Laboratory of Bacterial Genetics, Centre for DNA Fingerprinting and Diagnostics, Hyderabad, Telangana 500039, India.
Graduate Studies, Manipal Academy of Higher Education, Manipal, Karnataka 576104, India.
Nucleic Acids Res. 2018 Apr 20;46(7):3400-3411. doi: 10.1093/nar/gky118.
Transcription termination by Rho is essential for viability in various bacteria, including some major pathogens. Since Rho acts by targeting nascent RNAs that are not simultaneously translated, it also regulates antisense transcription. Here we show that RNase H-deficient mutants of Escherichia coli exhibit heightened sensitivity to the Rho inhibitor bicyclomycin, and that Rho deficiency provokes increased formation of RNA-DNA hybrids (R-loops) which is ameliorated by expression of the phage T4-derived R-loop helicase UvsW. We also provide evidence that in Rho-deficient cells, R-loop formation blocks subsequent rounds of antisense transcription at more than 500 chromosomal loci. Hence these antisense transcripts, which can extend beyond 10 kb in their length, are only detected when Rho function is absent or compromised and the UvsW helicase is concurrently expressed. Thus the potential for antisense transcription in bacteria is much greater than hitherto recognized; and the cells are able to retain viability even when nearly one-quarter of their total non-rRNA abundance is accounted for by antisense transcripts, provided that R-loop formation from them is curtailed.
Rho 介导的转录终止对于包括一些主要病原体在内的各种细菌的生存至关重要。由于 Rho 通过靶向同时未被翻译的新生 RNA 起作用，因此它还调节反义转录。在这里，我们表明大肠杆菌的 RNase H 缺陷突变体对 Rho 抑制剂双环霉素表现出更高的敏感性，并且 Rho 缺陷会引发 RNA-DNA 杂交（R 环）的形成增加，而噬菌体 T4 衍生的 R 环解旋酶 UvsW 的表达可以改善这种情况。我们还提供了证据表明，在 Rho 缺陷型细胞中，R 环的形成会阻止超过 500 个染色体基因座的后续反义转录。因此，只有在 Rho 功能缺失或受损且同时表达 UvsW 解旋酶时，才能检测到这些长度超过 10 kb 的反义转录本。因此，细菌中的反义转录的可能性比以前认识到的要大得多；并且只要它们的 R 环形成受到抑制，即使它们的总非 rRNA 丰度中有近四分之一被反义转录本占据，细胞也能够保持存活。
