Biology subjects

Conti, J.

Publications and source records attributed to Conti, J..

3 recordsLinked to original sources

bioRxiv2026

MinC influences the Min oscillation period in Escherichia coli by priming MinD on the membrane

Spatial organization during bacterial fission is critical to ensure that division results in identical, viable progeny cells. In Escherichia coli, one important system that accomplishes this is the Min system, comprising MinC, MinD, and MinE proteins. The Min system regulates assembly of the FtsZ-ring to ensure proper placement of the division machinery at midcell. We explored the dynamics of the Min system and demonstrate that Min oscillation and release of MinD from the membrane is modulated by MinC. Deletion of the minC gene from a strain expressing minD fused to the gene encoding green fluorescent protein (Gfp) resulted in faster pole-to-pole oscillation of Gfp-MinD in vivo, leading to a shorter period. This was also observed in a strain expressing the MinC(R133A), a variant mutated at the MinC-MinD interface. In phospholipid binding and release assays in vitro, we observed that the concentration of MinE required to release MinD was lower with MinC, suggesting that MinC activates MinD for MinE-dependent release. Oscillation is produced by a diffusion-capture-release mechanism occurring at the membrane with proteins proceeding toward a pole. To determine if an increase in the diffusive proportion of MinD leads to faster oscillation, we constructed MinD membrane targeting sequence (MTS) variants to reduce the membrane associated pool of MinD. MTS variants induced a range of oscillation defects, with MinD(KR265AA) oscillating faster in vivo. Together our results show that MinC is an effector that modifies Min oscillation by priming MinD for MinE-dependent membrane release and fine-tunes the Min oscillation rate in dividing cells.

microbiology↗

bioRxiv2021

Coregulated assembly of actin-like FtsA polymers with FtsZ during Z-ring formation and division in Escherichia coli

In Escherichia coli, the actin homolog FtsA localizes the cell division machinery, beginning with the Z-ring, to the cytoplasmic membrane through direct interaction with FtsZ. FtsZ polymers are first to assemble at the Z-ring at midcell, where they direct constriction and septation. While FtsZ polymerization is critical for establishing a functional Z-ring that leads to constriction, the assembly state of FtsA and the role of FtsA ATP utilization during division in E. coli remain unclear. Here, we show that ATP hydrolysis, FtsZ interaction, and phospholipid vesicle remodeling by FtsA are impaired by a substitution mutation at the predicted active site for hydrolysis. This mutation, Glu 14 to Arg, also impairs Z-ring assembly and division in vivo. To further investigate the role of phospholipid engagement and ATP utilization in regulating FtsA function, we characterized a truncated E. coli FtsA variant, FtsA({Delta}MTS), which lacks the region at the C-terminus important for engaging the membrane and is defective for ATP hydrolysis. We show that E. coli FtsA({Delta}MTS) forms ATP-dependent actin-like filaments and assembly is antagonized by FtsZ. Polymerization of FtsZ with GTP, or a non-hydrolyzable analog, blocks inhibition of ATP-dependent FtsA assembly, and instead favors coassembly of stable FtsA/FtsZ polymers. In the cell, FtsA/FtsZ coassembly is favored at midcell, where FtsZ polymerizes, and inhibited at regions where FtsZ polymers are destabilized by regulators, such as MinC at the poles or SlmA at the nucleoid. We show that MinC prevents recruitment of FtsZ, via FtsA, to phospholipids, suggesting that local interactions of MinC with FtsZ block membrane tethering and uncouple the Z-ring from its major membrane contact. During Z-ring formation, the coassembly of FtsZ polymers with FtsA is coordinated and is a critical early step in division. This step also serves as a checkpoint by responding to the suite of FtsZ assembly regulators in the cell that modulate Z-ring position and dynamics prior to initiating cell wall synthesis.

microbiology↗

bioRxiv2020

Disassembly and degradation of MinD oscillator complexes by Escherichia coli ClpXP

MinD is a cell division ATPase in Escherichia coli that oscillates from pole to pole and regulates the spatial position of the cell division machinery. Together with MinC and MinE, the Min system restricts assembly of the FtsZ-ring to midcell, oscillating between the opposite ends of the cell and preventing FtsZ-ring misassembly at the poles. Here, we show that the ATP-dependent bacterial proteasome complex ClpXP degrades MinD in reconstituted degradation reactions in vitro, through direct recognition of the MinD N-terminal region, and in vivo. MinD degradation is enhanced during stationary phase, suggesting that ClpXP regulates levels of MinD in cells that are not actively dividing. MinC and MinD are known to co-assemble into linear polymers, therefore we monitored copolymers assembled in vitro after incubation with ClpXP and observed that ClpXP promotes rapid MinCD copolymer disassembly as a result of direct MinD degradation by ClpXP. The N-terminus of MinD, including residue Arg 3, which is near the ATP-binding site, is critical for degradation by ClpXP. Together, these results demonstrate that ClpXP degradation modifies conformational assemblies of MinD in vitro and depresses Min function in vivo during periods of reduced proliferation.

microbiology↗