Journal
MOLECULAR MICROBIOLOGY
Volume 115, Issue 1, Pages 99-115Publisher
WILEY
DOI: 10.1111/mmi.14601
Keywords
Bacillussubtilis; NCIB 310; RNA-DNA hybrid; RNase HI; SOS response
Categories
Funding
- Division of Graduate Education [DEG 1256260]
- Division of Molecular and Cellular Biosciences [MCB 1714539]
- National Institute of General Medical Sciences [GM123635, GM131772, R35 GM131783]
- National Institutes of Health [GM123635]
- National Science Foundation [DEG 1256260, MCB 1714539]
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RNA-DNA hybrids can affect chromosome replication during cell growth, and various RNase H enzymes play important roles in bacterial cells. Bacillus subtilis NCIB 3610 has three functional RNase H enzymes, with the plasmid-encoded RNase HI contributing significantly to chromosome stability.
RNA-DNA hybrids form throughout the chromosome during normal growth and under stress conditions. When left unresolved, RNA-DNA hybrids can slow replication fork progression, cause DNA breaks, and increase mutagenesis. To remove hybrids, all organisms use ribonuclease H (RNase H) to specifically degrade the RNA portion. Here we show that, in addition to chromosomally encoded RNase HII and RNase HIII,Bacillus subtilisNCIB 3610 encodes a previously uncharacterized RNase HI protein, RnhP, on the endogenous plasmid pBS32. Like other RNase HI enzymes, RnhP incises Okazaki fragments, ribopatches, and a complementary RNA-DNA hybrid. We show that while chromosomally encoded RNase HIII is required for pBS32 hyper-replication, RnhP compensates for the loss of RNase HIII activity on the chromosome. Consequently, loss of RnhP and RNase HIII impairs bacterial growth. We show that the decreased growth rate can be explained by laggard replication fork progression near the terminus region of the right replichore, resulting in SOS induction and inhibition of cell division. We conclude that all three functional RNase H enzymes are present inB. subtilisNCIB 3610 and that the plasmid-encoded RNase HI contributes to chromosome stability, while the chromosomally encoded RNase HIII is important for chromosome stability and plasmid hyper-replication.
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