Pioneer transcription factor binding to sites in chromatin
We are exploring the mechanism of binding of the lineage-specific pioneer transcription factor PU.1 to its sites in chromatin. Lineage-specific transcription factors confer cellular identity and are often pioneer transcription factors, which have been suggested to be capable of binding to nucleosomal sites in chromatin. However, whether pioneer transcription factors bind to their sites on a nucleosomal surface or whether pioneer transcription factor binding displaces nucleosomes has not been investigated. Our previous results suggest that binding of the pioneer transcription factor PU.1 at least partially displaces nucleosomes, and we are investigating whether PU.1 removes nucleosomes by itself, or whether it collaborates with remodelers to create accessible chromatin for other transcription factors.
The role of the SWI/SNF remodelers BAF and PBAF in establishing accessible chromatin at cytokine enhancers
The two SWI/SNF family nucleosome remodelers BAF and PBAF share certain subunits, but each complex also contains unique subunits that have been suggested to play a role in targeting the remodelers to their sites on DNA. However, the roles of individual subunits in remodeler function have not been investigated in vivo. Previous studies have shown that BRG1, the catalytic subunit shared by both complexes plays a role in cellular differentiation, including myeloid and lymphoid differentiation (Choi et al., 2012; Hu et al., 2011; Kowenz-Leutz and Leutz, 1999). Other studies have demonstrated a role of BAF and/or PBAF in induction of pro-inflammatory cytokines (Ramirez-Carrozzi et al., 2009). We are analyzing the roles of BAF and PBAF subunits in macrophage differentiation and cytokine induction using a lentiviral shRNA delivery approach. We are testing the hypothesis that the BAF and PBAF complexes play roles in priming cytokine enhancers and creating accessible chromatin in resting macrophages, as well as in clearing nucleosomes from enhancers upon gene induction.