Role of Higher Order Chromatin Structure in Gamete Formation

Checchi, Paula

Abstract

Meiosis ensures the faithful segregation of genetic material to sperm and egg and is necessary for the propagation of all sexually reproducing species. Paradoxically, a requirement for this is the deliberate induction of genome-wide double-strand breaks (DSBs), which in other contexts are considered highly toxic. As a result, all organisms are equipped with an array of highly-conserved DSB repair mechanisms as well as checkpoint pathways. While it is mechanistically understood how repair pathways and checkpoints respond to meiotic DSBs, the interplay between this complex network of pathways is poorly defined. This gap in our knowledge is a significant barrier for human health, as perturbations in DSB repair cause fertility problems, birth defects, and can lead to cancer. Our long-term goal is to identify the regulatory mechanisms that control meiotic chromosome dynamics and prevent errors from being transmitted to offspring. The primary objective of this application is to determine how regulation of higher order chromatin structure coordinates timely and accurate repair of meiotic DSBs. To approach this problem, we will take advantage of the structural organization of the C. elegans germ line. Combined with the exceptional genetics and ease of obtaining mutant and transgenic strains, this established model is ideal for understanding how chromatin structure maintains the cellular integrity of germ cells during meiosis and prevents errors from being transmitted to offspring. The overarching hypothesis is that modifiers of higher order chromatin structure promote gamete quality by regulating meiotic DSB repair. The rationale behind this research is that our findings will produce a detailed understanding of how several regulatory complexes important for higher-order chromatin structure function during meiosis to ensure the production of gametes with the correct chromosome complement and an absence of DNA lesions. Guided by strong preliminary data, we will test our hypothesis by pursuing two major aims.
In Aim1, we will determine how the structural maintenance of chromosomes SMC-5/6 complex prevents defects in meiotic recombination that compromise oogenesis to a far greater degree than spermatogenesis. In this aim, we will use a combination of genetic, molecular and cytological assays which will identify the consequences of defective DSB repair in offspring and will also determine the nature of several sexually dimorphic aspects of SMC-5/6 activity that we recently discovered. Our experiments in Aim 2 will uncover the requirement for an ATP-dependent-chromatin remodeling complex, NuRD, in maintaining germ cell survival. To accomplish this, we will use genetic, cytological and biochemical approaches to determine its functional role in the context of meiotic recombination. The work outlined in this proposal will produce a detailed understanding of how several critical regulators of higher-order chromatin structure accommodate the timely and efficient repair of meiotic DSBs, which given the evolutionary conservation of all players involved, will provide significant insight as to how dysregulation of these processes manifests in human disease states.

Public Health Relevance

The proposed research is relevant to the general medical sciences because it will provide insight into the underlying causes of human reproduction problems and the significantly increased risk of chromosome disorders that arise from female meiosis. Thus, the proposed research is relevant the NIH's mission to increase our understanding of biological processes and provide the foundation for advances in disease diagnosis, treatment and prevention.