Molecular Mechanisms of Meiotic Chromosome Assembly, Architecture, and Interhomolog Recombination

Meiosis, a hallmark of sexual reproduction, entails the division of one diploid parent cell into four haploid gamete daughter cells. To support proper chromosome segregation in two successive meiotic divisions, meiotic chromosomes must undergo dramatic structural changes in meiotic prophase to identify and pair with their corresponding homolog. This chromosomal structural rearrangement is accomplished through meiotic recombination, a process that inflicts double strand break damage on the chromosomes and necessitates their repair using their homologous sequence, in some cases forming an inter-homolog crossover. This work begins by reviewing meiotic chromosome structure, and how it supports meiotic recombination. In order to understand the structural context of meiotic recombination I begin by reviewing higher-order data on how entire chromosomes are packaged and ordered during meiosis then zoom into well-known structures such as the chromosome axis, the synaptonemal complex, and their individual protein components. The chapters on my experimental work begin by examining a Chromatin Binding Region (CBR) in the yeast meiotic HORMAD Hop1, a component of the chromosome axis. I demonstrate how the CBR is responsible for a secondary mode of chromosome axis deposition through its capability to bind directly nucleosomes. This secondary mechanism functions alongside a previously defined cohesin dependent chromosome axis assembly mechanism, and I hypothesize that this second mode of axis assembly enables crossover formation in an as-yet unknown manner. I next examine the Msh4-Msh5 complex in an attempt to define its role in shuttling double strand DNA breaks towards a crossover fate. Through careful biochemistry, I demonstrate that the preferred DNA substrate of the S. cerevisiae Msh4-Msh5 is a Holliday junction, and I identify the critical regions in both Msh4 and Msh5 for this activity. My last study aims to examine meiotic recombination from a larger perspective. In order to determine how individual proteins interplay on the chromatin throughout the process of meiotic prophase I, I developed a mass spectroscopy protocol to assay a meiotic time-course of S. cerevisiae chromosomes. With this protocol I created a spatial and temporal atlas of chromatin associated proteins, which details how proteins assemble and disassemble throughout meiotic prophase I. As a whole this thesis provides an extensive review of the field of meiotic chromosome architecture, defines molecular mechanisms of individual protein assembly during meiosis, and demonstrates the proteomic context for interhomolog recombination