Biology subjects

Kureisaite-Ciziene, D.

Publications and source records attributed to Kureisaite-Ciziene, D..

5 recordsLinked to original sources

bioRxiv2023

PRC domain-containing proteins modulate FtsZ-based archaeal cell division

Dividing cells into two daughter cells is a complicated process that in bacteria and eukaryotes requires many proteins to work together. For archaea that divide via an FtsZ-based mechanism, only three proteins of the cell division machinery could so far be identified. These are two tubulin homologs, FtsZ1, FtsZ2 and the membrane anchor of FtsZ2, SepF. Here, we investigate additional archaeal cell division proteins that were identified by immunoprecipitation of SepF. These proteins comprise a single PRC-barrel domain and strictly co-occur with FtsZ. Two out of three PRC-barrel domain containing proteins found in Haloferax volcanii, CdpB1 and CdpB2 localize to the site of cell division in a SepF-dependent manner. Moreover, depletions and deletions cause severe cell division defects, generating drastically enlarged cells. Fluorescence microscopy of tagged FtsZ1, FtsZ2 and SepF in CdpB1/2 deletion strains revealed that the divisome is unusually disordered and not organized into a distinct ring-like structure at the cell centre. Biochemical analysis of CdpB homologs from different archaeal species showed that SepF interacts directly with CdpB1, which in turn binds to CdpB2, forming a tripartite complex. A crystal structure of CdpB1 and B2 recapitulated these interactions and suggested how these proteins might form filaments, possibly aligning SepF and therefore the FtsZ2 ring during cell division. In summary, we demonstrate that PRC domain proteins play essential roles in FtsZ based cell division in archaea.

microbiology↗

bioRxiv2021

The bacterial actin-like cell division protein FtsA forms curved antiparallel double filaments upon binding of FtsN

Cell growth and division of walled bacteria depend on the synthesis and remodelling of peptidoglycan (PG). These activities are carried out by two multiprotein complexes, the elongasome and the divisome during cell elongation and division, respectively. Filaments of tubulin-like FtsZ form the cytoplasmic scaffold for divisome assembly, the Z-ring. In E. coli, the actin homologue FtsA anchors the Z-ring to the membrane and recruits downstream divisome components, including bitopic FtsN. FtsN is recruited late and activates the periplasmic PG synthase FtsWI. To start unravelling the activation mechanism involving FtsA and FtsN, we showed that E. coli FtsA forms antiparallel double filaments on lipid monolayers when also binding FtsNs cytoplasmic tail, and that Vibrio maritimus FtsA crystallised as an equivalent double filament. We structurally located the FtsA-FtsN interaction site in FtsAs IA-IC interdomain cleft and confirmed FtsA double filament formation in vivo using site-specific cysteine cross-linking. FtsA-FtsN double filaments reconstituted on and in liposomes preferred negative Gaussian curvature, as was previously shown for the elongasomes actin, MreB. MreB filaments serve as curvature-sensing "rudders", orienting insertion of PG around the cells circumference. We propose that curved antiparallel FtsA double filaments function similarly in the divisome: FtsA filaments, together with dynamic FtsZ filaments orient and concentrate cell-constricting septal PG synthesis in the division plane.

