Sedivy Emma L, Smith Janet L, Grossman Alan D
Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America.
PLoS Genet. 2025 May 14;21(5):e1011625. doi: 10.1371/journal.pgen.1011625. eCollection 2025 May.
DnaA is the replication initiator and a transcription factor in virtually all bacteria. Although the synthesis and activity of DnaA are highly regulated, the mechanisms of regulation vary between organisms. We found that production of DnaA in Bacillus subtilis is regulated by an antisense RNA that overlaps with the 5' untranslated region upstream of the dnaA open reading frame. We initially observed this RNA in in vitro transcription experiments and found that its production was inhibited by DnaA. This RNA, now called ArrA for antisense RNA repressor of dnaA, is made in vivo. We identified the arrA promoter and made a mutation that greatly reduced (or eliminated) production of ArrA RNA in vitro and in vivo. In vivo, this arrA promoter mutation caused an increase in the amount of mRNA and protein from dnaA and dnaN, indicating that arrA expression normally inhibits expression of the dnaA-dnaN operon. The arrA mutation also caused a delay in sporulation that was alleviated by loss of sda, a sporulation-inhibitory gene that is directly activated by DnaA. arrA appears to be conserved in some members of the Bacillus genus, indicating that arrA has evolved in at least some endospore-forming bacteria to modulate production of DnaA and enable timely and robust sporulation.
DnaA是几乎所有细菌中的复制起始因子和转录因子。尽管DnaA的合成和活性受到高度调控，但调控机制在不同生物之间存在差异。我们发现，枯草芽孢杆菌中DnaA的产生受一种反义RNA的调控，该反义RNA与dnaA开放阅读框上游的5'非翻译区重叠。我们最初在体外转录实验中观察到这种RNA，并发现其产生受到DnaA的抑制。这种现在被称为ArrA（即dnaA的反义RNA阻遏物）的RNA在体内也会产生。我们鉴定出了arrA启动子，并制造了一个突变，该突变在体外和体内都极大地减少（或消除）了ArrA RNA的产生。在体内，这种arrA启动子突变导致来自dnaA和dnaN的mRNA和蛋白质数量增加，这表明arrA的表达通常会抑制dnaA - dnaN操纵子的表达。arrA突变还导致芽孢形成延迟，而sda基因（一个由DnaA直接激活的芽孢形成抑制基因）的缺失则缓解了这种延迟。arrA似乎在芽孢杆菌属的一些成员中是保守的，这表明arrA至少在一些形成芽孢的细菌中已经进化，以调节DnaA的产生并实现及时且强劲的芽孢形成。
