Determining the rate-limiting processes for cell division in Escherichia coli

A critical cell cycle checkpoint for most bacteria is the onset of constriction when the septal peptidoglycan synthesis starts. According to the current understanding, the arrival of FtsN to midcell triggers this checkpoint in Escherichia coli. Recent structural and in vitro data suggests that recruitment of FtsN to the Z-ring leads to a conformational switch in actin-like FtsA, which links FtsZ protofilaments to the cell membrane and acts as a hub for the late divisome proteins. Here, we investigate this putative pathway using in vivo measurements and stochastic cell cycle modeling. Quantitatively upregulating protein concentrations and determining the resulting division timings shows that FtsN and FtsA numbers are not rate-limiting for the division in E. coli. However, at higher overexpression levels, they affect divisions: FtsN by accelerating and FtsA by inhibiting them. At the same time, we find that the numbers of FtsZ in the cell are rate-limiting for cell divisions in addition to other processes. Altogether, these findings suggest that instead of FtsN, FtsZ protofilaments drive the conformational switch of FtsA and lead to the onset of constriction. Our data is also suggestive that FtsA minirings are not present at any significant numbers in wild-type cells.