EzrA promotes Z-ring formation through interaction of its QNR motif with FtsA.

Bacterial cell division requires precise placement and formation of the division machinery to ensure the accurate generation of identical daughter cells. This process is canonically initiated by the highly conserved FtsZ but also needs the involvement of a variety of FtsZ-binding proteins to orchestrate the spatial and temporal positioning and assembly of the Z-ring. However, the underlying molecular mechanisms remain poorly understood. In this study, we characterized the roles of an important FtsZ binding protein EzrA in the cell division of Streptococcus suis, an emerging zoonotic bacterial pathogen. Our results revealed that EzrA shares high subcellular dynamics with FtsZ during the entire cell division cycle and functions primarily as a positive regulator for Z-ring formation. Co-immunoprecipitation and bacterial two-hybrid data suggest that EzrA interacts with FtsZ and several early division proteins. Importantly, the conserved QNR motif in EzrA directly contributes to its interaction with FtsA. Disrupting this motif results in the mislocalization of EzrA itself at the division site rather than the localization of FtsA, which remains concentrated localization at the division site. Moreover, the interaction of EzrA through the QNR motif with FtsA is conserved among the Firmicutes. Taken together, these findings demonstrate EzrA as a regulator of Z-ring positioning to the division site through the interaction of its conserved QNR motif with FtsA.IMPORTANCEBacteria replicate through binary fission in which the FtsZ-ring positioning and assembly is a crucial process requiring precise spatial and temporal regulation. However, the mechanism of this process remains largely unknown, especially in ovoid-shaped bacteria, such as Streptococci, in which many members are important human and animal pathogens. In this study, we characterize the critical role of the cell division regulator EzrA in the formation of the Z-ring. Our data reveal a model in which EzrA interacts through its QNR motif with FtsA to be properly localized to the septum so as to facilitate the positioning and formation of the Z-ring of Streptococcus suis. This regulatory mechanism could be conserved in Firmicutes. This research provides insights into the regulation mechanism of the Z-ring formation and will contribute to the understanding of the cell division process in Streptococci.