Abstract
Lysobacter enzymogenes is an agriculturally important Gram-negative bacterium that employs a multitude of antifungal mechanisms to inhibit and infect filamentous fungal pathogens, through secretion of antifungal antibiotic HSAF (heat-stable antifungal factor), formation of T4P (type IV pilus)-mediated twitching motility, and production of extracellular chitinase. Interestingly, all such key antifungal factors seem to be controlled by Clp, a master regulator in L. enzymogenes; however, the underlying mechanisms are poorly understood. Here, employing strain OH11 as a working model, we show that Clp plays a dual role in controlling OH11 twitching motility. It controls transcription of pilA, a major T4P structure pilin gene, via directly binding to its promoter region, as well as regulates the gene transcription of pilMONOPQ operon, whose products were essential for T4P assembly, by directly binding to a similar promoter sequence. We also truncated the Clp-binding region of the pilA promoter fragment down to 41 bp to identify the potential Clp-binding sequence. In addition, the Clp-recognized pilM promoter motif of the L. enzymogenes strains is similarly conserved as the pilA promoter, both with a conserved 5′-GTG and a conserved CAC-3′, spaced by ten highly variable nucleotides. Thus, this study identified two direct and previously uncharacterized gene targets of Clp contributing to its regulation in the L. enzymogenes twitching motility. Overall, our findings further elucidate the molecular genetics of Clp-dependent twitching motility in Lysobacter.
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Acknowledgments
We thank Prof. Fengquan Liu in Jiangsu Academy of Agricultural Sciences for suggestions on manuscript organization. We also thank Dr. Wei Qian from Chinese Academy of Science for providing facilities to perform the MST assay and the anti-Clp antibody.
Funding
Funding in this study was supported by the Fundamental Research Funds for National Key Research and Development Program (2017YFD0201100 to G. Qian), the Central Universities (Y0201600126, KYT201805 and KYTZ201403 to G. Qian), National Natural Science Foundation of China (31572046 to G. Qian, and 31501589 to D. Shen), and Innovation Team Program for Jiangsu Universities (2017).
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G.Q., S.C., and B.O.O conceived the project and designed experiments. J.C., D.S., D.X., and S.H. carried out experiments. J.C. and G.Q. analyzed data. G.Q. and S.C. wrote the manuscript. All authors reviewed the manuscript.
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Chen, J., Shen, D., Odhiambo, B.O. et al. Two direct gene targets contribute to Clp-dependent regulation of type IV pilus-mediated twitching motility in Lysobacter enzymogenes OH11. Appl Microbiol Biotechnol 102, 7509–7519 (2018). https://doi.org/10.1007/s00253-018-9196-x
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DOI: https://doi.org/10.1007/s00253-018-9196-x