HETEROCHROMATIN REGULATION BY IMITATION SWITCH COMPLEXES

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Gene expression in eukaryotes is modulated by positional information and higher order repressive chromatin structure (heterochromatin). Chromatin remodeling complexes have been implicated in the disruption of such repressive chromatin resulting in modulation of transcription, DNA replication, and DNA repair. Mutations in the components of these chromatin modifying complexes leads to severe developmental defects (e.g. ATRx, MeCP2) and cancer development (e.g. SNF5, CBP). We are focused on chromatin remodeling complexes containing the human chromatin-remodeling factor Imitation Switch (ISWI) protein.We hypothesize that ISWI-containing complexes facilitate gene expression by altering the structure of a higher order chromatin structure, known as heterochromatin. The three Aims are designed to provide a thorough biochemical and functional characterization of human ISWI-containing complexes. Aim 1 will identify the remaining subunits of the 1-1.5 MDa ISWI/WSTF- containing complex and describe experiments that will begin to define the chromatin remodeling activity of the complex and the individual proteins in the complex. Aim 2 describes a detailed functional analysis of human ISWI-complexes with emphasis on their putative role in heterochromatin modulation. Aim 3 extends the work to tissue-specific chromatin remodeling complexes containing human SNF2L and use a cell line expressing FLAG-tagged SNF2L protein to identify and functionally characterize the SNF2L chromatin remodeling complex.