Pol II waiting in the starting gates: Regulating the transition from transcription initiation into productive elongation

Abstract

Proper regulation of gene expression is essential for the differentiation, development and survival of all cells and organisms. Recent work demonstrates that transcription of many genes, including key developmental and stimulus-responsive genes, is regulated after the initiation step, by pausing of RNA polymerase II during elongation through the promoter-proximal region. Thus, there is great interest in better understanding the events that follow transcription initiation and the ways in which the efficiency of early elongation can be modulated to impact expression of these highly regulated genes. Here we describe our current understanding of the steps involved in the transition from an unstable initially transcribing complex into a highly stable and processive elongation complex. We also discuss the interplay between factors that affect early transcript elongation and the potential physiological consequences for genes that are regulated through transcriptional pausing.

Figure 1. The importance of early elongation in the Pol II transcription cycle

A. Schematic view of Pol II transcription cycle.The “conventional” steps of the cycle are shown in black and early elongation in red, with the open arrow indicating termination of transcription and dissociation of Pol II. The dashed arrow represents the potential of a Pol II molecule to reinitiate transcription on the same DNA template as a result of Pol II recycling. Factors involved in regulating the transition between early and productive elongation are shown in colors, with arcs indicating the stages during which the given factor remains associated with Pol II complex. B. Steps and factors involved in the generation of a productive elongation complex. Pol II is shown in grey, with the CTD domain of its largest subunit protruding from polymerase. DNA is indicated in dark blue and RNA in red. Locally melted DNA within the transcription bubble is shown, including its presumably altered topology during abortive initiation. The status of CTD phosphorylation at each stage is shown in the inset on the right. GTFs are shown in brown. Factors involved in the transition to productive elongation, including NELF, DSIF, TFIIS, and P-TEFb are indicated.

Figure 2. Features of promoter-proximal sequences that contribute to pausing

A. Model for the two-stage mechanism of promoter-proximal Pol II pausing. The vertical axis represents A+T content, with the position of DNA along this axis representing its A+T vs. G+C-richness. Transcription through a region of high A+T content (shown as a peak), induces Pol II pausing (top panel). After the initial pausing, Pol II backtracks until it encounters the region with high G+C content (shown as a valley) that contains the pause button and DPE motifs (middle panel). Transcription factor TFIIS, shown in yellow, mediates cleavage of RNA in the backtracked complex, restoring transcriptional activity to the paused Pol II (bottom panel). We note that TFIIS mediated cleavage is necessary - but not sufficient - to release Pol II into productive elongation. B. Two most abundant GC-rich motifs found on genes that show evidence of paused polymerase, the Pause Button (top) and Downstream Promoter Element (bottom) [11, 96]. Logos depict the level of sequence conservation (shown as bits of information) at each position with respect to the TSS.

Figure 3. Transcription output can be affected through regulation of Pol II recruitment or promoter-proximal pausing

Top. Scheme of an inducible gene, showing that transcription output can be regulated both through Pol II recruitment and pause release. The two critical steps (shown by red arrows) can be regulated independently of each other: Pol II recruitment is enhanced by an Activator (blue), whereas pause release is stimulated by recruitment of P-TEFb (green), which can occur through a number of different mechanisms. Bottom. Simulated scenarios of gene transcription under different conditions. Numbers represent average rates of each step (in molecules/s), as well as the rate of transcript output (transcripts/s). Rate-limiting steps are shown in red.