LetR is a TetR family transcription factor from Lysobacter controlling antifungal antibiotic biosynthesis
- 449 Downloads
Heat-stable antifungal factor (HSAF) is a newly identified and broad-spectrum antifungal antibiotic from Lysobacter enzymogenes, a ubiquitous environmental proteobacterium. Yet, the regulatory mechanism for HSAF biosynthesis in L. enzymogenes remains poorly understood. Here, we report the identification of a TetR-family protein Le1552 (LetR) from L. enzymogenes strain OH11 that is involved in transcriptional repression of HSAF production. Bacterial one-hybrid and gel mobility shift assays show that LetR directly binds to PHSAF (the promoter region of the HSAF biosynthesis operon). A DNA truncation assay further reveals a core region in PHSAF that is responsible for LetR binding. In-frame deletion of letR in wild-type OH11 is found to significantly increase HSAF levels and key biosynthetic gene transcription, while overexpression of letR in the wild-type background remarkably reduces HSAF levels as well as related gene expression instead. Together, we have identified not only a new regulator for the HSAF biosynthesis but also constructed a higher HSAF-producing deletion strain (ΔletR) of L. enzymogenes, which shall be of great value in promoting HSAF production for pharmaceutical and biological control purposes.
KeywordsLysobacter HSAF TetR Regulation Antibiotics
We thank Prof. Liangcheng Du (University of Nebraska-Lincoln) for the suggestions on manuscript organization. We also thank Prof. Zheng-Guo He from Huazhong Agricultural University (China) for kindly providing the bacterial one-hybrid system. Author contributions: G.Q. and F.L. conceived the project and designed experiments. P.W and H.C carried out experiments. P.W., H. C., G.Q., and F.L. analyzed data. P.W and G.Q. wrote the manuscript draft. F.L. revised the manuscript.
Compliance with ethical standards
This study was supported by National Basic Research (973) program of China (2015CB150600 to G.Q.), the Fundamental Research Funds for the Central Universities (Y0201600126 and KYTZ201403 to G.Q.), Special Fund for Agro-Scientific Research in the Public Interest (no. 201303015 to G.Q. and F.L.), National Natural Science Foundation of China (31371981 and 31572046 to G.Q.) and the Jiangsu Provincial Key Technology Support Program (BE2014386 and BE2015354 to F.L.), the Basal Research Funds from JAAS [ZX(15)1006 to F. L.], Jiangsu Agricultural Science and Technology Innovation Funds [CX(16)1049 to F. L.), ‘948’ Project of the Ministry of Agriculture (2014-Z24 to F.L.), and National pear industry technology system (CARS-29-09 to F.L. and G.Q.).
Conflict of interest
The authors declare that they have no competing interests.
This article does not contain any studies with human participants or animals.
- Hoang TT, Karkhoff-Schweizer RR, Kutchma AJ, Schweizer HP (1998) A broad-host-range Flp-FRT recombination system for site-specific excision of chromosomally-located DNA sequences: application for isolation of unmarked Pseudomonas aeruginosa mutants. Gene 212(1):77–86. doi: 10.1016/S0378-1119(98)00130-9 CrossRefPubMedGoogle Scholar
- Kobayashi DY, Reedy RM, Palumbo JD, Zhou JM, Yuen GY (2005) A clp gene homologue belonging to the Crp gene family globally regulates lytic enzyme production, antimicrobial activity, and biological control activity expressed by Lysobacter enzymogenes strain C3. Appl Environ Microbiol 71(1):261–269. doi: 10.1128/AEM.71.1.261-269.2005 CrossRefPubMedPubMedCentralGoogle Scholar
- Li Y, Chen H, Ding Y, Xie Y, Wang H, Cerny RL, Shen YM, Du LC (2014) Iterative assembly of two separate polyketide chains by the same single-module bacterial polyketide synthase in the biosynthesis of HSAF. Angew Chem Int Ed Engl 53(29):7524–7530. doi: 10.1002/anie.201403500 CrossRefPubMedPubMedCentralGoogle Scholar
- Qian GL, Wang YS, Qian DY, Fan JQ, Hu BS, Liu FQ (2012) Selection of available suicide vectors for gene mutagenesis using chiA (a chitinase encoding gene) as a new reporter and primary functional analysis of chiA in Lysobacter enzymogenes strain OH11. World J Microbiol Biotechnol 28(2):549–557. doi: 10.1007/s11274-011-0846-8 CrossRefPubMedGoogle Scholar
- Qian GL, Wang YL, Liu YR, Xu FF, He YW, Du LC, Venturi V, Fan JQ, Hu BS, Liu FQ (2013) Lysobacter enzymogenes uses two distinct cell-cell signaling systems for differential regulation of secondary-metabolite biosynthesis and colony morphology. Appl Environ Microbiol 79(21):6604–6616. doi: 10.1128/AEM.01841-13 CrossRefPubMedPubMedCentralGoogle Scholar
- Stratigopoulos G, Cundliffe E (2002) Expression analysis of the tylosin-biosynthetic gene cluster: pivotal regulatory role of the tylQ gene product. Chem Biol 9(1): 71–78. doi: org/10.1016/S1074-5521(01)00095-3
- Wang YS, Zhao YX, Zhang J, Zhao YY, Shen Y, Su ZH, Xu GG, Du LC, Huffman JM, Venturi V, Qian GL, Liu FQ (2014) Transcriptomic analysis reveals new regulatory roles of Clp signaling in secondary metabolite biosynthesis and surface motility in Lysobacter enzymogenes OH11. Appl Microbiol Biotechnol 98(21):9009–9020. doi: 10.1007/s00253-014-6072-1 CrossRefPubMedPubMedCentralGoogle Scholar
- Yu FG, Zaleta-Rivera K, Zhu XC, Huffman J, Millet JC, Harris SD, Yuen G, Li XC, Du LC (2007) Structure and biosynthesis of heat-stable antifungal factor (HSAF), a broad-spectrum antimycotic with a novel mode of action. Antimicrob Agents Ch 51(1):64–72. doi: 10.1128/AAC.00931-06 CrossRefGoogle Scholar
- Zhang W, Li YY, Qian GL, Wang Y, Chen HT, Li YZ, Liu FQ, Shen YM, Du LC (2011) Identification and characterization of the anti-methicillin-resistant Staphylococcus aureus WAP-8294A2 biosynthetic gene cluster from Lysobacter enzymogenes OH11. Antimicrob Agents Ch 55(12):5581–5589. doi: 10.1128/AAC.05370-11 CrossRefGoogle Scholar