Meiotic recombination in budding yeast

有性生殖的基础是制造配子的能力，如精子和卵子。 人类，每个染色体只有一个拷贝。受精的结果是两个 单倍体配子来创造二倍体生物体。对于芽殖酵母和人类等生物体来说，这是一种 这是一项艰巨的任务，因为分别有16对和23对染色体，必须正确地分类成 每个配子减数分裂是一种特殊的细胞分裂，它通过使染色体数目减半， 一轮DNA复制后两轮染色体分离。未能正确 在减数分裂期间分离的染色体产生染色体不平衡的配子，导致不育 和出生缺陷，如21三体（唐氏综合症）。 减数分裂的一个关键部分是第一次减数分裂，同源染色体分离 到主轴的相反两极。交叉是由DNA的相互交换产生的， 同系物。交叉，结合姐妹染色单体的凝聚力，物理连接同源物， 可以在第一次减数分裂时对齐并正确分离。交叉是由双链修复引起的， 有意引入染色体以启动重组的断裂。因为未修复的双 链断裂是致命的，减数分裂重组是一个高度调控的过程，确保每对染色体都能在减数分裂中发生。 同源物接受至少一次交换，并且所有双链断裂在第一次减数分裂前被修复 师. 直接在哺乳动物中研究减数分裂是困难的，因为很难接近生殖细胞，而且细胞是 二倍体，这使得寻找隐性突变具有挑战性。芽殖酵母酿酒酵母 由于复杂的遗传，生化， 可用的分子和细胞生物学工具。我研究的重点是 减数分裂重组所需的，并确定这些基因功能的分子机制 和/或被调节。特别是，我的实验室已经开发了新的方法来研究磷酸化如何 调节减数分裂过程中的重组，重点是减数分裂特异性激酶Mek 1和 保守的细胞周期激酶Cdc 7-Dbf 4。 虽然我们对减数分裂重组的理解已经取得了巨大的进展，但仍然存在一些问题。 仍然是需要填补的关键空白。例如，有许多基因有助于 减数分裂双链断裂修复，仍有待发现。在接下来的五年里，我的实验室计划 研究了我们已经确定的两个基因，它们以前不知道在减数分裂中起作用：SEN 1和RRM 3。 Sen 1是一种解旋酶，在有丝分裂的细胞中解旋RNA/DNA杂合体（称为R环），是必不可少的 终身它的哺乳动物直系同源物Senataxin是减数分裂所必需的。Rrm 3是保守的Pif 1的成员 DNA解旋酶家族，以其在DNA复制中的作用而闻名。此外，我们还发现了一个 另外两种DNA解旋酶Sgs 1和Srs 2可能在调节 交叉编队 在许多生物体如酵母和哺乳动物中的减数分裂需要两种重组酶Rad 51， 对于有丝分裂重组和减数分裂特异性Dmc 1是必需的，Dmc 1介导大部分同源物之间的相互作用。 重组一个突出的问题是为什么需要两个重组酶。我的实验室和其他人的工作 Dmc 1已经进化到更好地处理可能出现的错配碱基对， 同源物间链侵入，因为同源物具有高度相似但不一定相同的DNA 序列的我的实验室已经开发了一种体内方法来测试这个有趣的假设。工作支持 这项资助将对我们理解减数分裂做出重要贡献， 最终可应用于人类以预防/治疗不孕症和出生缺陷。

Fundamental to sexual reproduction is the ability to make gametes, such as sperm and eggs in humans, which contain only one copy of each chromosome. Fertilization then results in the fusion of two haploid gametes to create a diploid organism. For organisms such as budding yeast and humans, this is a daunting task, as there are 16 and 23 pairs of chromosomes, respectively, that must be properly sorted into each gamete. Meiosis is the specialized cell division that divides the chromosome number in half by having one round of DNA replication followed by two rounds of chromosome segregation. Failure to properly segregate chromosomes during meiosis produces chromosomally imbalanced gametes, resulting in infertility and birth defects such as Trisomy 21 (Down Syndrome). A critical part of meiosis is the first meiotic division, where homologous chromosomes are segregated to opposite poles of the spindle. Crossovers are created by the reciprocal exchange of DNA between homologs. Crossovers, in combination with sister chromatid cohesion, physically connect homologs so they can align and properly segregate at the first meiotic division. Crossovers result from the repair of double strand breaks that are deliberately introduced into chromosomes to initiate recombination. Because unrepaired double strand breaks are lethal, meiotic recombination is a highly regulated process that ensures that every pair of homologs receives at least one crossover and that all double strand breaks are repaired before the first meiotic division. Studying meiosis directly in mammals is difficult as it is hard to access germ cells and the cells are diploid, making it challenging to find recessive mutations. The budding yeast, Saccharomyces cerevisiae, has been an excellent model system for studying meiosis because of the sophisticated genetic, biochemical, molecular and cell biological tools that are available. The focus of my research has been on identifying genes required for meiotic recombination and defining the molecular mechanisms by which these genes function and/or are regulated. In particular, my lab has developed novel approaches for studying how phosphorylation regulates recombination during meiosis, with an emphasis on the meiosis-specific kinase, Mek1 and the conserved cell cycle kinase, Cdc7-Dbf4. Although immense progress has been made in our understanding of meiotic recombination, there are still critical gaps that need to be filled. For example, there are many genes that contribute to the fidelity of meiotic double strand break repair which remain to be discovered. Over the next five years, my lab plans to study two genes we have identified that were previously unknown to play a role in meiosis: SEN1 and RRM3. Sen1 is a helicase that unwinds RNA/DNA hybrids called R-loops in mitotically dividing cells and is essential for life. Its mammalian ortholog, Senataxin, is required for meiosis. Rrm3 is a member of the conserved Pif1 DNA helicase family that is well known for its role in DNA replication. In addition, we have discovered a potentially novel role for two other DNA helicases, Sgs1 and Srs2, working together in the regulation of crossover formation. Meiosis in many organisms such as yeast and mammals requires two recombinases, Rad51, which is essential for mitotic recombination and the meiosis-specific Dmc1, which mediates the bulk of interhomolog recombination. An outstanding question is why two recombinases are necessary. Work from my lab and others has suggested that Dmc1 has evolved to better handle the mismatched basepairs that can arise by interhomolog strand invasion because homologs have highly similar, but not necessarily identical DNA sequences. My lab has developed an in vivo approach to test this interesting hypothesis. The work supported by this grant will make an important contribution to our understanding of meiosis, knowledge which may ultimately be applicable in humans for preventing/treating infertility and birth defects.