Topoisomerases I and III inhibit R-loop formation to prevent unregulated replication in the chromosomal Ter region of Escherichia coli

PLoS Genetics
Julien BrochuM Drolet

Abstract

Type 1A topoisomerases (topos) are the only ubiquitous topos. E. coli has two type 1A topos, topo I (topA) and topo III (topB). Topo I relaxes negative supercoiling in part to inhibit R-loop formation. To grow, topA mutants acquire compensatory mutations, base substitutions in gyrA or gyrB (gyrase) or amplifications of a DNA region including parC and parE (topo IV). topB mutants grow normally and topo III binds tightly to single-stranded DNA. What functions topo I and III share in vivo and how cells lacking these important enzymes can survive is unclear. Previously, a gyrB(Ts) compensatory mutation was used to construct topA topB null mutants. These mutants form very long filaments and accumulate diffuse DNA, phenotypes that appears to be related to replication from R-loops. Here, next generation sequencing and qPCR for marker frequency analysis were used to further define the functions of type 1A topos. The results reveal the presence of a RNase HI-sensitive origin of replication in the terminus (Ter) region of the chromosome that is more active in topA topB cells than in topA and rnhA (RNase HI) null cells. The S9.6 antibodies specific to DNA:RNA hybrids were used in dot-blot experiments to show the accumulation of R-loops in...Continue Reading

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Citations

Dec 20, 2018·Nucleic Acids Research·Phoolwanti Rani, Valakunja Nagaraja
Dec 14, 2018·Nucleic Acids Research·Sarah L Midgley-SmithChristian J Rudolph
Dec 18, 2019·The Journal of Biological Chemistry·Youssef A HegazyElizabeth J Tran
Apr 15, 2020·FEMS Microbiology Reviews·Anurag Kumar SinhaDavid R F Leach
Apr 9, 2019·Nucleic Acids Research·Nalini RaghunathanJayaraman Gowrishankar
May 1, 2020·Frontiers in Microbiology·Aisha H SyedaChristian J Rudolph
Apr 9, 2019·Nucleic Acids Research·Daniel R ReußJörg Stülke
Mar 19, 2019·Molecular Microbiology·Nicklas A HamiltonJustin Courcelle
Sep 1, 2019·DNA Repair·Marc Drolet, Julien Brochu
Mar 22, 2020·IScience·Li-Hong ZhangXiang Gao

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Datasets Mentioned

BETA
AP009048.1

Methods Mentioned

BETA
dot blots
dot blotting
dot-blot
dot-blots
Assay
PCR
Illumina sequencing
electrophoresis

Software Mentioned

PrimerQuest

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