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.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM094972-12
Application #
9069446
Study Section
Cellular Signaling and Regulatory Systems Study Section (CSRS)
Program Officer
Melillo, Amanda A
Project Start
2005-07-01
Project End
2018-05-31
Budget Start
2016-06-01
Budget End
2017-05-31
Support Year
12
Fiscal Year
2016
Total Cost
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
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
Jones, Mathew J K; Jallepalli, Prasad V (2016) Engineering and Functional Analysis of Mitotic Kinases Through Chemical Genetics. Methods Mol Biol 1413:349-63
Kim, Minhee; O'Rourke, Brian P; Soni, Rajesh Kumar et al. (2016) Promotion and Suppression of Centriole Duplication Are Catalytically Coupled through PLK4 to Ensure Centriole Homeostasis. Cell Rep 16:1195-1203
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Rahman, Sadia; Jones, Mathew J K; Jallepalli, Prasad V (2015) Cohesin recruits the Esco1 acetyltransferase genome wide to repress transcription and promote cohesion in somatic cells. Proc Natl Acad Sci U S A 112:11270-5
Chen, Yu-Hung; Jones, Mathew J K; Yin, Yandong et al. (2015) ATR-mediated phosphorylation of FANCI regulates dormant origin firing in response to replication stress. Mol Cell 58:323-38
Rodriguez-Bravo, Veronica; Maciejowski, John; Corona, Jennifer et al. (2014) Nuclear pores protect genome integrity by assembling a premitotic and Mad1-dependent anaphase inhibitor. Cell 156:1017-31
Jones, Mathew J K; Jallepalli, Prasad V (2012) Chromothripsis: chromosomes in crisis. Dev Cell 23:908-17
Oppermann, Felix S; Grundner-Culemann, Kathrin; Kumar, Chanchal et al. (2012) Combination of chemical genetics and phosphoproteomics for kinase signaling analysis enables confident identification of cellular downstream targets. Mol Cell Proteomics 11:O111.012351

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