The reverse gyrase TopR1 is responsible for the homeostatic control of DNA supercoiling in the hyperthermophilic archaeon Sulfolobus solfataricus

Abstract

Maintaining an appropriate DNA topology with DNA‐based processes (DNA replication, transcription and recombination) is crucial for all three domains of life. In bacteria, the homeostatic regulation for controlling DNA supercoiling relies on antagonistic activities of two DNA topoisomerases, TopoI and gyrase. In hyperthermophilic crenarchaea, the presence of such a regulatory system is suggested as two DNA topoisomerases, TopoVI and reverse gyrase, catalyze antagonistic activities. To test this hypothesis, we estimated and compared the number of the TopoVI with that of the two reverse gyrases, TopR1 and TopR2, in Sulfolobus solfataricus cells maintained either at 80 or at 88°C, or reciprocally shifted from one temperature to the other. From the three DNA topoisomerases, TopR1 is the only one exhibiting significant quantitative variations in response to the up‐ and down‐shifts. In addition, the corresponding intrinsic activities of these three DNA topoisomerases were tested in vitro at both temperatures. Although temperature modulates the three DNA topoisomerases activities, TopR1 is the sole topoisomerase able to function at high temperature. Altogether, results presented in this study demonstrate, for the first time, that the DNA topological state of a crenarchaeon is regulated via a homeostatic control, which is mainly mediated by the fine‐tuning of TopR1.

G-Quadruplexes in the Archaea Domain

“…It also highlights one of the consequences of a high GC %: G4-prone motifs become more frequent ( Figure 5 ). In addition, all hyperthermophilic organism genomes encode a reverse gyrase, which positively supercoil DNA, possibly to protect the genome [ 86 ]. In future studies, it would be very interesting to carry out a genome-wide wet-lab experiment, for example, direct DNA sequencing of G-quadruplex loci as described in [ 87 , 88 ] or direct visualization of G-quadruplexes in living cells using specific antibodies, such as BG4 [ 89 ].…”

Direct observation of helicase–topoisomerase coupling within reverse gyrase

“…Coupling between these two subunits provides RG the ability to exclusively increase DNA linking number through a strandpassage reaction that relaxes negative supercoils and introduces positive supercoils (3). However, the strength of this coupling can vary: It is weak in the regulated Sulfolobus solfataricus (Sso) reverse gyrase 1 (RG1), which can relax negative supercoils even in the absence of ATP hydrolysis, but it is strong in the constitutive Sso reverse gyrase 2 (RG2), which cannot (13)(14)(15).…”

“…Here, we analyze Sso RG2 at single-turnover resolution, allowing us to observe discrete substeps in the catalytic cycle and extract from them a mechanistic understanding of the RG's function (24)(25)(26)(27)(28)(29). In addition to the tight coupling existing between the helicase and the topoisomerase domain, RG2 is the most highly processive reverse gyrase and it is able to work well at a temperature as low as 45°C (14,15).…”

Archaea: A Gold Mine for Topoisomerase Diversity

“…TopR2 is not involved in this regulation while TopoVI assumes the removal of the excess of positive supercoils when necessary. TopA is not either involved in the homeostatic control of the DNA supercoiling because both topA mRNA and TopoA protein amounts remain very low or even undetectable in Saccharolobus in all the conditions tested (Garnier and Nadal, 2008;Couturier et al, 2014Couturier et al, , 2019. Nevertheless, TopA could help or even replace TopoVI to decatenate DNA as it is still very efficient at unknotting or decatenating DNA at high temperature (Bizard et al, 2018).…”

“…Thus, from a biochemical point of view, the particularity of the Topo VI is to have two base pairs between the two cleavage sites ( Figure 3 ) instead of four base pairs for the type IIA enzymes ( Schoeffler and Berger, 2008 ). It was recently reported that the Topo VI from Saccharolobus shibatae (previously named Sulfolobus shibatae ) is able to relax negatively or positively supercoiled DNA with a preference for the positive supercoils, but is not very efficient at decatenating DNA ( Couturier et al, 2019 ). Finally, these enzymes are sensitive to the drugs that interfere with the ATPase activity, like radicicol that is a powerful Topo VI topoisomerases inhibitor ( Gadelle et al, 2005 ).…”