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

Abstract

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.

Diversity, Origin and Evolution of the ESCRT Systems

“…Two independent recent studies reported PRC barrel protein to be essential for cell division in Haloferax volcanii (51, 69). It has been shown that two PRC barrel proteins form a tripartite complex with SepF which recruits FtsZ to the divisosome (51, 69). It has been thought previously that Thermoproteales had a unique cell division apparatus because neither FtsZ nor ESCRT system genes were identified in these genomes (24).…”

“…During the recent decade, considerable progress has been reached in understanding the functionality of both FtsZ-based and ESCRT-based molecular machineries in archaea (68). Two independent recent studies reported PRC barrel protein to be essential for cell division in Haloferax volcanii (51, 69). It has been shown that two PRC barrel proteins form a tripartite complex with SepF which recruits FtsZ to the divisosome (51, 69).…”

“…Two independent recent studies reported PRC barrel protein to be essential for cell division in Haloferax volcanii (51,69). It has been shown that two PRC barrel proteins form a tripartite complex with SepF which recruits FtsZ to the divisosome (51,69).…”

Differential GTP-dependentin-vitropolymerization of recombinant Physcomitrella FtsZ proteins

“…The CDS of FtsZ1-2 and FtsZ2-1 lacking the N-terminal transit peptides (86 and 42 amino acids, respectively) were amplified via PCR (Supplemental Table S4) and subcloned into the linearized pSVA13429 vector (Nußbaum et al, 2023) via Gibson cloning (Gibson et al, 2009). The final vectors were sequenced with SUMO_fwd and T7_rev (Supplemental Table S4).…”

“…Cleavage was performed overnight at 4 °C using 20 ng/mL His-Ulp1 protease (SUMO-protease) in the presence of 2 mM DTT, 0.1 % ND-40, 200 μM GDP, and 1 mM MgCl 2 . SUMO protease was produced in-house (Lau et al, 2018; Nußbaum et al, 2023). Following the removal of the His 6 _SUMO-tag, the pH was increased to pH 10 for FtsZ1-2, and kept at 9.5 for FtsZ2-1.…”