microbiology↗

bioRxiv2019

Bactofilins form non-polar filaments that bind to membranes directly

Bactofilins are small beta-helical proteins that form cytoskeletal filaments in a range of bacteria. Bactofilins have diverse functions: filaments in Caulobacter crescentus are involved in cell stalk formation whereas Myxococcus xanthus filaments aid chromosome segregation and motility. The precise molecular architecture of bactofilin filaments has remained unclear. Here we revealed by sequence analyses and electron microscopy that in addition to wide distribution across bacteria and archaea, bactofilins are also present in a few eukaryotic cells such as oomycetes. The sole bactofilin from Thermus thermophilus was demonstrated to form constitutive filaments and cryo-EM analysis revealed that protofilaments formed through end-to-end association of the beta-helical domains. Using a nanobody against Thermus bactofilin we determined the near-atomic filament structure, showing that the filaments are non-polar, with subunits arranged head-to-head and tail-to-tail. A polymerisation-impaired mutant F105R, that disrupts one of the two protofilament interfaces, enabled crystallisation. The crystal structure also revealed non-polar protofilaments, and the dominance of the beta-stacking interface that formed despite the inhibiting mutation. To confirm the generality of the lack of polarity, we performed co-evolutionary analysis of a large set of sequences. Finally, using Thermus bactofilin, we determined that the N-terminal disordered tail of the protein is responsible for direct binding to lipid membranes both on liposomes and by electron cryotomography in E. coli cells. The tail is conserved, suggesting that membrane binding is likely a general feature of these very common but only recently discovered filaments of the prokaryotic cytoskeleton.

biochemistry↗

bioRxiv2018

Structural analysis of the interaction between the bacterial cell division proteins FtsQ and FtsB

Most bacteria and archaea use similar proteins within their cell division machinery, which uses the tubulin homologue FtsZ as its central organiser. In Gram-negative Escherichia coli bacteria, FtsZ recruits cytosolic, transmembrane, periplasmic and outer membrane proteins, assembling the divisome that facilitates bacterial cell division. One such divisome component, FtsQ, a bitopic membrane protein with a globular domain in the periplasm, has been shown to interact with many other divisome proteins. Despite its otherwise unknown function, it has been shown to be a major divisome interaction hub. Here, we investigated the interactions of FtsQ with FtsB and FtsL, two small bitopic membrane proteins that act immediately downstream of FtsQ. In biochemical assays we show that the periplasmic domains of E. coli FtsB and FtsL interact with FtsQ, but not with each other. Our crystal structure of FtsB bound to the {beta} domain of FtsQ shows that only residues 64-87 of FtsB interact with FtsQ. A synthetic peptide comprising those 24 FtsB residues recapitulates the FtsQ:FtsB interactions. Protein deletions and structure-guided mutant analyses validate the structure. Furthermore, the same structure-guided mutants show cell division defects in vivo that are consistent with our structure of the FtsQ:FtsB complex that shows their interactions as they occur during cell division. Our work provides intricate details of the interactions within the divisome and also provides a tantalising view of a highly conserved protein interaction in the periplasm of bacteria that is an excellent target for cell division inhibitor searches.\n\nImportanceCells in most bacteria and archaea divide through a cell division process that is characterised through its filamentous organiser, FtsZ protein. FtsZ forms a ring structure at the division site and starts the recruitment of 10-20 downstream proteins that together form an elusive multi-protein complex termed divisome. The divisome is thought to facilitate many of the steps required to make two cells out of one. FtsQ and FtsB are part of the divisome, with FtsQ being a central hub, interacting with most of the other divisome components. Here we show for the first time how FtsQ interacts with its downstream partner FtsB and show that mutations that disturb the interface between the two proteins effectively inhibit cell division.

microbiology↗

bioRxiv2016

A polymerisation-associated conformational switch in FtsZ that enables treadmilling

Bacterial cell division in many organisms involves a constricting cytokinetic ring that is orchestrated by the tubulin-like protein FtsZ. FtsZ forms dynamic filaments close to the membrane at the site of division that have recently been shown to treadmill around the division ring, guiding septal wall synthesis.\n\nHere, using X-ray crystallography of Staphylococcus aureus SaFtsZ we reveal how an FtsZ can adopt two functionally distinct conformations, open and closed. The open form is found in SaFtsZ filaments formed in crystals and also in soluble filaments of E. coli FtsZ as deduced by cryoEM. The closed form is found within several crystal forms of two non-polymerising SaFtsZ mutants and corresponds to many previous FtsZ structures from other organisms.\n\nWe argue that FtsZ undergoes a polymerisation-associated conformational switch. We show that such a switch provides explanations for both how treadmilling may occur within a single-stranded filament, and why filament assembly is cooperative.

microbiology↗