Abstract
Cell division in bacteria is an essential process that is carried out at mid-cell by a group of cell division proteins referred to as the divisome. In Escherichia coli, over two dozen cell division proteins have been identified of which ten are essential. These division proteins localize sequentially and interdependently to the division site, after which constriction eventually produces two daughter cells. Various genetic and biochemical techniques have identified many interactions amongst cell division proteins, however the existence of the divisome as a large multi-protein complex has never been shown. Here, we identify a 1 MDa protein complex by native page that contains seven essential cell division proteins (FtsZ, ZipA, FtsK, FtsQ, FtsB, FtsL, and FtsN). The 1 MDa complex is present in rapidly dividing cells, but absent when cultures enter the stationary growth phase. Slight overexpression of the ftsQ D237N mutation that blocks cell division prevents formation of this 1 MDa complex. In cells depleted of FtsN, the 1 MDa complex is not assembled. Combined, our findings indicate that a large protein complex containing many different cell division proteins indeed exists. We note that this complex is very fragile and sensitive to the expression of tagged versions of FtsQ.
Highlights
Impairment or even overexpression of one protein in the interdependent sequence described above can cause other downstream proteins to fail to localize to the division site
An extensive Bacterial Two Hybrid (BACTH) study found that FtsZ interacts with FtsA, ZipA, FtsK; FtsW interacts with FtsL, FtsN, FtsI; FtsL interacts with FtsK, FtsQ, FtsW; FtsI interacts with FtsA, FtsQ, FtsN, FtsI; and that FtsQ interacts with FtsK, FtsI, FtsN, FtsL31
To study whether cell division proteins form a large multi-protein complex in the cellular envelope of Escherichia coli, total cell fractions of mildly solubilized bacteria were analyzed by native page
Summary
Impairment or even overexpression of one protein in the interdependent sequence described above can cause other downstream proteins to fail to localize to the division site. When a downstream division protein is moved upstream by fusing it to an upstream protein it is able to back-recruit other upstream proteins and division can proceed, resulting in viable cells[11] These findings provide hints towards the existence of the divisome as a protein super-complex or a temporary assembly that is comprised of all cell division proteins. Oligomeric interactions comprising multiple proteins that prove that the divisome is present as a large multi-protein machinery in the cell, have so far not been shown. We describe the identification of a large 1 MDa cell division protein complex, that includes at least 7 essential division proteins; FtsZ, ZipA, FtsK, FtsQ, FtsB, FtsL, and FtsN. Our findings indicate that a large protein complex containing cell division proteins exists. Our findings have implications for the reconstitution of the divisome in vitro as well as for the interpretation of data obtained with GFP-fusions to cell division proteins
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