Analysis of meiotic chromosome synapsis in yeast

描述(申请人提供)：减数分裂是有性生殖生物体用来从二倍体细胞产生单倍体配子的一种特殊的细胞分裂。染色体数量的减少是通过在一轮染色体复制之后进行两轮染色体分离来实现的。人类减数分裂染色体分离的失败会导致不孕不育和出生缺陷，如21三体，这是美国智力低下的主要原因。此外，减数分裂染色体分离需要修复程序性双链断裂(DSB)。有丝分裂细胞中的DSB修复对于基因组的完整性和癌症的预防至关重要。芽殖酵母是研究减数分裂的良好模式系统。突变为同源减数分裂基因的小鼠和酵母的表型相似，表明对酵母减数分裂的研究可能对人类的减数分裂具有启发性。第一次减数分裂(MI)的独特之处在于姐妹染色单体分离到同一极点(称为还原分离)。适当的MI分离要求同源染色体通过非姊妹染色单体之间的交叉以及姊妹染色单体的凝聚力相互连接。最近的工作表明，在减数分裂过程中，非姐妹染色单体之间的交叉在一定程度上是通过主动抑制姐妹染色单体之间的DSB修复而促进的。这种抑制是由MEK1介导的，MEK1是一种减数分裂特异的丝氨酸/苏氨酸激酶。要了解MEK1抑制减数分裂姐妹染色单体修复的分子机制，需要鉴定由该激酶磷酸化的底物。我们将使用新的生化和遗传策略来识别MEK1靶标，并对这些蛋白质的磷酸化缺陷版本进行表征，阐明这一过程。此外，我们将测试一个特定的模型，研究粘附素如何参与抑制减数分裂间DSB修复。减数分裂DSB的修复利用两种重组酶进行链侵袭：Dmc1，它是减数分裂特有的；RAD51，营养细胞中的主要重组酶。这两个链交换蛋白在减数分裂重组过程中如何协调作用尚不清楚。最近的生化实验表明了一种新的机制，通过辅助蛋白Rad54的MEK1磷酸化来调节RAD51的活性。一个目标是确定这种机制是否在减数分裂细胞中起作用。 公共卫生相关性：减数分裂在进化上保守的细胞分裂失败会导致不孕不育和出生缺陷，如唐氏综合症。适当的减数分裂染色体分离需要双链断裂修复--这一过程也是维持基因组完整性和预防癌症所必需的。这笔赠款的重点是了解减数分裂过程中正确的染色体行为的分子基础--这些知识最终可能导致诊断和/或预防某些类型的不孕不育、出生缺陷甚至癌症。

DESCRIPTION (provided by applicant): Meiosis is a specialized cell division used by sexually reproducing organisms to produce haploid gametes from diploid cells. This reduction in chromosome number is accomplished by having two rounds of chromosome segregation follow a single round of chromosome duplication. Failures in meiotic chromosome segregation in humans lead to infertility and birth defects such as Trisomy 21, the leading cause of mental retardation in the United States. In addition, meiotic chromosome segregation requires the repair of programmed double strand breaks (DSBs). DSB repair in mitotically dividing cells is critical for genome integrity and the prevention of cancer. Budding yeast is an excellent model system for studying meiosis. Mice and yeast mutated for orthologous meiotic genes are phenotypically similar, demonstrating that studies of meiosis in yeast are likely to be illuminating with regard to meiosis in humans. The first meiotic division (MI) is unique in that sister chromatids segregate to the same pole (called reductional segregation). Proper MI segregation requires that homologous chromosomes be connected to each other by crossovers between non-sister chromatids in combination with sister chromatid cohesion. Recent work has shown that crossing over between non-sister chromatids during meiosis is promoted in part by active suppression of DSB repair between sister chromatids. This suppression is mediated by Mek1, a meiosis-specific serine/threonine kinase. Understanding the molecular mechanism by which Mek1 inhibits meiotic sister chromatid repair requires identification of the substrates phosphorylated by this kinase. We will use novel biochemical and genetic strategies to identify Mek1 targets and characterize phosphorylation- defective versions of these proteins elucidate this process. In addition we will test a specific model for how cohesins may be involved in suppressing meiotic intersister DSB repair. Repair of meiotic DSBs utilizes two recombinases for strand invasion: Dmc1, which is meiosis-specific and Rad51, the major recombinase in vegetative cells. How the action of these two strand exchange proteins is coordinated during meiotic recombination not yet understood. Recent biochemical experiments have suggested a novel mechanism for regulating Rad51 activity mediated by Mek1 phosphorylation of an accessory protein, Rad54. One goal is to determine whether this mechanism functions in meiotic cells. PUBLIC HEALTH RELEVANCE: Failures in the evolutionarily conserved cell division of meiosis result in infertility and birth defects such as Down syndrome. Proper meiotic chromosome segregation requires double strand break repair-a process also required to maintain genome integrity and prevent cancer. This grant is focused on understanding the molecular basis for proper chromosome behavior during meiosis-knowledge that may ultimately lead to the diagnosis and/or prevention of certain types of infertility, birth defects and even cancer.