Accurate chromosome segregation is vital for cell proliferation, tissue homeostasis, embryonic development, and tumor suppression. One key regulatory pathway is the spindle assembly checkpoint (SAC), which prevents the separation of sister chromatids and exit from mitosis until all chromosomes are linked to both spindle poles by microtubule fibers. Even a single chromosome lacking bipolar attachment is sufficient to trigger the SAC, as its kinetochores recruit and activate downstream factors that not only communicate with the core cell-cycle machinery, but also alter the microtubule-binding properties of the kinetochore itself, so that incorrect microtubule attachments are destabilized. Recent work from our lab has implicated the protein kinase Mps1 in both of these outputs, as well as in a third pathway that acts as a mitotic 'clock' or 'timer' independently of kinetochores. These insights emerged through combined application of gene editing and chemical genetics techniques pioneered in our lab, whereby endogenous Mps1 in cultured human cells was deleted from the genome and replaced by a variant kinase allele sensitized to bulky purine analogs. Using this system, we have performed global and targeted proteomics screens and generated an extensive suite of phosphospecific antibodies, revealing the landscape of Mps1-dependent phosphorylation at the kinetochore-microtubule interface.
In Aim 1, we will mine this information to analyze how Mps1 and counteracting phosphatases regulate kinetochore-microtubule attachments, such that only proper bipolar attachments are stabilized.
In Aim 2, we dissect how Mps1-catalyzed phosphorylation fuels the recruitment and activation of SAC effectors at kinetochores, resulting in the production of biochemical inhibitors of the APC/C-Cdc20 ubiquitin ligase.
In Aim 3, we use chemical genetics to ask how Mps1 and other kinases interact to maintain the M phase state when the SAC is engaged. These studies will illuminate the molecules and mechanisms underlying M phase quality control, and in the long term will empower development of therapeutic agents that target aneuploidy-associated diseases such as cancer.

Public Health Relevance

Errors in transmitting chromosomes to dividing cells are strongly linked to human diseases, including infertility, spontaneous miscarriages, birth defects, and malignancy. The studies in this project seek to understand how the process of chromosome transmission is controlled at the molecular level. This information is necessary to understand how errors in chromosome transmission arise in the course of disease, and to develop new therapies that exploit this trait as an 'Achilles heel' of most cancer cells, providing more effective and less toxic cures for this disorder.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Cellular Signaling and Regulatory Systems Study Section (CSRS)
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Melillo, Amanda A
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Sloan-Kettering Institute for Cancer Research
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Combes, Guillaume; Barysz, Helena; Garand, Chantal et al. (2018) Mps1 Phosphorylates Its N-Terminal Extension to Relieve Autoinhibition and Activate the Spindle Assembly Checkpoint. Curr Biol 28:872-883.e5
Maciejowski, John; Drechsler, Hauke; Grundner-Culemann, Kathrin et al. (2017) Mps1 Regulates Kinetochore-Microtubule Attachment Stability via the Ska Complex to Ensure Error-Free Chromosome Segregation. Dev Cell 41:143-156.e6
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