Biomedical subjects
Publications and source records attributed to William Margolin.
Formation of the polar Z ring is a crucial step in the establishment of cellular asymmetry during sporulation of Bacillus subtilis. New results suggest that the transition from medial to polar Z rings involves a dynamic FtsZ spiral structure that may transfer FtsZ from medial to polar sites.
The MinCDE proteins help to select cell division sites in normal cylindrical Escherichia coli by oscillating along the long axis, preventing unwanted polar divisions. To determine how the Min system might function in cells with multiple potential division planes, we investigated its role in a round-cell rodA mutant. Round cells lacking MinCDE were viable, but growth, morphology and positioning of cell division sites were abnormal relative to Min+ cells. In round cells with a long axis, such as those undergoing cell division, green fluorescent protein (GFP) fusions to MinD almost always oscillated parallel to the long axis. However, perfect spheres or irregularly shaped cells exhibited MinD movement to and from multiple sites on the cell surface. A MinE-GFP fusion exhibited similar behavior. These results indicate that the Min proteins can potentially localize anywhere in the cell but tend to move a certain maximum distance from their previous assembly site, thus favoring movement along the cell's long axis. A new model for the spatial control of division planes by the Min system in round cells is proposed.
Min proteins are involved in the correct placement of division septa in many bacterial species. In Escherichia coli (Ec) cells, these proteins oscillate from pole to pole, ostensibly to prevent unwanted polar septation. Here, we show that Min proteins from the coccus Neisseria gonorrhoeae (Ng) also oscillate in E. coli. Green fluorescent protein (GFP) fusions to gonococcal MinD and MinE localized dynamically in different E. coli backgrounds. GFP-MinDNg moved from pole to pole in rod-shaped E. coli cells with a 70 +/- 25 s localization cycle when MinENg was expressed in cis. The oscillation time of GFP-MinDNg was reduced when wild-type MinENg was replaced with MinENg carrying a R30D mutation, but lengthened by 15 s when activated by MinEEc. Several mutations in the N-terminal domain of MinDNg, including K16Q and 4- and 19-amino acid truncations, prevented oscillation; these MinDNg mutants showed decreased or lost interaction with themselves and MinENg. Like MinEEc-GFP, MinENg-GFP formed MinE rings and oscillated in E. coli cells when MinDEc was expressed in cis. Finally, in round E. coli cells, GFP-MinDNg appeared to move in a plane parallel to completed septa. This pattern of movement is predicted to be similar in gonococcal cells, which also divide in alternating perpendicular planes.
DivIVA of Bacillus subtilis and FtsZ of Escherichia coli were used to target heterologous protein complexes to cell division sites of E. coli and Agrobacterium tumefaciens. DivIVA and FtsZ that were fused to the dimerizing leucine zipper (LZ) domain of the yeast transcription activator GCN4 directed the green fluorescent protein (GFP) that was fused to an LZ domain to E. coli division sites, resulting in fluorescence patterns identical to those observed with DivIVA::GFP and FtsZ::GFP. These cell division proteins also targeted the VirE1 chaperone and VirE2 secretion substrate complex to division sites of E. coli and A. tumefaciens. Coproduction of the native VirE1 or VirE2 proteins inhibited the dihybrid interaction in both species, as judged by loss of GFP targeting to division sites. The VirE1 chaperone bound independently to N- and C-terminal regions of VirE2, with a requirement for residues 84 to 147 and 331 to 405 for these interactions, as shown by dihybrid studies with VirE1::GFP and DivIVA fused to N- and C-terminal VirE2 fragments. DivIVA also targeted homo- and heterotypic complexes of VirB8 and VirB10, two bitopic inner membrane subunits of the A. tumefaciens T-DNA transfer system, in E. coli and homotypic complexes of VirB10 in A. tumefaciens. VirB10 self-association in bacteria was mediated by the C-terminal periplasmic domain, as shown by dihybrid studies with fusions to VirB10 truncation derivatives. Together, our findings establish a proof-of-concept for the use of cell-location-specific proteins for studies of interactions among cytosolic and membrane proteins in diverse bacterial species.