Factor-stimulated RNA polymerase II transcribes at physiological elongation rates on naked DNA but very poorly on chromatin templates.

Abstract

We used an in vitro assay system based on HeLa cell core transcription components to examine transcript elongation by RNA polymerase II on either naked DNA or chromatin templates as a function of the three known elongation factors, IIS, TFIIF, and TFIIX. We demonstrate for the first time that mammalian RNA polymerase II can achieve physiological elongation rates on naked DNA templates in vitro. The addition of TFIIF alone gave this rate, although IIS was required to minimize the block to elongation at intrinsic termination sites. However, IIS and TFIIF provided only a slight increase in the very poor elongation efficiency of RNA polymerase II on chromatin templates. The addition of TFIIX to reactions containing IIS and TFIIF reduced the elongation rate on naked DNA templates but slightly increased the elongation efficiency on chromatin. The ability of elongation factors either separately or in combination to stimulate transcription on naked DNA and chromatin templates was also examined.