Uncovering the Role of Synaptonemal Complex in Meiotic Recombination by Cryo-electron Tomography

通过冷冻电子断层扫描揭示联会复合体在减数分裂重组中的作用

• 批准号: 10460739
• 负责人: Vignesh Kasinath
• 金额: $ 24.9万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-14 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10460739
• 关键词: 3-Dimensional; Actins; Address; Advisory Committees; Aneuploidy; Animal Model; Architecture; Biochemical; Caenorhabditis elegans; Cell Nucleus; Cells; Chromatin Remodeling Factor; Chromosome Pairing; Chromosome Segregation; Chromosomes; Communities; Complex; Congenital Abnormality; Congenital chromosomal disease; Coupled; Cryo-electron tomography; Cryoelectron Microscopy; Cytoplasm; Defect; Development; Devices; Down Syndrome; Dynein ATPase; Electron Microscopy; Elements; Environment; Event; Experimental Designs; Foundations; Future; Genetic; Genetic Crossing Over; Genetic Recombination; Genetic Variation; Germ Cells; Goals; Gonadal structure; Homologous Gene; In Situ; Infertility; Infrastructure; Ions; Lead; Mediating; Meiosis; Meiotic Prophase I; Meiotic Recombination; Mentorship; Methods; Microtubules; Modality; Modeling; Molecular; Molecular Biology; Morphology; Movement; Multiprotein Complexes; Nodule; Noonan Syndrome; Nuclear; Nuclear Envelope; Organism; Outcome Study; Phase; Phase Transition; Play; Polycomb; Preparation; Process; Production; Protein Kinase; Proteins; Regulation; Research; Resolution; Rest; Role; Saccharomyces cerevisiae; Sampling; Scanning Electron Microscopy; Site; Structure; Synaptonemal Complex; System; Three-Dimensional Imaging; Time; TimeLine; Training; Transcriptional Regulation; Work; Yeasts; biochemical tools; career; chromosome movement; cohesin; comparative; computerized tools; developmental disease; electron tomography; env Gene Products; homologous recombination; in vivo; innovation; insight; mutant; notch protein; particle; spatiotemporal; three dimensional structure; tool; zygote

PROJECT SUMMARY/ABSTRACT Meiosis in sexually reproducing organisms is critical for generating genetic diversity and the production of healthy gametes. The synaptonemal complex is a dynamic and conserved macromolecular protein complex which is crucial for chromosome pairing, meiotic recombination and proper chromosome segregation. Errors or defects in these processes result in zygotes, which die early in development, infertility and aneuploidy, resulting in developmental disorders like Turner syndrome, Down syndrome. The lack of structural framework for the synaptonemal complex has hindered mechanistic insights into its role in critical meiotic processes. My proposal combines innovative electron tomography methods and molecular biology tools to define the complete molecular architecture of the synaptonemal complex in paired chromosomes using the tractable systems, C. elegans and S. cerevisiae. During the K99/R00 period I will, 1) Construct the spatiotemporal landscape of synaptonemal complex architecture changes from early to late pachytene in C. elegans for all six chromosome pairs and highlight the role played by key protein factors in regulating the concerted change in synaptonemal complex architecture and meiotic recombination processes. 2) Determine a high-resolution molecular view of the synaptonemal complex for both S. cerevisiae and C. elegans in order to (a) provide insight into its subunit organization and evolutionary conservation, (b) examine its interactions with both chromosome axis proteins and recombination nodules - crossover machinery, to delineate its role in homologous recombination and chromosome architecture changes during pachytene. 3) Provide mechanistic insight into homologous chromosome recognition by elucidating the role played by nuclear envelope proteins in regulating cytoskeletal forces (microtubules, dynein) and chromosome movements critical for homolog recognition and subsequent initiation of chromosome pairing. During my postdoctoral work in the Nogales lab, I have obtained training in single-particle cryo-electron microscopy and provided structural insights into the regulation of chromatin modifier, Polycomb Repressive Complex 2. During my K99/R00 phase, I will undertake training in electron tomography methods and genetics tools that will enable me to study critical processes during meiosis, in vivo. I am confident that my training in electron tomography coupled with the excellent mentorship of Eva Nogales, Elizabeth Villa, Abby Dernburg, and the rest of my advisory team, will help me transition to an independent research career. I believe my access to top notch scientific infrastructure and a truly collaborative scientific community at UC Berkeley makes it the ideal environment for my K99/R00 training. During my R00 phase, I will provide insight into the molecular mechanisms governing homologous chromosome recognition and how defects in these mechanisms lead to chromosomal disorders. I envisage developing a cross-disciplinary research group utilizing electron microscopy, biochemistry, and computational tools to address problems concerning chromosome architecture and transcription regulation.

项目摘要/摘要 有性生殖生物体的减数分裂对于产生遗传多样性和生产健康的 配子。联会复合体是一种动态的、保守的大分子蛋白质复合体，是 对染色体配对、减数分裂重组和适当的染色体分离至关重要。错误或缺陷 在这些过程中产生受精卵，受精卵在发育早期死亡，不育和非整倍体，导致 发育障碍，如特纳综合征，唐氏综合症。缺乏结构框架， 联会复合体阻碍了对其在关键减数分裂过程中作用的机械性洞察。 我的建议结合了创新的电子断层扫描方法和分子生物学工具来定义 用易处理的方法研究配对染色体中联会复合体的完整分子结构 系统、线虫和酿酒酵母。在K99/R00期间，我将， 1)构建了联会复合体建筑由早到晚的时空景观 线虫中所有六对染色体的粗线期，并强调关键蛋白质因子在 调节联会复合体结构和减数分裂重组过程的协同变化。 2)确定了酿酒酵母和啤酒酵母的联会复合体的高分辨率分子视图。 为了(A)深入了解其亚基组织和进化保护，(B)审查 它与染色体轴蛋白和重组结节的相互作用-交换机制，以 描述其在粗线期同源重组和染色体结构变化中的作用。 3)通过阐明同源染色体识别所扮演的角色，提供对同源染色体识别的机械洞察力 核膜蛋白在调节细胞骨架力量(微管、动力蛋白)和染色体运动中的作用 对于同源基因识别和随后的染色体配对的启动至关重要。 在诺加莱斯实验室的博士后工作期间，我接受了单粒子冷冻电子的培训。 显微镜观察和提供了对染色质修饰物、多梳抑制的调节的结构洞察 在我的K99/R00阶段，我将接受电子断层扫描方法和遗传学方面的培训 这些工具将使我能够在体内研究减数分裂过程中的关键过程。我相信我在中国的训练 电子断层扫描与伊娃·诺加莱斯、伊丽莎白·维拉、艾比·邓伯格和 我的顾问团队的其余成员将帮助我过渡到独立的研究生涯。我相信我能接触到 加州大学伯克利分校一流的科学基础设施和真正的协作科学社区使其成为理想的 我的K99/R00培训的环境。在我的R00阶段，我将提供对分子机制的见解 控制同源染色体识别以及这些机制中的缺陷如何导致染色体 精神错乱。我设想建立一个跨学科的研究小组，利用电子显微镜，生物化学， 以及解决有关染色体结构和转录调控问题的计算工具。