Replacement of Widefield Imaging System

项目总结 亲本资助的中心目标是确定蛋白酶体在减数分裂染色体上的作用。这个 26S蛋白酶体是真核生物蛋白质降解的主要部位。它是一个隔间的， 存在于细胞质和细胞核中的多组分的蛋白酶。胞质蛋白酶体 人们很早就认识到了消除调节蛋白和错误折叠蛋白的功能。功能 对细胞核中的蛋白酶体的了解要少得多。我们最近发现，在减数分裂过程中， 26S蛋白酶体以进化保守的方式招募到它控制的染色体上 同源配对、突触和重组。结合使用高分辨率显微镜，基因 和蛋白质组学方法，我们正在追求建立在这一发现基础上的三个目标：第一，我们正在使用 严密控制的条件等位基因系统地表征不同品种减数分裂过程中的作用 蛋白酶体成分。第二，我们正在鉴定与以下相关的蛋白酶体的底物 染色体配对和重组，提供了对一类需要 被降解以确保正常的减数分裂进程。第三，我们正在确定蛋白酶体的决定因素 染色体上的募集。功能缺失的减数分裂染色体分离缺陷 蛋白酶体强调了更好地了解染色体对生殖健康的重要性- 相关的蛋白分解。我们的分析也与癌症治疗有关，在癌症治疗中，蛋白酶体抑制剂具有 变得越来越重要。我们的许多方法都涉及到高分辨率和活细胞成像 真核生物发芽酵母的模型。在这份补充提案中，我们要求为一项高级 高分辨率的实时成像系统，作为我们老化的当前显微镜的替代品。我们目前的制度 给了我们有价值的见解，但已经变得不可靠，即将结束。最新情况-- ART成像系统将使我们能够保持并进一步增强一个高影响力的研究计划 染色体传递的机制。

PROJECT SUMMARY The central aim of the parent grant is to identify roles of the proteasome along meiotic chromosomes. The 26S proteasome is the main site of protein degradation in all eukaryotes. It is a compartmentalized, multicomponent protease that resides both in the cytoplasm and the nucleus. Cytoplasmic proteasome functions in eliminating regulatory and misfolded proteins have been recognized for a long time. Functions of the proteasome in the nucleus are much less understood. We recently discovered that during meiosis I, the 26S proteasome is recruited in an evolutionarily conserved manner to chromosomes where it controls homolog pairing, synapsis and recombination. Using a combination of high-resolution microscopy, genetic and proteomic approaches, we are pursuing three aims that build on this discovery: First, we are using tightly controlled conditional alleles to systematically characterize roles during meiosis of different proteasome components. Second, we are identifying substrates of the proteasome relevant to chromosome pairing and recombination, providing insights into a class of molecules that need to be degraded to ensure normal meiotic progression. Third, we are identifying determinants for proteasome recruitment to chromosomes. Meiotic chromosome segregation defects in absence of a functional proteasome emphasize the importance for reproductive health of a better understanding of chromosome- associated proteolysis. Our analysis is also relevant to cancer therapy where proteasome inhibitors have become increasingly important. Many of our approaches involve high resolution and live-cell imaging in the model eukaryote budding yeast. In this supplement proposal, we are requesting funds for an advanced high-resolution, live imaging system as replacement for our aging current microscope. Our current system has given us valuable insights but has become unreliable and is near the end of its lifetime. A state-of-the- art imaging system will enable us to maintain and further enhance a high impact research program into mechanisms of chromosome transmission.