In all eukaryotes, cytokinesis depends on the contraction of the actomyosin ring, a ring of actin, myosin, and several other proteins that forms at the division site. The timing of contraction and subsequent disassembly of the actomyosin ring are poorly understood. The anaphase-promoting complex (APC) is an E3 ubiquitin ligase that controls several key steps in mitosis by targeting specific proteins for degradation. We hypothesized that the APC also controls cytokinesis by regulating the actomyosin ring.
We began by searching for novel APC substrates among proteins known to function in cytokinesis. Out of over 75 proteins tested, we found that Iqg1, an essential component of the actomyosin ring, is an APC substrate in vitro and is degraded in an APC-dependent manner after cytokinesis in vivo. To characterize the functional interaction between the APC and the actomyosin ring, we used live cell microscopy to analyze the behavior of the actomyosin ring components in apc mutants. In wild-type cells, fluorescent versions of actomyosin ring components visibly form a ring at the bud neck, then contract and disappear from the site of cytokinesis. In cells lacking CDH1, the APC activator during the time of cytokinesis, we find that these proteins behave normally during actomyosin ring contraction but then co-localize in multiple foci, rather than disappearing. From this we infer that the mutants cannot disassemble the actomyosin ring, and that APCCdh1 may be essential for this process.
Actomyosin disassembly defects have been reported in septin (a structural scaffold for cytokinesis proteins) and myosin regulatory light chain (mlc2) mutant cells. We find that deletion of CDH1 exacerbates these defects, which suggests that APCCdh1 functions in parallel to both septins and MLC2. In addition, we find that APCCdh1-mediated degradation of Iqg1 and Mlc2 activity are both required for actomyosin ring disassembly. Therefore, the APC, septins, and myosin regulatory light chain cooperate to ensure complete disassembly of the cytokinesis machinery.
The APC controls entry to and exit from mitosis. The results from these experiments demonstrate a function of the APC in cytokinesis, regulating disassembly of the actomyosin ring.