Chromatin was first defined by Flemming in 1882 as the 'substance in the cell nucleus which is readily stained' that is retractile to digestion'. The term stuck, and now is taken to refer to DNA (the readily stained component) and the associated proteins that make it retractile to digestion. Over the past two decades, it has become apparent that the associated proteins play a key role in regulating transcription, replication and recombination of the DNA. The basic structural component of chromatin, the nucleosome, which was treated for years as a 'dumb' building block, has been shown to be dynamic and regulated at several levels. Nucleosomes can be evicted or shifted in position by ATP-dependent remodeling proteins, opening up DNA sequences. They can be covalently modified in different ways, allowing differential abilities to interact with the regulatory machinery. The purpose of the experiments described in this application is to characterize regulatory proteins that modulate chromatin structure to effect gene regulation. The three Aims examine three separate complexes involved in these processes, all of which are human homologs of proteins originally identified in screens done in Drosophila for mutations that impact developmental progression. These screens identified the Polycomb-Group (PcG) of genes, responsible for maintaining repression of master regulatory genes during development, and the trithorax-Group (trxG), isolated as suppressor of a Polycomb mutant.
Aims 1 and 2 examine two mammalian ATP-dependent remodeling proteins in the trxG family and Aim 3 examines a mammalian protein in the PcG family.
Aim 1 will determine the crystal structure of the ATPase domain of BRG1;
Aim 2 will examine the impact of human disease mutations of function of the CHD7-ATP-dependent
The CHD7 gene is mutated in 90% of human patients that have CHARGE syndrome, a serious developmental defect that impact 1 in 10,000 live births. EZH2 has been implicated in numerous human malignancies and is a key therapeutic target in cancer treatment. BRGI and other members ofthe human SWI/SNF family complexes are also strongly implicated in numerous cancers.
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