The objective is to study the function of the SWI/SNF ATP-dependent chromatin remodeling complex in promoter proximal pausing of RNA polymerase II and determination of transcriptional directionality, key regulatory steps in transcription important in development and diseases. In the last several years, researchers have found that in mammals most genes have RNA polymerase (RNAP) II paused 25-60 nts from the transcription start site and that transcription can resume once the appropriate signal is provided. Regulation of promoter proximal pausing has been uncovered to be as crucial, or perhaps more, of a regulatory step in gene expression than formation of the preinitiation complex. There is currently very little known about connections between RNAPII pausing and ATP-dependent chromatin remodelers such as SWI/SNF, even though the chromatin structure at the promoter region is known to be an important determining factor in RNAPII pausing. Polymerase pausing is also connected to the occurrence of divergent transcription, another recent finding in transcription. Most non-coding RNA comes from divergent or bidirectional transcription in which transcription occurs upstream of the coding region in the anti-sense direction. Similar to RNAP II pausing the directionality of transcription is a developmentally controlled process. Preliminary data shows SWI/SNF?s involvement in the regulation of both of these processes and the goals of this proposal are to delineate the molecular means of SWI/SNF regulation of these relatively new aspects of transcription. SWI/SNF is one of the most frequently mutated epigenetic factors in cancer, occurring in ~20% of all cancers, and mutations in SWI/SNF are the molecular drivers for two neurological disorders, Nicolaides-Baraister and Coffin-Siris syndromes. The SWI/SNF complex is also crucial for both pluripotency and differentiation. As a master gatekeeper of chromatin, it has been assumed to be primarily involved in making DNA accessible for formation of the transcription initiation complex. The premise of this proposal is that the functions of SWI/SNF in regulation of RNAPII pausing and directionality is as vital for its role in development and human disease as promoting formation of the transcription initiation complex. In addition, we will examine the important question of how SWI/SNF regulates the synthesis of eRNAs, short RNAs transcribed at enhancer regions, given the remarkable similarity with which both enhancers and promoters are transcribed, often bi-directionally, and using the same transcription and elongation factors. eRNAs facilitate long-range interactions between promoters and enhancers, and are synthesized prior to the synthesis of its corresponding coding RNAs. This proposal examines how SWI/SNF influences RNAPII pausing and directionality at enhancer regions, which is likely to be important for enhancer activity.
The proposed research is relevant to public health because chromatin remodelers regulate gene expression, DNA replication, and chromosome integrity; have important roles in cell development and alterations of these complexes cause different human diseases. This project is relevant to NIH's mission because it focuses on understanding the function of the SWI/SNF chromatin remodeler in the transcription regulation and RNA polymerase II elongation. We want to address the gap in knowledge of how SWI/SNF regulates promoter proximal pausing and divergent transcription in pluripotency and mammalian development both at promoters and enhancers.
