Supercoiling, R-loops, Replication and the Functions of Bacterial Type 1A Topoisomerases

Abstract

Type 1A topoisomerases (topos) are the only topos that bind single-stranded DNA and the only ones found in all cells of the three domains of life. Two subfamilies, topo I and topo III, are present in bacteria. Topo I, found in all of them, relaxes negative supercoiling, while topo III acts as a decatenase in replication. However, recent results suggest that they can also act as back-up for each other. Because they are ubiquitous, type 1A enzymes are expected to be essential for cell viability. Single topA (topo I) and topB (topo III) null mutants of Escherichia coli are viable, but for topA only with compensatory mutations. Double topA topB null mutants were initially believed to be non-viable. However, in two independent studies, results of next generation sequencing (NGS) have recently shown that double topA topB null mutants of Bacillus subtilis and E. coli are viable when they carry parC parE gene amplifications. These genes encode the two subunits of topo IV, the main cellular decatenase. Here, we discuss the essential functions of bacterial type 1A topos in the context of this observation and new results showing their involvement in preventing unregulated replication from R-loops.

Keywords: PriA; R-loop; oriC; replication; supercoiling; topA; topB; topoisomerase I; topoisomerase III; topoisomerases.

Figure 1

Model for the effects of deleting topA and topB on replication in E. coli cells. (A) Topological problems associated with replication elongation (1), replication termination (2 and 3) and head-on conflicts between replication and transcription (4). (B) Supercoiling during transcription. (C) In wild-type cells, bi-directional replication is initiated at oriC and is terminated when replication forks converge in the Ter region. Replication forks are trapped in the Ter region via the Ter/Tus barriers [106]. In Escherichia coli cells lacking type 1A topos, the topological problems illustrated in A are exacerbated because of over-replication from oriC and PriA-dependent replication initiation (e.g., R-loops) that takes place outside of oriC. Black and red arrows indicate the direction of, respectively, replication and transcription (rrn operons). The absence of type 1A topos activity during replication further aggravates the topological problems. See text for details.

Figure 1

Model for the effects of deleting topA and topB on replication in E. coli cells. (A) Topological problems associated with replication elongation (1), replication termination (2 and 3) and head-on conflicts between replication and transcription (4). (B) Supercoiling during transcription. (C) In wild-type cells, bi-directional replication is initiated at oriC and is terminated when replication forks converge in the Ter region. Replication forks are trapped in the Ter region via the Ter/Tus barriers [106]. In Escherichia coli cells lacking type 1A topos, the topological problems illustrated in A are exacerbated because of over-replication from oriC and PriA-dependent replication initiation (e.g., R-loops) that takes place outside of oriC. Black and red arrows indicate the direction of, respectively, replication and transcription (rrn operons). The absence of type 1A topos activity during replication further aggravates the topological problems. See text for details.