Biology subjects

Ferreira, C. N.

Publications and source records attributed to Ferreira, C. N..

4 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↗

bioRxiv2023

Protein coopted from a phage restriction system dictates orthogonal cell division plane selection in Staphylococcus aureus

The spherical bacterium Staphylococcus aureus, a leading cause of nosocomial infections, undergoes binary fission by dividing in two alternating orthogonal planes, but the mechanism by which S. aureus correctly selects the next cell division plane is not known. To identify cell division placement factors, we performed a chemical genetic screen that revealed a gene which we termed pcdA. We show that PcdA is a member of the McrB family of AAA+ NTPases that has undergone structural changes and a concomitant functional shift from a restriction enzyme subunit to an early cell division protein. PcdA directly interacts with the tubulin-like central divisome component FtsZ and localizes to future cell division sites before membrane invagination initiates. This parallels the action of another McrB family protein, CTTNBP2, which stabilizes microtubules in animals. We show that PcdA also interacts with the structural protein DivIVA and propose that the DivIVA/PcdA complex recruits unpolymerized FtsZ to assemble along the proper cell division plane. Deletion of pcdA conferred abnormal, non-orthogonal division plane selection, increased sensitivity to cell wall-targeting antibiotics, and reduced virulence in a murine infection model. Targeting PcdA could therefore highlight a treatment strategy for combatting antibiotic-resistant strains of S. aureus.

microbiology↗

bioRxiv2022

Active site mutations in the bacterial actin homolog FtsA impair direct interactions with the divisome in Escherichia coli

During cell division in Escherichia coli, the highly conserved tubulin homolog FtsZ polymerizes and assembles into a ring-like structure, called the Z-ring, at the site of septation early in the division pathway. For recruitment to the membrane surface, FtsZ polymers directly interact with membrane-associated proteins. In E. coli, membrane recruitment and tethering of FtsZ are predominantly carried out by FtsA. FtsA shares structural homology with actin and, like actin, hydrolyzes ATP. Yeast actin detects nucleotide occupancy through a sensor region adjacent to the nucleotide binding site and adopts distinct conformations in monomeric and filamentous actin. Accordingly, bacterial actin homologs also display considerable conformational flexibility across different nucleotide-bound states and adopt a polymerized conformation. Here, we show that a cluster of amino acid residues in the central region of FtsA and proximal to the nucleotide binding site are critical for FtsA function in vitro and in vivo. Each of these residues are important for ATP hydrolysis, phospholipid (PL) binding, ATP-dependent vesicle remodeling, and recruitment to the divisome in vivo, to varying degrees. Notably, we observed that Ser 84 and Glu 14 are essential for ATP-dependent vesicle remodeling and magnesium-dependent membrane release of FtsA from vesicles in vitro, and these defects likely underlie the loss of function by FtsA(E14R) and FtsA(S84L) in vivo. Finally, we demonstrate that FtsA(A188V), which is associated with temperature-sensitive growth in vivo, is defective for rapid ATP hydrolysis and ATP-dependent remodeling of PL vesicles in vitro. Together, our results show that loss of nucleotide-dependent activities by FtsA, such as ATP hydrolysis, ATP-dependent PL vesicle remodeling, and membrane release, lead to failed Z-ring assembly and division defects in cells.

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↗