Homolog bi-orientation and segregation in oocyte acentrosomal meiosis

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During the first meiotic division, homologous chromosomes linked by chiasmata attach to microtubules from opposite spindle poles (bi-orientation) and then segregate. In humans, errors in chromosome segregation in the oocyte lead to aneuploidy and are the leading cause of miscarriage, infertility and birth defects. Our long-term goal is to understand the mechanisms that promote accurate chromosome segregation, and the features of the oocyte spindle that make it susceptible to chromosome segregation errors. Our previous research using Drosophila melanogaster females has led to a model in which two types of microtubule attachment are used for bi-orientation. Lateral attachments, where the kinetochores interact with the sides of microtubules, establish bi-orientation. Then end-on attachments, where the kinetochores attach to the end of microtubules, maintain and segregate bi-orientated homologs. A prominent feature of the Drosophila oocyte is the metaphase I central spindle, which functions as a “backbone”, organizing the microtubules into a bipolar structure in the absence of centrosomes. Our work has shown that the central spindle has an important role in bi-orientation during pro-metaphase. Studies in C. elegans and mouse oocytes indicate that the metaphase central spindle may be a conserved element required for the bi-orientation of homologous chromosomes during acentrosomal meiosis. In the previous funding period, we developed several tools to study the mechanisms of bi- orientation in oocytes. These tools include RNAi resistant transgenes in order to make germline-specific mutants of key proteins. Furthermore, we have the reagents, either transgenes or antibodies, to detect many of the important proteins that regulate chromosome segregation, including centromere, kinetochore, checkpoint and spindle proteins. With these tools, we will investigate the mechanisms by which the central spindle interacts with the kinetochores to promote bi-orientation. It is likely that premature stabilization of end-on attachments leads to bi-orientation defects. Therefore, we will investigate the mechanisms of lateral attachments and bi-orientation, and how the transition to end-on attachments is regulated. These studies will focus on two kinetochore proteins, CENP-C and SPC105R, which are required to load several other kinetochore and checkpoint proteins. We will also investigate how the central spindle interacts with the kinetochores and promotes accurate bi-orientation. These studies will include experiments to model in Drosophila, central spindle mutations that decrease fertility in human females. The Aims of this proposal are linked by a goal to understand the mechanisms of chromosome segregation important to oocytes. In completing this work, we will have gained insights into how kinetochores regulate the transition from lateral and end-on microtubule attachment.