Assembly, Disassembly and Function of the Mitotic Checkpoint Complex

University Of Toledo, Toledo OH

Investigators

Abstract

Intellectual merit: The research objective of the project is to investigate the assembly, disassembly and function of a fundamentally important protein complex that prevents erroneous chromosome segregation. The genome of a eukaryotic cell is packaged into individual chromosomes. Faithful transmission of a full complement of chromosomes into daughter cells requires delicate control mechanisms during cell division (mitosis). Of these critical mechanisms, the mitotic checkpoint (or spindle assembly checkpoint) ensures that duplicated sister chromatids are not separated until they are properly attached by microtubules emanating from opposite spindle poles. At the molecular level, the checkpoint is executed by inhibiting an enzyme termed the Anaphase Promoting Complex/Cyclosome (APC/C) whose activation underlies anaphase onset and exit from mitosis. How the APC/C is inhibited during early mitosis is not yet resolved. A four-subunit Mitotic Checkpoint Complex (MCC), comprised of BUBR1 (MAD3 in yeast), BUB3, CDC20 and MAD2, was found to bind and inhibit the APC/C efficiently under physiologically relevant protein concentrations in cells. However, there have been continuous debates in the past decade concerning whether MAD2 is an integral component of the MCC. Partly due to this identity crisis', how MCC activity is regulated remains elusive. The PI's group has recently demonstrated that a specific conformation of MAD2 (C-MAD2) and a previously unknown direct interaction between BUBR1 and C-MAD2 are essential for the assembly and function of the MCC. The discoveries, made by graduate and undergraduate researchers, help resolve earlier inconsistencies and open the door to further characterization of the MCC. Three specific aims will be addressed in the proposal to investigate how MCC is assembled; how MCC inhibits APC/C; and how MCC disassembly is regulated. A combination of molecular, cellular, and biochemical approaches will be used in the project. Cell division is one of the most fundamental processes for life. If successful, the project will significantly advance current understandings of the control mechanisms for the timing of chromosome segregation. A coherent model of the MCC:APC/C interaction will further accelerate discoveries of additional factors that impact the fidelity of chromosome segregation through modulating MCC and APC/C activities. Broader impact: The primary educational objective of the project is to retain aspiring young researchers in science careers through research-based engagement. The natural beauty of mitosis is appealing to graduate, undergraduate and high school students, but how to help young people to convert transient interest in science into lasting passion for research is still challenging. Level-appropriate career guidance and research activities will be bundled to engage students, particularly those from underrepresented groups, in scientific exploration. Different programs managed by the University of Toledo Office of Undergraduate Research (OUR-UT) and the UT High School Outreach Initiatives Office will help the PI to identify and recruit enthusiastic young researchers. The PI is also interested in exploring efficient channels to communicate scientific progress with the public. An interactive lab website and a potential museum exhibit are proposed as means to introduce their research to a broader audience. The results of proposed educational activities will not only support realization of specific research goals, but will also aid in design of effective curricula and activities to stimulate the interest in science careers among young people, and promote public understanding of science.