Pol III-Related Mechanism of Transcription Through Chromatin

Rutgers, The State University Of New Jersey-Rbhs-Robert Wood, Piscataway NJ

Investigators

Abstract

The genome of higher organisms is organized in DNA-protein complexes called chromatin. Chromatin is designed to allow tight DNA packaging and proper functioning of DNA, as well as regulation of enzymes such as nuclear DNA and RNA polymerases. RNA polymerases transcribe chromatin (make RNA copies of one DNA strand). Transcribed genes are organized in chromatin raising the question of how RNA polymerases deal with chromatin. The primary features of this process have been recapitulated in vitro. In particular, the mechanism used by eukaryotic RNA polymerase III (pol III) is accompanied by transient chromatin remodeling at the sites of transcription followed by recovery of chromatin structure. This project focuses on further mechanistic analysis of this mechanism and its regulation using defined chromatin templates. The mechanism of transcription through chromatin by pol III will be analyzed using biochemical approaches, electron cryo-microscopy and fluorescence resonance energy transfer (FRET). Completion of this project will result in development of novel defined chromatin templates and new techniques useful for analysis of various DNA-protein interactions in chromatin, including interaction of specific regulatory factors and enzymes of DNA metabolism (such as enzymes involved in DNA replication and repair) with chromatin. The data obtained will lead to better understanding of the mechanism of chromatin transcription in terms of the structures of the DNA-protein complexes formed during this process. Since the mechanisms of DNA replication, ATP-dependent chromatin remodeling and the mechanism of transcription through chromatin by pol III have many common features, the results obtained during this research will help to understand all these processes.