Determinants of FtsZ C-terminal linker-dependent regulation of cell wall metabolism in Caulobacter crescentus

Abstract

Bacterial cell division requires assembly of a multi-protein machinery or \"divisome\" that remodels the cell envelope to cause constriction. The cytoskeletal protein FtsZ forms a ring-like scaffold for the divisome at the incipient division site. FtsZ has three major regions - a conserved, polymerizing GTPase domain; a C-terminal conserved (CTC) peptide required for binding membrane-anchoring proteins; and a C-terminal linker (CTL) of poor length and sequence conservation. We previously demonstrated that, in Caulobacter crescentus, the CTL regulates FtsZ polymerization in vitro and cell wall metabolism in vivo. To understand the mechanism of CTL-dependent regulation of cell wall metabolism, here we investigated the impact of the CTL on Z-ring structure in cells and employed genetics to identify molecular determinants of the dominant lethal effects of {Delta}CTL. Deleting the CTL specifically resulted in formation of dense, asymmetric, non-ring FtsZ assemblies in vivo. Moreover, we observed that production of an FtsZ variant with the GTPase domain of Escherichia coli FtsZ fused to the CTC of C. crescentus FtsZ phenocopied the effects of C. crescentus {Delta}CTL, suggesting the CTC mediates signaling to cell wall metabolism. Finally, whereas overproduction of ZapA, FzlC, or FtsEX had slight protective effects against {Delta}CTL, depletion of FtsA partially suppressed the effects of {Delta}CTL. From these results, we propose that the cell wall misregulation downstream of {Delta}CTL results from its aberrant assembly properties and is propagated through the interaction between the CTC of FtsZ and FtsA. Our study provides mechanistic insights into CTL-dependent regulation of cell wall enzymes downstream of FtsZ.\n\nImportanceBacterial cell division is essential and requires the recruitment and regulation of a complex network of proteins needed to initiate and guide constriction and cytokinesis. FtsZ serves as a master regulator for this process, and its function is highly dependent on both its self-assembly into a canonical \"Z-ring\" and interaction with protein binding partners, which results in the activation of enzymes that remodel the cell wall to drive constriction. Using mutants of FtsZ and its binding partners, we have established the role of its C-terminal linker domain in regulating Z-ring organization, as well as the requirement for its C-terminal conserved peptide and interaction with the membrane-anchoring protein FtsA for regulating cell wall remodeling for constriction.