Characterization of the Synapis Checkpoint in C. elegans Meiosis

描述（由申请人提供）：减数分裂从二倍体细胞产生单倍体配子，使得在受精后恢复二倍体基因组。减数分裂过程中染色体的正确分离取决于减数分裂前期的事件，如同源染色体的配对和联会以及交换重组。染色体分离的错误通常对受精卵是致命的，但也可能导致癌症倾向或严重的发育障碍。我已经确定了一个减数分裂检查点，响应同源突触的缺陷，独立的DNA损伤/重组检查点，并激活细胞凋亡，以避免非整倍体配子的产生。并不是所有的非突触序列都有能力触发这个检查点;相反，这个通路是由非突触配对中心（PC）特异性激活的，非突触配对中心是促进C染色体中突触的染色体位点。优雅此外，检查点需要C。线虫PCH-2的同源物，芽殖酵母粗线期检查点基因，这表明检测突触失败的分子机制是广泛保守的。 我计划进一步描述这个突触检查点。我特别感兴趣的是PC对突触检查点激活的贡献。与PC功能所需的因子相互作用的蛋白质的鉴定和表征将提供对该位点在未突触时如何激活检查点的深入了解。还将进行研究，解决非突触染色体上异染色质的调节以及PC如何抑制DNA损伤检查点。我将通过表征与联会复合体（SC）相互作用的两个基因来确定联会复合体（SC）在突触检查点中的作用，并且通过初步RNA干扰（RNAi）实验似乎是检查点所需的。我将研究已知的检查点组件pch-2的功能和调节; GFP-PCH-2融合蛋白将定位于各种遗传背景中，并将提供试剂以生物化学方式识别相互作用的蛋白质。此外，我将通过进行RNAi筛选来确定检查点的其他组成部分，该筛选将关注满足特定表达和表型特征标准的候选基因。这些互补的方法将使我能够从分子和机制上理解同源物突触是如何被监测的，以及未突触或不适当突触的同源物如何产生最终转化为凋亡反应的检查点信号。 减数分裂产生配子，如卵子和精子。检查点监测减数分裂事件，以确保配子具有正确的染色体数目。如果一个配子的染色体数目不正确，受精产生的胚胎通常是不能存活的。偶尔，胚胎会遗传一条额外的染色体，这条染色体不是致命的，但可能导致癌症易感性或严重的发育缺陷。

DESCRIPTION (provided by applicant): Meiosis generates haploid gametes from a diploid cell such that a diploid genome is restored upon fertilization. The proper segregation of chromosomes during the meiotic divisions depends on events in meiotic prophase, such as the pairing and synapsis of homologous chromosomes and crossover recombination. Errors in chromosome segregation are usually fatal to the fertilized zygote but can also result in cancer predisposition or serious developmental disorders. I have identified a meiotic checkpoint that responds to defects in homolog synapsis, independent of a DNA damage/recombination checkpoint, and activates apoptosis to avoid the generation of aneuploid gametes. Not all unsynapsed sequences have the capacity to trigger this checkpoint; rather, this pathway is specifically activated by unsynapsed pairing centers (PCs), chromosome sites that promote synapsis in C. elegans. Furthermore, the checkpoint requires the C. elegans homolog of PCH-2, a budding yeast pachytene checkpoint gene, suggesting that the molecular mechanism that detects synaptic failure is widely conserved. I plan to further characterize this synapsis checkpoint. I am particularly interested in the PC's contribution to synapsis checkpoint activation. The identification and characterization of proteins that interact with factors required for PC function will provide insight into how this locus activates the checkpoint when unsynapsed. Studies that address the regulation of heterochromatin on unsynapsed chromosomes and how the PC may inhibit the DNA damage checkpoint will also be undertaken. I will determine the role of the synaptonemal complex (SC) in the synapsis checkpoint by characterizing two genes that interact with the SC and appear to be required for the checkpoint by preliminary RNA inteferference (RNAi) experiments. I will investigate the function and regulation of the known checkpoint component, pch-2; a GFP-PCH-2 fusion protein will be localized in a variety of genetic backgrounds and a reagent will be provided to identify interacting proteins biochemically. Furthermore, I will identify additional components of the checkpoint by undertaking an RNAi screen that will focus on candidate genes that fulfill specific expression and phenotypic profile criteria. These complementary approaches will enable me to gain a molecular and mechanistic understanding of how homolog synapsis is monitored and how an unsynapsed or inappropriately synapsed homolog generates a checkpoint signal that is ultimately translated into an apoptotic response. Meiosis produces gametes, such as eggs and sperm. Checkpoints monitor meiotic events to ensure that gametes have the correct number of chromosomes. If a gamete has an incorrect number of chromosomes, the embryo that results from fertilization is often inviable. Occasionally, an embryo inherits an extra chromosome that is not lethal but can cause cancer predisposition or serious developmental defects.