Our lab is interested in understanding the mechanisms of ATP-dependent chromatin remodeling, the characteristic features of remodeled products and the impact of different remodeling mechanisms on biological processes. The information gained from these studies will not only reveal how ATP-dependent chromatin remodelers regulate diverse nuclear processes, but will shed light on the underlying causes of disease and the principles of epigenetic regulation.
To understand how ATP-dependent chromatin remodelers regulate biological processes such as transcription, we are interested in defining chromatin states inside cells before and after a remodeling event, and assessing their effects 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 size of the region that can be analyzed, about 1-2 Kbs, and in resolution (Figure 1). Furthermore, a Cot-enrichment step is incorporated into our protocol to remove repetitive DNA and reduce the complexity of mammalian genomes, and thus, increase the signals from signal copy genes. As an experimental model, we are using two remodelers, BRG1 and BSB, which create different remodeled products in vitro but are targeted to the same genomic loci (Figure 2). 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.
Cockayne syndrome is an inherited disorder in which patients are highly sensitive to sunlight, have neurological and developmental defects, and suffer from premature aging. Mutations in the CSB protein lead to Cockayne syndrome; however, how defects in this protein lead to this diverse constellation of clinical features remains largely unknown. The CSB protein is an ATP-dependent chromatin remodeler and plays a pivotal role in transcription-coupled DNA repair. 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. We are addressing such questions as: What is the biochemical mechanism that CSB uses to alter chromatin structure? How is CSB remodeling activity integrated into the biological process of transcription-coupled DNA repair? How do auxiliary proteins modulate CSB activity? These studies will unveil the mechanistic bases for the genetic defects associated with Cockayne syndrome and give us a broader understanding of how ATP-dependent remodelers regulate diverse biological processes.