Posttranslational modification with ubiquitin chains is essential for cell division in all eukaryotes. Ubiquitin chains are synthesized by a cascade of E1 ubiquitin-activating, E2 ubiquitin-conjugating, and E3 ubiquitin ligase enzymes, with the latter defining the substrate specificity of this modification. Among ~600 human E3s, the anaphase-promoting complex (APC/C) is essential for mitosis, and the aberrant expression of APC/C- regulators or substrates has been tightly linked to tumorigenesis. Moreover, the indirect inhibition of the APC/C by paclitaxel, a small molecule that activates the APC/C-inhibitory spindle checkpoint, has become a mainstay of chemotherapy. We have recently discovered that the APC/C assembles chains of a novel topology, linked through Lys11 of ubiquitin. The APC/C achieves K11-linkage formation by employing an initiating E2, Ube2C, and an elongating E2, Ube2S. Similar to the APC/C, K11-linked chains and the E2-module composed of Ube2C and Ube2S are required for cell division. Therefore, dissecting the APC/C-dependent assembly and function of K11-linked chains is critical to our understanding of cell division. Here, we propose t dissect the mechanism of K11-linked ubiquitin chain formation by using a set of unique biochemical and structural assays combined with ubiquitin- and E2-mutants derived from extensive biochemical screens. We will directly test for functions of K11-linked ubiquitin chains by employing the novel approach of E3-reprogramming that allows us to switch the topology of ubiquitin chains that are assembled by the APC/C in cells. Finally, we will investigate the regulation of K11-linked chain formation, using a group of cancer-related APC/C-substrates, the spindle assembly factors HURP, NuSAP, and Tpx2, as our model system. Together, we expect that our studies will reveal core principles of ubiquitylation and cell cycle regulation. In this manner, our work will likely provide guidance for the development of novel classes of chemotherapeutics that target essential E2 enzymes.
Errors in cell division lead to aneuploidy, a condition that is tightly linked to birth defects and cancer. Ubiquitin chain formation by the anaphase-promoting complex, APC/C, is essential for accurate cell division. We are, therefore, studying how the tightly regulated APC/C enables flawless cell division, allowing us to determine the molecular basis of several proliferative diseases.
|Cappell, Steven D; Mark, Kevin G; Garbett, Damien et al. (2018) EMI1 switches from being a substrate to an inhibitor of APC/CCDH1 to start the cell cycle. Nature 558:313-317|
|Yau, Richard G; Doerner, Kerstin; Castellanos, Erick R et al. (2017) Assembly and Function of Heterotypic Ubiquitin Chains in Cell-Cycle and Protein Quality Control. Cell 171:918-933.e20|
|Werner, Achim; Manford, Andrew G; Rape, Michael (2017) Ubiquitin-Dependent Regulation of Stem Cell Biology. Trends Cell Biol 27:568-579|
|Werner, Achim; Iwasaki, Shintaro; McGourty, Colleen A et al. (2015) Cell-fate determination by ubiquitin-dependent regulation of translation. Nature 525:523-7|
|Meyer, Hermann-Josef; Rape, Michael (2014) Enhanced protein degradation by branched ubiquitin chains. Cell 157:910-21|
|Song, Ling; Craney, Allison; Rape, Michael (2014) Microtubule-dependent regulation of mitotic protein degradation. Mol Cell 53:179-92|
|Williamson, Adam; Werner, Achim; Rape, Michael (2013) The Colossus of ubiquitylation: decrypting a cellular code. Mol Cell 49:591-600|
|Jin, Lingyan; Pahuja, Kanika Bajaj; Wickliffe, Katherine E et al. (2012) Ubiquitin-dependent regulation of COPII coat size and function. Nature 482:495-500|
|Meyer, Hermann-Josef; Rape, Michael (2011) Processive ubiquitin chain formation by the anaphase-promoting complex. Semin Cell Dev Biol 22:544-50|
|Wickliffe, Katherine E; Lorenz, Sonja; Wemmer, David E et al. (2011) The mechanism of linkage-specific ubiquitin chain elongation by a single-subunit E2. Cell 144:769-81|
Showing the most recent 10 out of 18 publications