Eukaryotic transcription mechanisms: from nuclear RNA polymerases to general initiation factors, gene‐specific activators, coactivators and chromatin

Abstract

Transcriptional regulation underlies key events in development, differentiation, homeostasis and cell growth and transformation. The last 45 years have revealed an extraordinary complexity to the eukaryotic transcription machinery, far beyond that evident in prokaryotes, as well as diverse regulatory mechanisms. Our own biochemical studies over this period led to the discovery of (i) three nuclear RNA polymerases (Pols I, II and III) with distinct subunit compositions and functions, (ii) cognate general initiation factors that are essential for RNA polymerase recruitment and initiation, (iii) the first gene‐specific transcriptional activator (and prototype zinc finger protein) in eukaryotes, (iv) diverse general and cell‐specific coactivators and (v) underlying mechanisms of action1. Notably, our early demonstration of robust, accurate in vitro transcription of cell‐specific genes (assayed as DNA templates) by Pol II and corresponding (ubiquitous) general initiation factors also led to the prediction of a general repression mechanism, later identified primarily as assembly into chromatin, and gene/tissue‐specific factors, many examples of which now exist, that could selectively reverse this repression. Our more recent studies have emphasized mechanistic analyses of transcriptional coactivators in biochemically defined cell‐free systems reconstituted with purified factors and either DNA templates (to analyze coactivators, such as the 30‐subunit Mediator and the TAF subunits of TFIID, that directly link activators to Pol II and cognate initiation factors) or with recombinant chromatin templates (to analyze the functions of histone modifying factors that act indirectly on the transcription machinery). Among other significant findings, these studies have (i) provided important insights into mechanisms underlying Mediator recruitment (interactions with enhancer‐bound activators) and subsequent functions, (ii) provided the first evidence that specific histone modifications are causal for (rather than simply correlated with) transcriptional activation, (iii) established direct synergistic functions of diverse histone modifying factors, through new mechanisms, in transcription and (iv) established a novel mechanism for activation of higher order (condensed) linker histone H1‐containing chromatin. Selected aspects of this work will be discussed.