Functional and structural studies of the transition from transcription initiation to elongation

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We extend our structural mechanistic studies into eukaryotic transcription to determine the functions of the conserved and essential Mediator complex during assembly and disassembly of the RNA polymerase II (pol II) transcription pre-initiation complex (PIC) and in the divergent transcription that arises from pol II promoters. Building on our recent success in developing highly efficient in vitro reconstituted transcription system, we will leverage this system to isolate and dissect a series of PIC-Mediator complexes representing a range of states, from initiation, to early elongation, to complexes that might function on divergent promoters. We will address the molecular mechanisms underlying transitions between these states through structural and biochemical means involving cryo- EM, cross-linking and mass spectrometry (XL-MS), and multi-wavelength analytical ultracentrifugation.