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Identification of a gene from Streptomyces rimosus M527 negatively affecting rimocidin biosynthesis and morphological differentiation

  • Applied microbial and cell physiology
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Abstract

The polyene macrolide rimocidin, produced by Streptomyces rimosus M527, was found to be highly effective against a broad range of fungal plant pathogens. Current understanding of the regulatory mechanism of rimocidin biosynthesis and morphological differentiation in S. rimosus M527 is limited. NsdA is considered a negative regulator involved in morphological differentiation and biosynthesis of secondary metabolites in some Streptomyces species. In this study, nsdAsr was cloned from S. rimosus M527. The role of nsdAsr in rimocidin biosynthesis and morphological differentiation was investigated by gene deletion, complementation, and over-expression. A ΔnsdAsr mutant was obtained using CRISPR/Cas9. The mutant produced more rimocidin (46%) and accelerated morphological differentiation than the wild-type strain. Over-expression of nsdAsr led to a decrease in rimocidin production and impairment of morphological differentiation. Quantitative RT-PCR analysis revealed that transcription of rim genes responsible for rimocidin biosynthesis was upregulated in the ΔnsdAsr mutant but downregulated in the nsdAsr over-expression strain. Similar effects have been described for Streptomyces coelicolor M145 and the industrial toyocamycin-producing strain Streptomyces diastatochromogenes 1628.

Key points

• A negative regulator for sporulation and rimocidin production was identified.

• The CRISPR/Cas9 system was used for gene deletion in S. rimosus M527.

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References

  • Bednarz B, Kotowska M, Pawlik KJ (2019) Multi-level regulation of coelimycin synthesis in Streptomyces coelicolor A3(2). Appl Microbiol Biotechnol 103(16):6423–6434

    CAS  PubMed  PubMed Central  Google Scholar 

  • Bruheim P, Sletta H, Bibb MJ, White J, Levine DW (2002) High-yield actinorhodin production in fed-batch culture by a Streptomyces lividans strain overexpressing the pathway-specific activator gene actII-ORF4. J Ind Microbiol Biotechnol 28(2):103–111

    CAS  PubMed  Google Scholar 

  • Caffrey P, Aparicio JF, Malpartida F, Zotchev SB (2008) Biosynthetic engineering of polyene macrolides towards generation of improved antifungal and antiparasitic agents. Curr Top Med Chem 8(8):639–653

    CAS  PubMed  Google Scholar 

  • Cobb RE, Wang Y, Zhao H (2015) High-efficiency multiplex genome editing of Streptomyces species using an engineered CRISPR/Cas system. ACS Synth Biol 4(6):723–728

    CAS  PubMed  Google Scholar 

  • Escudero L, Al-Refai M, Nieto C, Laatsch H, Malpartida F, Seco EM (2015) New rimocidin/CE-108 derivatives obtained by a crotonyl-CoA carboxylase/reductase gene disruption in Streptomyces diastaticus var. 108: substrates for the polyene carboxamide synthase PcsA. PLoS One 10(8):e0135891

    PubMed  PubMed Central  Google Scholar 

  • Fedoryshyn M, Petzke L, Welle E, Bechthold A, Luzhetskyy A (2008) Marker removal from actinomycetes genome using Flp recombinase. Gene 419(1-2):43–47

    CAS  PubMed  Google Scholar 

  • Gibson DG, Benders GA, Axelrod KC, Zaveri J, Algire MA, Moodie M, Montague MG, Venter JC, Smith HO, Hutchison CA (2008) One-step assembly in yeast of 25 overlapping DNA fragments to form a complete synthetic Mycoplasma genitalium genome. Proc Natl Acad Sci U S A 105(51):20404–20409

    CAS  PubMed  PubMed Central  Google Scholar 

  • Hou B, Tao L, Zhu X, Wu W, Guo M, Ye J, Wu H, Zhang H (2018) Global regulator BldA regulates morphological differentiation and lincomycin production in Streptomyces lincolnensis. Appl Microbiol Biotechnol 102(9):4101–4115

    CAS  PubMed  Google Scholar 

  • Imai Y, Sato S, Tanaka Y, Ochi K, Hosaka T (2015) Lincomycin at subinhibitory concentrations potentiates secondary metabolite production by Streptomyces spp. Appl Environ Microbiol 81(11):3869–3879

    CAS  PubMed  PubMed Central  Google Scholar 

  • Jeon BJ, Kim JD, Han JW, Kim BS (2016) Antifungal activity of rimocidin and a new rimocidin derivative BU16 produced by Streptomyces mauvecolor BU16 and their effects on pepper anthracnose. J Appl Microbiol 120(5):1219–1228

    CAS  PubMed  Google Scholar 

  • Kang Y, Wang Y, Hou B, Wang R, Ye J, Zhu X, Wu H, Zhang H (2019) AdpAlin, a pleiotropic transcriptional regulator, is involved in the cascade regulation of lincomycin biosynthesis in Streptomyces lincolnensis. Front Microbiol 10:2428

