Free SepF interferes with recruitment of late cell division proteins

Abstract

The conserved cell division protein SepF aligns polymers of FtsZ, the key cell division protein in bacteria, during synthesis of the (Fts)Z-ring at midcell, the first stage in cytokinesis. In addition, SepF acts as a membrane anchor for the Z-ring. Recently, it was shown that SepF overexpression in Mycobacterium smegmatis blocks cell division. Why this is the case is not known. Surprisingly, we found in Bacillus subtilis that SepF overproduction does not interfere with Z-ring assembly, but instead blocks assembly of late division proteins responsible for septum synthesis. Transposon mutagenesis suggested that SepF overproduction suppresses the essential WalRK two-component system, which stimulates expression of ftsZ. Indeed, it emerged that SepF overproduction impairs normal WalK localization. However, transcriptome analysis showed that the WalRK activity was in fact not reduced in SepF overexpressing cells. Further experiments indicated that SepF competes with EzrA and FtsA for binding to FtsZ, and that binding of extra SepF by FtsZ alleviates the cell division defect. This may explain why activation of WalRK in the transposon mutant, which increases ftsZ expression, counteracts the division defect. In conclusion, our data shows that an imbalance in early cell division proteins can interfere with recruitment of late cell division proteins.

Systematic Identification of Target Genes for Cellular Morphology Engineering in Synechococcus elongatus PCC7942

“…This might subsequently impair the formation and function of cell divisome and elongate the mutant cells. A similar mechanism could also explain the phenomenon that the overexpression of Cdv3 and ZipN resulted in significantly increased cell size (5.8- and 6.2-folds increased lengths than that of the wild type control, as shown in Table 1 and Figures 3B,C ), because excessive abundances of specific factors for the cell division might competitively block and inhibit the interaction and affinity of the subsequent cells, subsequently impaired the normal cell division and generated elongated cells ( Gao et al, 2017 ). An additional phenotype supporting this hypothesis is the increased frequency of uneven cell divisions ( Figure 3A ).…”

“…Excessive accumulation of other factors potentially involved in cell morphogenesis, including Cdv1, RodA, FtsE, FtsI, FtsW, SulA, and Ftn6, caused similar effects on cell length, with increase ranging from 1.2- to 1.4-fold, while rod-shapes of the cells were maintained ( Table 1 and Figure 3A ). Although it has been reported previously that overexpression of SepF in B. subtilis resulted in filamentation of the mutant cells and eventually cell deaths ( Gao et al, 2017 ), the excessive abundance of the homologous protein Cdv2 in PCC7942 caused minor effects on cellular morphology (with 10% increased cell lengths and 10% decreased cell area, Table 1 ). This indicated that the Cdv2/SepF was not as essential as other components such as ZipN and Cdv3 for Z-ring stability and functionality in PCC7942.…”

Branching of sporogenic aerial hyphae in sflA and sflB mutants of Streptomyces coelicolor correlates to ectopic localization of DivIVA and FtsZ in time and space

“…1), only SepF interacts with FtsZ (Schlimpert et al, 2017). Interestingly, in Mycobacterium smegmatis and B. subtilis, overexpression of SepF is lethal and largely blocks cell division, and it was suggested that this was due to interference of free SepF with the assembly of lateral cell division proteins (Gao et al, 2017;Gola et al, 2015). In S. coelicolor however, overexpression of SepF barely showed any effect, while overexpression of its paralogs SflA or SflB let to growth inhibited.…”

Control of septum thickness by the curvature of SepF polymers

“…The medium was supplemented with 0.1 mM IPTG to induce expression of SepF variants, where appropriate. It is important not to use more IPTG since SepF overproduction causes membrane deformations that obscure septa (41). Overnight cultures were grown with appropriate antibiotic concentrations (100 μg/mL spectinomycin, 10 μg/mL chloramphenicol, 5 μg/mL kanamycin, 1 μg/mL erythromycin), where necessary.…”