The interplay between the polar growth determinant DivIVA, the segregation protein ParA, and their novel interaction partner PapM controls the Mycobacterium smegmatis cell cycle by modulation of DivIVA subcellular distribution

Abstract

ABSTRACT Bacterial chromosome segregation is facilitated by the ParABS system. The ParB protein binds centromere-like parS sequences and forms nucleoprotein complexes. These nucleoprotein complexes are segregated by the dynamic ATPase ParA. In mycobacteria, ParA also interacts with the polar growth determinant DivIVA (Wag31). This interaction was earlier shown not only to facilitate the segregation of ParB complexes but also to affect cell extension. Here, we identify an additional partner of ParA in Mycobacterium smegmatis, named PapM. Using E. coli-based analysis, we show that PapM likewise interacts with DivIVA and that the tripartite interaction of ParA-PapM-DivIVA is phosphorylation dependent: ParA binding to DivIVA is diminished, while PapM binding is promoted upon phosphorylation of DivIVA. The presence of PapM enhances the dissociation of ParA from the DivIVA complex upon its phosphorylation. Studies of M. smegmatis mutant strains reveal that altered PapM levels influence chromosome segregation and cell length. The elimination of PapM affects ParA dynamics. Furthermore, ParA and, to a lesser extent, PapM modulate the subcellular distribution of DivIVA. Altogether, our studies show that the tripartite interplay between ParA-DivIVA and PapM controls the switch between cell division and cell elongation and, in this way, affects the mycobacterial cell cycle. IMPORTANCE The genus of Mycobacterium includes important clinical pathogens (M. tuberculosis). Bacteria of this genus share the unusual features of their cell cycle such as asymmetric polar cell elongation and long generation time. Markedly, control of the mycobacterial cell cycle still remains not fully understood. The main cell growth determinant in mycobacteria is the essential protein DivIVA, which is also involved in cell division. DivIVA activity is controlled by phosphorylation, but the mechanism and significance of this process are unknown. Here, we show how the previously established protein interaction partner of DivIVA in mycobacteria, the segregation protein ParA, affects the DivIVA subcellular distribution. We also demonstrate the role of a newly identified M. smegmatis DivIVA and ParA interaction partner, a protein named PapM, and we establish how their interactions are modulated by phosphorylation. Demonstrating that the tripartite interplay affects the mycobacterial cell cycle contributes to the general understanding of mycobacterial growth regulation.

Keywords

• chromosome segregation
• mycobacteria
• polar growth
• ParA