The long-term goal of this program is to understand the mechanisms of establishment and maintenance of transcriptionally silent chromatin in eukaryotes. The experimentally tractable Saccharomyces cerevisiae will be used to examine how silencers initiate the establishment of silent chromatin, how barriers restrict silent chromatin to limited domains, and how silent chromatin responds to different regulations. A mystery about silencer function is that some silencers act bidirectionally while others unidirectionally. Both the intrinsic property and the genomic context of a silencer play roles in determining its directionality. How silencer- binding proteins organize local chromatin, and how special DMA elements called protosilencers cooperate with the silencer to influence its directionality will be examined. Yeast barrier elements are akin to metazoan insulators that can block the propagation of silent chromatin. How barriers demarcate the silent HM loci will be investigated by examining the protein factors involved and testing proposed mechanisms of their functions. Silent chromatin is dynamic. It is abrogated by a histone acetylase recruited to a nearby site, and enhanced by thermal stress. How these stimuli alter the state of silent chromatin pertaining to the covalent modifications of histones and/or the silencing proteins, the association of silencing proteins with chromatin, and the primary chromatin structure will be investigated. Results from these studies should lead to a better understanding of the formation, maintenance and dynamics of silent chromatin domains in yeast, and shed light on similar processes in other organisms including humans. As silent chromatin is key to epigenetic regulation of gene expression during development and cellular differentiation, a better understanding of its structure and function should provide clues to diseases that arise due to disruptions of this regulation. Moreover, barrier elements can be used to protect transgenes from the repressive effect of silent chromatin in the genome in gene therapy.
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