LTA). The composition of TA varies greatly but the presence of both WTA and LTA is highly conserved, hinting at an underlying fundamental function that is distinct from their specific roles in diverse organisms. We report the observation of a periplasmic space in Streptococcus pneumoniae by cryo-electron microscopy of vitreous sections. The thickness and appearance of this region change upon deletion of genes involved in the attachment of TA, supporting their role in the maintenance of a periplasmic space in Gram-positive bacteria as a possible universal function. Consequences of these mutations were further examined by super-resolved microscopy, following metabolic labeling and fluorophore coupling by click chemistry. This novel labeling method also enabled in-gel analysis of cell fractions. With this approach, we were able to titrate the actual amount of TA per cell and to determine the ratio of WTA to LTA. In addition, we followed the change of TA length during growth phases, and discovered that a mutant devoid of LTA accumulates the membrane-bound polymerized TA precursor.
