Our lab is interested in understanding the mechanisms of ATP-dependent chromatin remodeling, the characteristic features of remodeled chromatin and the impact of different remodeling mechanisms on biological processes and disease.
To understand how ATP-dependent chromatin remodelers regulate biological processes such as transcription, we are defining chromatin states inside cells before and after a remodeling event, and assessing the effects of these changes on gene expression. We have adapted a microarray approach to map nucleosome structure in human cell lines using tiled genomic DNA arrays. This approach improves upon classic methods, which are limited in resolution and the size of regions that can be analyzed(Figure to the left).
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Furthermore, a Cot-enrichment step is incorporated into our protocol to remove repetitive DNA; thus reducing the complexity of mammalian genomes and increasing signals from signal copy genes. As an experimental model, we are examining chromatin structure remodeled by BRG1 and BSB, a chimeric remodeler that has a SNF2h-lke activity but is incorporated into BRG1-containing complexes. These two remodelers create different remodeled products in vitro but are targeted to the same genomic loci (Figure to the right). We are comparing BRG1 and BSB-created nucleosome structures over 100 Mbs of human DNA at high resolution. Based upon our in vitro studies of the differences in remodeling activities, this level of resolution is necessary to understand the biological importance of different remodeling mechanisms. Top
Investigate the Role of Chromatin Remodeling in Transcription-Coupled DNA Repair and Cockayne Syndrome
The Cockayne syndrome B (CSB) protein is an ATP-dependent chromatin remodeler that plays a pivotal role in transcription-coupled DNA repair and transcription regulation in general. Mutations in CSB lead to Cockayne syndrome, an inherited disorder in which patients have neurological and developmental defects, are sensitive to sunlight, and suffer from premature aging. How defects in CSB activity lead to this diverse constellation of clinical features remains largely unknown. We are using CSB as an experimental paradigm to elucidate the relationship between the biochemical activities and biological functions of ATP-dependent chromatin remodeling complexes. By defining the activities of this enzymes and using disease-associated mutations that disrupt specific activities, we will be able to determine how CSB is utilized in different steps of transcription-coupled DNA repair. Consequently, we will gain greater insights into the mechanisms by which chromatin remodelers function in the maintenance of genome integrity, and elucidate the contribution of their activities to normal development and disease.