Long-Distance Cooperative and Antagonistic RNA Polymerase Dynamics via DNA Supercoiling

Abstract

Genes are often transcribed by multiple RNA polymerases (RNAPs) at densities that can vary widely across genes and environmental conditions. Here, we provide in vitro and in vivo evidence for a built-in mechanism by which co-transcribing RNAPs display either collaborative or antagonistic dynamics over long distances (>2 kb) through transcription-induced DNA supercoiling. In Escherichia coli, when the promoter is active, co-transcribing RNAPs translocate faster than a single RNAP, but their average speed is not altered by large variations in promoter strength and thus RNAP density. Environmentally induced promoter repression reduces the elongation efficiency of already-loaded RNAPs, causing premature termination and quick synthesis arrest of no-longerneeded proteins. This negative effect appears independent of RNAP convoy formation and is abrogated by topoisomerase I activity. Antagonistic dynamics can also occur between RNAPs from divergently transcribed gene pairs. Our findings may be broadly applicable given that transcription on topologically constrained DNA is the norm across organisms.

Nascent transcript folding plays a major role in determining RNA polymerase elongation rates

“…Basic parameters were taken from published literature (as described in Extended Text), defining the initiation frequency, overall elongation rates, the torque generated by torsion in the DNA and the time distribution of the stochastic elongation and backtracking steps. Effects of DNA torsion were previously included in models for RNAPII convoys visualized following promoter bursting (Lesne et al, 2018;Tantale et al, 2016) and the cooperative behavior of bacterial polymerases (Heberling et al, 2016;Kim et al, 2019). Other features were fitted to the experimental data, including the negative correlation of elongation rate with high G+C content in the transcription bubble and the positive effects of stable folding of the nascent transcript.…”

“…Positive supercoils in front of the polymerase overwind DNA and strongly resist strand opening during elongation. Even modest changes in relative position can exert sufficient force to effectively block transcription elongation in vitro (Heberling et al, 2016;Kim et al, 2019;Lesne et al, 2018;Tantale et al, 2016).…”

Control of RNA polymerase II promoter-proximal pausing by DNA supercoiling

“…Thus, (-) supercoiling at promoter regions facilitates transcription initiation by allowing promoter melting and the formation of Pol II open complexes. In this same line, although upstream (-) supercoiling can stall bacterial RNA polymerase (Ma et al, 2013), when encountered downstream, it can favor transcriptional elongation by cancelling the (+) supercoiling generated (Kim et al, 2019). In this context, we propose that, by removing (-) supercoiling generated behind advancing Pol II complexes, TOP2A limits initiation and processivity of subsequent transcriptional cycles, imposing a strong requirement for TOP1-mediated removal of (+) supercoiling ahead of each elongating polymerase ( Figure 7).…”

“…It may seem counter-intuitive that a topoisomerase, which relieves torsional stress, acts to negatively regulate transcription. One must bear in mind however that (-) supercoiling is a well-established stimulator of transcription initiation, at least in vitro and in prokaryotic models (Chong et al, 2014;Kim et al, 2019;Parvin and Sharp, 1993;Revyakin et al, 2004;Tabuchi et al, 1993), and has been proposed to operate similarly in yeast and mammalian cells (Baranello et al, 2016;Bermudez et al, 2010). Thus, (-) supercoiling at promoter regions facilitates transcription initiation by allowing promoter melting and the formation of Pol II open complexes.…”

“…This could explain how merbarone treatment leads to a strong transcriptional stimulation of c-FOS without increased TOP1 activity ( Figure 6A). Interestingly, long-range communication between RNA polymerases by supercoiling cancelation and interference has been recently reported in bacteria (Kim et al, 2019). In any case, the accumulation of (-) supercoiling at promoter regions seems a wide-spread characteristic of eukaryotic gene organization (Bermudez et al, 2010;Kouzine et al, 2013;Matsumoto and Hirose, 2004;Naughton et al, 2013;Teves and Henikoff, 2014), and could serve additional functions such as, for example, favoring the formation of non-B DNA structures with regulatory functions (Kouzine et al, 2004;Kouzine et al, 2008), providing an appropriate 3D conformation to favor interaction with regulatory elements (Liu et al, 2001;Racko et al, 2019) or as a means to isolate and maintain the topology of the transcriptional unit (Achar et al, 2020).…”