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Error Correction in Mammalian Mitosis: Defining Physical Cues and Integration Mechanisms

哺乳动物有丝分裂中的错误纠正：定义物理线索和整合机制

基本信息

批准号：
10674003
负责人：
Megan Kaiulani Chong
金额：
$ 4.31万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10674003
关键词：
3-Dimensional Affinity Aneuploidy Binding Biological Assay Breast Breast Cancer Cell Cancer Etiology Cell division Cells Chromosomal Instability Chromosome Segregation Chromosomes Color Compensation Congenital Abnormality Crowding Cues Development Drug resistance Ensure Event Fiber Grasshoppers Heart Image Individual Inherited Kinetochores Laboratories MCF10A cells MDA MB 231 Malignant Neoplasms Mammalian Cell Mammals Measures Microtubule Stabilization Microtubules Mitosis Mitotic Modeling Molecular Monitor Needles Normal Cell Oncogenic Patients Phosphotransferases Polymerase Positioning Attribute Proteins Resolution Role Signal Transduction Sister Spermatocytes Structure Testing Therapeutic Time Tumor stage Up-Regulation Work cancer cell cancer therapy daughter cell experimental study improved insight live cell imaging mutant new therapeutic target novel novel therapeutics overexpression prevent protein expression real-time images response segregation sensor therapy development tool tumor

项目摘要

Project Summary/Abstract Errors in chromosome segregation give rise to aneuploidy, a hallmark of cancer. Breakdown of mitotic fidelity correlates with both tumor stage and patient drug resistance. Identifying mechanisms that prevent errors in chromosome segregation, and determining how they go wrong in cancer, are essential to developing therapies to either decrease or increase segregation error rates in cancer. The kinetochore attaches chromosomes to spindle microtubules. It segregates chromosomes and monitors their microtubule attachments, stabilizing correct attachments and destabilizing incorrect ones. We now know nearly all mammalian kinetochore proteins, and many have dysregulated expression in cancer. How does the kinetochore detect and correct attachment errors, and fail to do so in cancer? The idea that tension from bi-orientation signals correct attachments is decades-old, originating in Nicklas' pioneering experiments in grasshopper spermatocytes. Yet, how the kinetochore monitors tension and robustly integrates information across its many bound microtubules to regulate attachment stability is not known. In large part, this is due to challenges in applying tension on kinetochores inside cells, in quantitatively tuning kinetochore composition, and in imaging short-lived error correction events in real-time. Our laboratory has recently overcome these challenges, uniquely positioning us to answer these questions. Notably, two candidate kinetochore proteins have been proposed for sensing tension, the kinase AurKB and microtubule polymerase chTOG, and the expression of both is dysregulated in cancer, as is that of the main microtubule binder Hec1. Here, we test defining hypotheses on how normal and cancer cells detect and correct mitotic errors, combining high resolution 3D live-cell imaging, state-of-the-art physical perturbations, and molecular tools in normal and breast cancer cells. In Aim 1, we test the hypothesis that AurKB and chTOG sense tension. We use microneedles to directly apply force to kinetochore-microtubules, measure how attachment stability responds, and assess how these proteins' dysregulated expression alters this response in cancer. In Aim 2, we test models for whether microtubules respond independently or cooperatively to attachment cues such as tension, and test the hypothesis that Hec1 overexpression in cancer cells leads to hyper-stable attachments that may be more challenging to properly correct. We do so by quantitatively tuning kinetochore microtubule binding capacity using mixed Hec1 mutants, and measuring microtubule attachment lifetime using photomarks. In defining critical mechanisms for correcting mitotic errors, and how they are modified in cancer, we expect to identify adapted mechanisms of error correction in cancer cells. For example, some cancer cells may be deficient in error correction, leading to aneuploidy, or have improved error correction to compensate for extra chromosomes. Mechanisms uniquely or preferentially employed in cancer would offer a new therapeutic window.

项目总结/文摘