The mammalian oocyte is extremely susceptible to chromosome segregation errors leading to aneuploidy. As the single most common cause of congenital birth defects and pregnancy loss, aneuploidy is an important clinical condition with a significant impact on human health. Meiotic centromeres are epigenetically determined chromosomal domains that exhibit unique structural and functional properties for accurate chromosome segregation. Little is known concerning the epigenetic control of centromere function. We provide novel evidence that the chromatin remodeling protein LSH exhibits a germ cell-specific function in centromere cytosine methylation and silencing of centromeric non-coding RNAs. Notably, LSH is required to establish critical epigenetic marks for inner centromere and kinetochore formation. We test the hypothesis that LSH provides a molecular link between the epigenetic control of centromere function, kinetochore assembly and chromosome segregation during female meiosis.
Aim 1. Using a conditional deletion in fetal oocytes and RNA sequencing, we will test the hypothesis that LSH regulates the expression of repetitive elements such as centromeric transcripts and IAP transposons to protect the integrity of the oocyte genome and will establish direct targets of LSH epigenetic silencing during meiotic prophase-I.
Aim 2 will determine the mechanisms by which loss of LSH function disrupts DNA methylation at repetitive elements. Using a genome-wide single base resolution DNA methylome analysis, we test the hypothesis that LSH is required to recruit the DNA methylation machinery for the epigenetic silencing of transposons in female germ cells.
Aim 3. Using an LSH conditional deletion in preovulatory oocytes, transcription activator-like effectors for epigenome editing and chromatin accessibility assays, we will determine the role of LSH in regulating kinetochore chromatin determinants for accurate chromosome segregation and genome wide nucleosome organization. We test the hypothesis that LSH is a novel component of the kinetochore that is required to integrate histone H3 modifications with chromosome segregation and timely anaphase-I onset. This proposal shifts the existing paradigm of LSH function by revealing a previously unrecognized, germ cell specific role in the epigenetic control of centromere function and chromosome segregation. Outcome: This proposal addresses for the first time the epigenetic mechanisms regulating centromere organization during meiosis and will contribute to determine how the chromatin that defines the inner centromere and kinetochore domains is epigenetically regulated. Impact: More than 40% of DNA sequences in the mouse genome are derived from repetitive elements, yet the mechanisms regulating deleterious transposons during female meiosis remain to be established. Abnormal centromere function is a major cause of human oocyte aneuploidy. These studies will provide molecular insight into the underlying mechanisms of centromere instability, chromosomal non-disjunction and the epigenetic origins of oocyte aneuploidy. !
Public Health Relevance:Abnormal chromosome segregation during meiotic division leads to aneuploidy, the single most important cause of congenital birth defects and pregnancy loss in humans. The mechanisms involved in this process are not known. The proposed studies address a previously unrecognized role of chromatin remodeling proteins in the epigenetic and molecular origins of chromosomal non-disjunction, and have important implications for the identification of mechanisms leading to human trisomies, chromosome instability, oocyte aneuploidy and female infertility.