    PubMed  PubMed Central  Google Scholar 

  • Kemung HM, Tan LT, Khan TM, Chan KG, Pusparajah P, Goh BH, Lee LH (2018) Streptomyces as a prominent resource of future anti-MRSA drugs. Front Microbiol 9:2221

    PubMed  PubMed Central  Google Scholar 

  • Kieser T, Bibb MJ, Buttner MJ, Chater KF, Hopwood DA (2000) Practical Streptomyces genetics. John Innes Foundation, Norwich

    Google Scholar 

  • Li W, Ying X, Guo Y, Yu Z, Zhou X, Deng Z, Kieser H, Chater KF, Tao M (2006) Identification of a gene negatively affecting antibiotic production and morphological differentiation in Streptomyces coelicolor A3(2). J Bacteriol 188(24):8368–8375

    CAS  PubMed  PubMed Central  Google Scholar 

  • Liu R, Deng Z, Liu T (2018) Streptomyces species: ideal chassis for natural product discovery and overproduction. Metab Eng 50:74–84

    CAS  PubMed  Google Scholar 

  • Lu D, Ma Z, Xu X, Yu X (2016) Isolation and identification of biocontrol agent Streptomyces rimosus M527 against Fusarium oxysporum f. sp. cucumerinum. J Basic Microbiol 56(8):929–933

    CAS  PubMed  Google Scholar 

  • Luo S, Chen XA, Mao XM, Li YQ (2018) Transposon-based identification of a negative regulator for the antibiotic hyper-production in Streptomyces. Appl Microbiol Biotechnol 102(15):6581–6592

    CAS  PubMed  Google Scholar 

  • Martín JF, Ramos A, Liras P (2019) Regulation of geldanamycin biosynthesis by cluster-situated transcription factors and the master regulator PhoP. Antibiotics (Basel) 8(3):87

    Google Scholar 

  • Ma Z, Liu J, Bechthold A, Tao L, Shentu X, Bian Y, Yu X (2014a) Development of intergeneric conjugal gene transfer system in Streptomyces diastatochromogenes 1628 and its application for improvement of toyocamycin production. Curr Microbiol 68(2):180–185

    CAS  PubMed  Google Scholar 

  • Ma Z, Tao L, Bechthold A, Shentu X, Bian Y, Yu X (2014b) Overexpression of ribosome recycling factor is responsible for improvement of nucleotide antibiotic-toyocamycin in Streptomyces diastatochromogenes 1628. Appl Microbiol Biotechnol 98(11):5051–5058

    CAS  PubMed  Google Scholar 

  • Mo J, Wang S, Zhang W, Li C, Deng Z, Zhang L, Qu X (2019) Efficient editing DNA regions with high sequence identity in actinomycetal genomes by a CRISPR-Cas9 system. Synth Syst Biotechnol 4(2):86–91

    PubMed  PubMed Central  Google Scholar 

  • Neldeborg S, Lin L, Stougaard M, Luo Y (2019) Rapid and efficient gene deletion by CRISPR/Cas9. Methods Mol Biol 1961:233–247

    CAS  PubMed  Google Scholar 

  • Novakova R, Núñez LE, Homerova D, Knirschova R, Feckova L, Rezuchova B, Sevcikova B, Menéndez N, Morís F, Cortés J, Kormanec J (2018) Increased heterologous production of the antitumoral polyketide mithramycin A by engineered Streptomyces lividans TK24 strains. Appl Microbiol Biotechnol 102(2):857–869

    CAS  PubMed  Google Scholar 

  • Olanrewaju OS, Babalola OO (2019) Streptomyces: implications and interactions in plant growth promotion. Appl Microbiol Biotechnol 103(3):1179–1188

    CAS  PubMed  Google Scholar 

  • Qiu J, Zhuo Y, Zhu D, Zhou X, Zhang L, Bai L, Deng Z (2011) Overexpression of the ABC transporter AvtAB increases avermectin production in Streptomyces avermitilis. Appl Microbiol Biotechnol 92(2):337–345

    CAS  PubMed  Google Scholar 

  • Rezuchova B, Homerova D, Sevcikova B, Núñez LE, Novakova R, Feckova L, Skultety L, Cortés J, Kormanec J (2018) An efficient blue-white screening system for markerless deletions and stable integrations in Streptomyces chromosomes based on the blue pigment indigoidine biosynthetic gene bpsA. Appl Microbiol Biotechnol 102(23):10231–10244

    CAS  PubMed  Google Scholar 

  • Sambrook J, Russel DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory, New York

