Naomi Goldman

"Exploring the mechanisms that establish the chromatin state of T cells"

Cell fate-specific gene expression programs are established in part by alterations in chromatin accessibility via the action of lineage-determining transcription factors (TFs). Work in our lab has recently identified that the transcription factor TCF-1—integral for normal thymic development—targets and is essential for the opening of repressed chromatin in T cells1. In other cell-types, LDTFs are known to collaborate to organize a fully active enhancer2,3. However, the mechanism through which TCF-1 acts at enhancers in order to regulate genes during T cell development is unknown. To characterize the role of TCF-1 in the establishment of the T cell epigenome, I immunoprecipitated TCF-1 in thymocytes followed by a mass spectrometry (MS) analysis to identify interacting proteins. The proteins that were enriched in the TCF-1 IP relative to a IgG pull down included multiple subunits of the SWI/SNF chromatin remodeling complex. In order to validate this finding I performed Brg1(the ATPase subunit of the swi/snf complex) ChIP-seq in thymocytes and found pervasive binding of this protein at key T cell loci. Previous work has shown that exogenous expression of TCF-1 in NIH3T3 fibroblasts leads to a gain in chromatin accessibility at T cell regulatory regions that are normally silent in fibroblasts. Given this observation, I have developed a system to determine whether these epigenetic changes are dependent on SWI/SNF or other chromatin remodeling proteins by utilizing CRISPR/cas9 to knock down remodelers in fibroblasts prior to TCF-1 transduction followed by RNAseq and ATAC-seq to assess gene expression and chromatin state. Additionally, I have delineated changes in Swi/Snf binding after TCF-1 transduction suggesting this protein is recruited away from endogenous fibroblast loci to sites that gain accessibility after TCF-1 expression. These studies aim to provide mechanistic insight into how transcription factors work to establish cell identity in addition to adding to our understanding of how cell fate might be reprogramed at will for therapeutic purposes.