Abstract

Several quorum sensing systems occurring in Bacillus subtilis, e.g. Rap-Phr systems, were reported to interact with major regulatory proteins, such as ComA, DegU, and Spo0A, in order to regulate competence, sporulation, and synthesis of secondary metabolites. In this study, we characterized a novel Rap-Phr system, RapA4-PhrA4, in Bacillus velezensis NAU-B3. We found that the rapA4 and phrA4 genes were co-transcribed in NAU-B3. When rapA4 was expressed in the heterologous host Bacillus subtilis OKB105, surfactin production and sporulation were severely inhibited. However, when the phrA4 was co-expressed, the RapA4 activity was inhibited. The transcription of the surfactin synthetase srfA gene and sporulation-related genes were also regulated by the RapA4-PhrA4 system. In vitro results obtained from electrophoretic mobility shift assay (EMSA) proved that RapA4 inhibits ComA binding to the promoter of the srfA operon, and the PhrA4 pentapeptide acts as anti-activator of RapA4. We also found that the F24 residue plays a key role in RapA4 function. This study indicated that the novel RapA4-PhrA4 system regulates the surfactin synthesis and sporulation via interaction with ComA, thereby supporting the bacterium to compete and to survive in a hostile environment.

Key points

•Bacillus velezensis NAU-B3 has a novel Rap-Phr quorum sensing system, which does not occur in model strains Bacillus subtilis 168 and B. velezensis FZB42.

•RapA4-PhrA4 regulates surfactin production and sporulation.

•RapA4-PhrA4 interacts with the ComA protein from ComP/ComA two-component system.

中文翻译：

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velezensis velacensis中的新型Rap-Phr系统NAU-B3通过与ComA相互作用调节表面活性素的产生和孢子形成

摘要

据报道，发生在枯草芽孢杆菌中的几种群体感应系统（例如Rap-Phr系统）与主要的调节蛋白（例如ComA，DegU和Spo0A）相互作用，以调节能力，孢子形成和次级代谢产物的合成。在这项研究中，我们表征了一种新的Rap-Phr系统，RapA4-PhrA4，在Velezensis芽孢杆菌NAU-B3中。我们发现rapA4和phrA4基因在NAU-B3中共转录。当rapA4在异源宿主枯草芽孢杆菌OKB105中表达时，表面活性素的产生和孢子形成被严重抑制。但是，当phrA4共表达时，RapA4活性受到抑制。表面活性蛋白合成酶srfA基因和孢子形成相关基因的转录也受RapA4-PhrA4系统调控。从电泳迁移率变动分析（EMSA）获得的体外结果证明，RapA4抑制ComA与srfA操纵子的启动子结合，而PhrA4五肽充当RapA4的抗激活剂。我们还发现F24残基在RapA4功能中起关键作用。这项研究表明，新型RapA4-PhrA4系统通过与ComA的相互作用来调节表面活性素的合成和孢子形成，从而支持细菌在不利的环境中竞争和生存。

关键点

•维尔茨芽孢杆菌NAU-B3具有新颖的Rap-Phr群体感应系统，在枯草芽孢杆菌168和维尔茨芽孢杆菌FZB42模型菌株中不存在。

•RapA4-PhrA4调节表面活性素的产生和孢子形成。

•RapA4-PhrA4与来自ComP / ComA两组分系统的ComA蛋白相互作用。