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A novel Rap-Phr system in Bacillus velezensis NAU-B3 regulates surfactin production and sporulation via interaction with ComA

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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.

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Funding

This work was supported by the National Key R&D Program of China (grants 2017YFD0201101), the Natural Science Foundation of Jiangsu Province, China (grant BK20181322), the National Natural Science Foundation of China (grants 31972325, 31660543), and the Special Fund for the Fundamental Research Funds for Central Universities (grant KYZ201708, JCQY201904).

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Author notes

  1. Zhao Liang, Jun-Qing Qiao, and Ping-Ping Li contributed equally to this work and share the first authorship.

Authors and Affiliations

  1. Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, Nanjing, 210095, Jiangsu Province, People’s Republic of China

    Zhao Liang, Ping-Ping Li, Lu-Lu Zhang, Zi-Xuan Qiao, Ling Lin, Chen-Jie Yu, Yang Yang, Muhammad Zubair, Qin Gu, Hui-Jun Wu & Xue-Wen Gao

  2. Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing, 210095, Jiangsu Province, People’s Republic of China

    Zhao Liang, Ping-Ping Li, Lu-Lu Zhang, Zi-Xuan Qiao, Ling Lin, Chen-Jie Yu, Yang Yang, Muhammad Zubair, Qin Gu, Hui-Jun Wu & Xue-Wen Gao

  3. Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, Jiangsu Province, People’s Republic of China

    Jun-Qing Qiao

  4. Nordreet UG Greifswald, Greifswald, Germany

    Rainer Borriss

  5. Institute of Marine Biotechnology e.V. (IMaB), Greifswald, Germany

    Rainer Borriss

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  1. Zhao Liang
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Contributions

ZL, JQ, PL, and LZ performed the major experiments and wrote the manuscript. ZQ and LL performed RT-PCR and real-time PCR analysis. CY and YY provided the bioinformatics data. HW designed the research content, analyzed the data, and wrote the manuscript. QG analyzed the data and discussed the data. MZ, RB, and XG analyzed the data and modified the manuscript. All authors read and approved the final manuscript.

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Correspondence to Hui-Jun Wu.

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Liang, Z., Qiao, JQ., Li, PP. et al. A novel Rap-Phr system in Bacillus velezensis NAU-B3 regulates surfactin production and sporulation via interaction with ComA. Appl Microbiol Biotechnol 104, 10059–10074 (2020). https://doi.org/10.1007/s00253-020-10942-z

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