Absence of the Polar Organizing Protein PopZ Results in Reduced and Asymmetric Cell Division in Agrobacterium tumefaciens

Abstract

Agrobacterium tumefaciens is a rod-shaped bacterium that grows by polar insertion of new peptidoglycan during cell elongation. As the cell cycle progresses, peptidoglycan synthesis at the pole ceases prior to insertion of new peptidoglycan at midcell to enable cell division. The A. tumefaciens homolog of the Caulobacter crescentus polar organelle development protein PopZ has been identified as a growth pole marker and a candidate polar growth-promoting factor. Here, we characterize the function of PopZ in cell growth and division of A. tumefaciens. Consistent with previous observations, we observe that PopZ localizes specifically to the growth pole in wild-type cells. Despite the striking localization pattern of PopZ, we find the absence of the protein does not impair polar elongation or cause major changes in the peptidoglycan composition. Instead, we observe an atypical cell length distribution, including minicells, elongated cells, and cells with ectopic poles. Most minicells lack DNA, suggesting a defect in chromosome segregation. Furthermore, the canonical cell division proteins FtsZ and FtsA are misplaced, leading to asymmetric sites of cell constriction. Together, these data suggest that PopZ plays an important role in the regulation of chromosome segregation and cell division.IMPORTANCE A. tumefaciens is a bacterial plant pathogen and a natural genetic engineer. However, very little is known about the spatial and temporal regulation of cell wall biogenesis that leads to polar growth in this bacterium. Understanding the molecular basis of A. tumefaciens growth may allow for the development of innovations to prevent disease or to promote growth during biotechnology applications. Finally, since many closely related plant and animal pathogens exhibit polar growth, discoveries in A. tumefaciens may be broadly applicable for devising antimicrobial strategies.KEYWORDS Agrobacterium, PopZ, cell polarity, chromosome segregation, cell division, growth polarity A grobacterium tumefaciens, the causative agent of crown gall disease in flowering plants, has been studied extensively with regard to pathogenesis and its ability to transfer engineered DNA to plant cells using the type IV secretion system encoded by multiple vir genes. More recent observations of A. tumefaciens growth have revealed that this bacterium exhibits polar growth during elongation (1, 2). In A. tumefaciens, as well as in other Rhizobiales, peptidoglycan is inserted at the new poles created by cell division until the cell doubles in length. The peptidoglycan synthesis is then directed to midcell to enable septum formation and cell division. The growth patterning of A.

Agrobacterium tumefaciens divisome proteins regulate the transition from polar growth to cell division

“…In WT A. tumefaciens , deletion of popZ has been shown to cause ectopic poles and cells devoid of DNA, demonstrating a role in coordinating cell division with chromosome segregation (Ehrle et al ., ; Howell et al ., ). We hypothesize that PopZ‐dependent coordination of cell division likely involves FtsZ.…”

“…These observations indicate that FtsZ AT is required for recruitment of PopZ from the growth pole to mid‐cell. Remarkably, both FtsZ AT and FtsA are mislocalized in the absence of PopZ, leading to the establishment of asymmetric constriction sites and a broad range of cell lengths (Howell et al ., ). Together, these data suggest that the presence of both PopZ and FtsZ are important for proper positioning and functioning of the divisome.…”

“…While the mid-cell localization of PopZ is dependent on the presence of FtsZ AT (Fig. 4), FtsZ AT -ring stability and placement are impacted by the absence of PopZ (Howell et al, 2017a). Furthermore, deletion of popZ impairs termination of polar growth and results in cell division defects (Ehrle et al, 2017;Grangeon et al, 2017;Howell et al, 2017a).…”

“…4), FtsZ AT -ring stability and placement are impacted by the absence of PopZ (Howell et al, 2017a). Furthermore, deletion of popZ impairs termination of polar growth and results in cell division defects (Ehrle et al, 2017;Grangeon et al, 2017;Howell et al, 2017a). The apparent co-dependence of FtsZ and PopZ for localization may suggest that these proteins function together during the early stages of cell division, particularly the termination of polar growth and onset of mid-cell PG biosynthesis.…”

“…In WT, PopZ-YFP localizes to the growing pole during elongation and is recruited to mid-cell just prior to cell separation (Fig. 4B, top panel) (Grangeon et al, 2015;Ehrle et al, 2017;Howell et al, 2017a). In some freshly divided cells PopZ-YFP can be visualized at both the growing pole as well as the old pole resulting in an average of 1.12 PopZ-YFP labeled poles per cell (100 cells analyzed SD ± 0.33).…”