RNA polymerase backtracking in gene regulation and genome instability

Abstract

RNA polymerase is a ratchet machine that oscillates between productive and backtracked states at numerous DNA positions. Since its first description 15 years ago, backtracking--the reversible sliding of RNA polymerase along DNA and RNA--has been implicated in many critical processes in bacteria and eukaryotes, including the control of transcription elongation, pausing, termination, fidelity, and genome instability.

Figure 1. Multifaceted role of RNAP backtracking in the cell

Schematics depict the ternary elongation complex (EC) in active and backtracked configurations. The catalytic site (star) looses the 3′-OH end of RNA (red), which is extruded through the secondary channel during backtracking.

Figure 2. RNAP backtracking and genome instability

Schematics show a model of DSB formation as a result of co-directional collisions between the replisome and backtracked RNAP in bacteria (Dutta et al., 2011). The pink arrow indicates a single strand break (SSB) due to replisome switching from the leading DNA strand (blue) to the RNA (red). The latter forms a stable R-loop upon displacement of the backtracked EC. Transcript cleavage factor (Gre) in the secondary channel and trailing active ribosome prevent RNAP backtracking and R-loop formation, thus preserving genome integrity.

Figure 3. Structural basis of RNAP backtracking

Views of nucleic acids in the backtracked (3PO2) and active (1Y1W) RNAP II structures. Non-template DNA = blue cartoon, template DNA = black cartoon, RNA = red cartoon, RNA extruded past the active site = hotpink cartoon, Mg(I) = magenta sphere, catalytic site residues = yellow sticks, trigger loop = orange cartoon. The RNA:DNA hybrid is partially unwound and tilted in the backtracked EC, whereas the trigger loop is stabilized in the trapped conformation by interactions with the extruded RNA, which is also contacted by the interior of the secondary channel (Cheung and Cramer, 2011).