In eukaryotes, epigenetic events govern diverse processes ranging from gene expression to other aspects of global chromosome architecture, essential for preserving the integrity of the genome. They allow discrete subsets of genetic information to be expressed while others are not, in such a fashion that the various patterns of expression are transmitted during cell division. While patterning in higher eukaryotes is complex, evidence from several model systems suggests that heritable repression of transcription might often be mediated by self-templating chromatin structures. The main goal is to understand the molecular mechanisms responsible for the inheritance of epigenetic states. This information is essential for comprehending development in general and expected to contribute to our understanding of the genetic disorders of human, such as Prader Willi syndrome, Angelman syndrome, Beckwith-Wiedemann syndrome and Wilm's tumor, all diseases in which epigenetic imprinting plays an important role. By using the fission yeast silent mating-type region, the investigator recently showed that the transcriptional and recombinational suppression in this region is regulated by an epigenetic mechanism, involving the assembly of a heterochromatic structure. To understand the molecular basis of how a silenced state is established and propagated during cell division, he proposes to study the mechanism of heterochromatin assembly. Chromatin immunoprecipitation analysis will be used to study the localization of silencing factors, such as a chromodomain-containing Swi6 protein, at the mating-type interval. The possible role of histone acetylation pattern and protein-protein interactions in the targeting of Swi6 will be analyzed. Biochemical analysis will be employed to characterize histone deacetylase homologues, Clr3 and Clr6, which also affect transcriptional repression. Additionally, he will identify critical cis-and trans-acting elements essential for the inheritance of the epigenetic states and explore whether heritability of the repressed state depends upon the nuclear compartmentalization of the mating-type region. Their analysis will also define the stage(s) of the cell cycle where a repressed chromatin state can be assembled. Finally, they plan to characterize recently identified factor affecting silencing and will extend our analysis showing that a mutation in DNA identified factors affecting silencing and will extend their analysis showing that a mutation in DNA polymerase-alpha can affect heritability of a repressed chromatin state. Results from these experiments will lead to comprehensive models about how distinct chromatin domains are established and propagated.
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