Microtubule dynamics and error correction

微管动力学和误差校正

• 批准号: 10640162
• 负责人: Linda Wordeman
• 金额: $ 37.93万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-07 至 2026-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10640162
• 关键词: ATP phosphohydrolase; Antineoplastic Agents; Biological Assay; Brain; CRISPR/Cas technology; Cell Culture Techniques; Cell Line; Cell Survival; Cell division; Cells; Centrosome; Chromosomal Instability; Chromosome Segregation; Chromosomes; Classification; Clustered Regularly Interspaced Short Palindromic Repeats; Congenital Abnormality; Congresses; Coupled; Development; Embryo; Embryonic Development; Engineering; Enhancers; Enzymatic Biochemistry; Fluorescence; Fluorescence Microscopy; Functional disorder; Future; Generations; Genome; Geometry; Goals; Human; Image; Imaging Device; Impairment; Investigation; Kinesin; Kinetochores; Length; MXD1 gene; Malignant Neoplasms; Microscopy; Microtubules; Mitosis; Mitotic; Motor; Pathway interactions; Pharmaceutical Preparations; Pharmacology; Phosphotransferases; Positioning Attribute; Predisposition; Proteasome Inhibitor; Proteins; Resistance; Role; Sirolimus; Testing; Therapeutic; Transgenic Organisms; Tubulin; Vertebrates; Zebrafish; cell fixing; cohesion; embryo cell; hatching; knowledge base; mutant; nerve stem cell; neurodevelopment; new therapeutic target; overexpression; premature; preservation; prevent; public health relevance; single molecule; small molecule; stem cell division; stem cells; targeted agent; therapeutic target; tumorigenic

Modified Project Summary/Abstract Section The broad goal of this project is to understand how force generation by kinesin-related motors and dynamic microtubules controls the fidelity of chromosome segregation. Both microtubules and motors represent excellent targets for anti-cancer drugs but to make wise choices for developing therapeutics it is necessary to understand their contribution to cell division in detail. MCAK/Kif2C is a MT depolymerizing kinesin that controls MT length within the spindle and supresses chromosome instability (CIN). Using CRISPR/Cas9 engineered cells, rapamycin-dependent relocalization and long-term live imaging we have pinpointed the precise contribution that this protein makes to enhance the fidelity of chromosome segregation which will enable us to understand why this protein rescues CIN in tumorigenic cells and its future potential as a therapeutic target. MCAK/Kif2C is also strongly associated with centrosomes where it has the potential to suppress MT outgrowth and influence centrosome separation. We have identified two other poorly studied centrosome-associated kinesins, Kif25 and Kif9, that function in centrosome cohesion and centrosome satellite positioning respectively and that also impact centrosome separation and positioning. Altered centrosome positioning in cell culture has limited effects on cell viability. However, Both Kif9 and Kif25 are widely expressed in vertebrates. Because Kif25, in particular, is most highly expressed in brain it affords an opportunity to investigate the role of centrosome cohesion in vertebrate development and neural stem cell division using morpholinos directed against Kif25 introduced into early zebrafish embryos. We aim to use live and fixed-cell imaging tools and transgenic cell lines to investigate, with precision, the contribution that these kinesins make to preserving the fidelity of the genome and to long-term cell fate. Ultimately this knowledge base can be leveraged to develop new therapies that target kinesin enzymology or MT dynamics.

修改后的项目摘要/摘要部分 这个项目的主要目标是了解动蛋白相关的马达和动态微管产生的力是如何控制染色体分离的保真度的。微管和马达都是抗癌药物的极佳靶点，但要为开发治疗药物做出明智的选择，有必要详细了解它们对细胞分裂的贡献。MCAK/KIF2C是一种MT解聚运动蛋白，控制纺锤体内MT的长度，抑制染色体不稳定性(CIN)。利用CRISPR/Cas9工程细胞、雷帕霉素依赖的重定位和长期的活体成像，我们已经准确地确定了该蛋白对提高染色体分离的保真度所做出的准确贡献，这将使我们能够理解该蛋白为什么拯救肿瘤细胞中的CIN及其作为治疗靶点的未来潜力。MCAK/KIF2C也与中心体密切相关，它有可能抑制MT的生长并影响中心体的分离。我们已经确定了另外两个研究较少的中心体相关Kinesin，Kif25和Kif9，它们分别在中心体凝聚和中心体卫星定位中发挥作用，也影响中心体分离和定位。细胞培养中中心体位置的改变对细胞活力的影响有限。然而，Kif9和Kif25在脊椎动物中广泛表达。由于Kif25在大脑中的表达最高，这为研究中心体凝聚力在脊椎动物发育和神经干细胞分裂中的作用提供了机会，将针对Kif25的吗啡引入斑马鱼早期胚胎。我们的目标是使用活的和固定的细胞成像工具和转基因细胞系来精确地研究这些动蛋白对保持基因组的保真度和对长期细胞命运的贡献。最终，这个知识库可以被用来开发针对动蛋白酶学或MT动力学的新疗法。