Molecular Basis of Centromere Specification and Inheritance

New York University, New York NY

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Abstract

PROJECT SUMMARY/ABSTRACT A fundamental but poorly understood process in eukaryotic cells is how cells structure their genomes into distinct functional domains. This project addresses this gap in knowledge by studying the centromere, a specific chromatin domain found in all eukaryotes. This stably propagated locus guides the assembly of kinetochores to ensure proper segregation of chromosomes during mitosis and meiosis. Mis-regulation of centromeres adversely affects chromosome segregation resulting in aneuploidy, a condition found in more than 90% of all cancers. Aneuploidy contributes to the development of many diseases, such as cancer and Down syndrome. The goal of this project is to understand the molecular mechanisms underlying the specification and inheritance of centromeres. In most eukaryotes, centromeres are epigenetically governed by the centromere-specific histone H3 variant, CENP-A. CENP-A partially replaces canonical histone H3 at centromeres, and provides the foundation for the assembly of kinetochores. Mislocalization of CENP-A to non-centromeric regions has a devastating impact on chromosome segregation, and has been linked to a variety of cancers. However, how CENP-A chromatin is assembled at centromeres remains poorly understood. Centromeres in most eukaryotes can be transcribed at low level, and centromere transcription has been implicated in centromere structure and function. But how centromere transcription is regulated is still largely unexplored. Interestingly, centromeres in eukaryotes are usually embedded in epigenetically distinct heterochromatin, the transcriptionally silenced chromatin domain. Heterochromatin is mainly composed of tandem DNA repeats, and has been shown to contribute to centromere formation. Nonetheless, how heterochromatin repeats are organized and the precise role of heterochromatin in centromere assembly remains elusive. In addition, centromeres and heterochromatin domains in the eukaryotic nucleus typically exhibit distinct patterns of spatial organization, which have been implicated in 3D genome architecture and regulation. But how centromeres and heterochromatin are spatially organized remains unclear. We propose to use fission yeast (Schizosaccharomyces pombe) to address these outstanding questions. Fission yeast is a simple eukaryotic model organism with many aspects of centromere regulation conserved with humans. It is particularly suited to an interdisciplinary approach that includes genetics, genomics, cytology, biochemistry, and structural biology. We propose to: 1) define the mechanisms for how centromere transcription is regulated and its role in centromere function; 2) determine how the heterochromatin repeats are organized and their contribution to CENP-A chromatin assembly; 3) identify regulatory mechanisms underlying spatial organization of centromeres and heterochromatin. Given that epigenetic regulation in fission yeast is conserved, our studies will shed light on the processes governing chromosome segregation in human cells, and contribute to a better understanding of human diseases resulting from centromere misregulation.