Protein characterization of Saccharomyces cerevisiae RNA polymerase II after in vivo cross-linking.

ABSTRACT: To characterize proteins associated with active transcription complexes, we purified RNA polymerase II (pol II) from Saccharomyces cerevisiae after fixing live cells with formaldehyde. The approach mimics ChIP and requires solubilizing cross-linked complexes with sonication. Pol II was affinity-purified, and associated proteins were identified by MS. Several classes of proteins depended on cross-linking, including Mediator, general transcription factors, elongation factors, ribonucleoprotein particle (RNP) proteins, and histones. A tagged RNP protein reciprocally purified pol II under identical cross-linking conditions, and the association between RNP proteins and pol II was largely RNase-sensitive. The data indicate that the cross-linked Pol II purification contains elongating pol II with

SUBMITTER: Tardiff DF 

PROVIDER: S-EPMC2148403 | biostudies-literature | 2007 Dec

REPOSITORIES: biostudies-literature