ClpXP and ClpAP control the Escherichia coli division protein ZapC by proteolysis

Abstract

The bacterial FtsZ-ring is an essential cytokinetic structure under tight spatiotemporal regulation. In Escherichia coli, FtsZ polymerization and assembly into the Z-ring is controlled on multiple levels through interactions with positive and negative regulators. Among these regulatory factors are ZapC, a Z-ring stabilizer, and the conserved protease ClpXP, which has been shown to degrade FtsZ protofilaments in preference to FtsZ monomers. Here we report that ZapC and ClpX interact in a protein-protein interaction assay, and that ZapC is degraded in a ClpXP-dependent manner in vivo. The SspB adaptor protein is not required for targeting ZapC to the ClpXP proteolytic machinery. A mutation disrupting the zapC ssrAlike sequence (zapC DD ) stabilizes ZapC consistent with a reduction in ClpXP-mediated ZapC degradation. ZapC DD retains the ability to interact with FtsZ and to promote bundling in vitro indicating that WT ZapC contains discrete FtsZ and ClpX recognition motifs. Additionally, ClpAP complexes are sufficient for degradation of ZapC in the absence of ClpX in vivo. Further, chromosomal expression of zapC DD suppresses filamentation of the temperature-sensitive ftsZ84 mutant, confirming the role of ZapC as a Z-ring stabilizer. Lastly, changes in ClpXP and ZapC levels lead to cell division effects, likely through their roles in modulating FtsZ assembly dynamics. Taken together, our results indicate that the Zring stabilizer ZapC is a substrate of both ClpXP and ClpAP in vivo. Our data also point to a more complex regulatory circuit that integrates FtsZ, ClpXP and ZapC in achieving Z-ring stability in E. coli and related species.

Proteolysis-Dependent Remodeling of the Tubulin Homolog FtsZ at the Division Septum in Escherichia coli

“…By replacing the clpP gene with one that encodes a ClpP proteolytic mutant, ClpP(S97A), we confirmed that ClpXP degradation, not just ATP-dependent remodeling by ClpX, is critical for regulating division. Through direct degradation of FtsZ and the additional degradation of the Z-ring stabilizer ZapC, ClpXP likely has potent Z-ring destabilizing activity [59]. …”

Sequestration from Protease Adaptor Confers Differential Stability to Protease Substrate

“…The AAA+ family protease machines contribute to protein quality control by promoting proteolysis of a variety of proteins in all domains of life (Bonnet et al, 2013; Buczek et al, 2016; Mogk et al, 2007; Song et al, 2015; Williams et al, 2014). In bacteria, the AAA+ protease ClpAP is composed of the serine protease ClpP and the regulatory ATPase ClpA (Thompson and Maurizi, 1994).…”

Survival of Anaerobic Fe 2+ Stress Requires the ClpXP Protease

“…ClpXP is one of five AAAϩ proteases encoded in the S. oneidensis genome, the other four being ClpAP, Lon, HslVU, and FtsH (26)(27)(28)(29)(30). ClpXP has several established roles in bacteria, including regulation of entry into stationary phase via degradation of the stress response regulator S , degradation of cell division proteins, promoting release of the envelope stress response regulator E , and turning over ribosomes by degrading proteins stalled during translation (25,(31)(32)(33)(34)(35)(36). We demonstrate that the role of ClpXP in mediating resistance to anaerobic Fe 2ϩ stress is independent of these previously established roles.…”