    Google Scholar 

  • Seco EM, Miranzo D, Nieto C, Malpartida F (2010) The pcsA gene from Streptomyces diastaticus var. 108 encodes a polyene carboxamide synthase with broad substrate specificity for polyene amides biosynthesis. Appl Microbiol Biotechnol 85(6):1797–1807

    CAS  PubMed  Google Scholar 

  • Seco EM, Pérez-Zúñiga FJ, Rolón MS, Malpartida F (2004) Starter unit choice determines the production of two tetraene macrolides, rimocidin and CE-108, in Streptomyces diastaticus var. 108. Chem Biol 11(3):357–366

    CAS  PubMed  Google Scholar 

  • Sohoni SV, Fazio A, Workman CT, Mijakovic I, Lantz AE (2014) Synthetic promoter library for modulation of actinorhodin production in Streptomyces coelicolor A3(2). PLoS One 9(6):e99701

    PubMed  PubMed Central  Google Scholar 

  • Song ZQ, Liao ZJ, Hu YF, Ma Z, Bechthold A, Yu XP (2019) Development and optimization of an intergeneric conjugation system and analysis of promoter activity in Streptomyces rimosus M527. J Zhejiang Univ Sci B 20(11):891–900

    CAS  PubMed  PubMed Central  Google Scholar 

  • Song ZQ, Ma Z, Bechthold A, Yu XP (2020) Effects of addition of elicitors on rimocidin biosynthesis in Streptomyces rimosus M527. Appl Microbiol Biotechnol 104(10):4445–4455

    CAS  PubMed  Google Scholar 

  • Sowiński P, Pawlak J, Borowski E, Gariboldi P (1995) Stereostructure of rimocidin. J Antibiot (Tokyo) 48(11):1288–1291

    Google Scholar 

  • Tong Y, Charusanti P, Zhang L, Weber T, Lee SY (2015) CRISPR-Cas9 based engineering of actinomycetal genomes. ACS Synth Biol 4(9):1020–1029

    CAS  PubMed  Google Scholar 

  • Wang T, Bai L, Zhu D, Lei X, Liu G, Deng Z, You D (2012) Enhancing macrolide production in Streptomyces by coexpressing three heterologous genes. Enzym Microb Technol 50(1):5–9

    CAS  Google Scholar 

  • Wang XJ, Guo SL, Guo WQ, Xi D, Xiang WS (2009) Role of nsdA in negative regulation of antibiotic production and morphological differentiation in Streptomyces bingchengensis. J Antibiot (Tokyo) 62(6):309–313

    CAS  Google Scholar 

  • Wei J, Tian J, Pan G, Xie J, Bao J, Zhou Z (2017) Development and application of a T7 RNA polymerase-dependent expression system for antibiotic production improvement in Streptomyces. Biotechnol Lett 39(6):857–864

    CAS  PubMed  Google Scholar 

  • Wu H, Liu W, Shi L, Si K, Liu T, Dong D, Zhang T, Zhao J, Liu D, Tian Z, Yue Y, Zhang H, Xuelian B, Liang Y (2017) Comparative genomic and regulatory analyses of natamycin production of Streptomyces lydicus A02. Sci Rep 7(1):9114

    PubMed  PubMed Central  Google Scholar 

  • Xia H, Zhan X, Mao XM, Li YQ (2020) The regulatory cascades of antibiotic production in Streptomyces. World J Microbiol Biotechnol 36(1):13

    PubMed  Google Scholar 

  • Xu J, Song Z, Xu X, Ma Z, Bechthold A, Yu X (2019) ToyA, a positive pathway-specific regulator for toyocamycin biosynthesis in Streptomyces diastatochromogenes 1628. Appl Microbiol Biotechnol 103(17):7071–7084

    CAS  PubMed  Google Scholar 

  • Xu X, Wang J, Bechthold A, Ma Z, Yu X (2017) Selection of an efficient promoter and its application in toyocamycin production improvement in Streptomyces diastatochromogenes 1628. World J Microbiol Biotechnol 33(2):30

    PubMed  Google Scholar 

  • Yu J, Liu Q, Chen C, Qi X (2017) Antifungal activity change of Streptomyces rimosus MY02 mediated by confront culture with other microorganism. J Basic Microbiol 57(3):276–282

    CAS  PubMed  Google Scholar 

  • Zeng H, Wen S, Xu W, He Z, Zhai G, Liu Y, Deng Z, Sun Y (2015) Highly efficient editing of the actinorhodin polyketide chain length factor gene in Streptomyces coelicolor M145 using CRISPR/Cas9-CodA(sm) combined system. Appl Microbiol Biotechnol 99(24):10575–10585

    CAS  PubMed  Google Scholar 

  • Zhang MM, Wong FT, Wang Y, Luo S, Lim YH, Heng E, Yeo WL, Cobb RE, Enghiad B, Ang EL, Zhao H (2017) CRISPR-Cas9 strategy for activation of silent Streptomyces biosynthetic gene clusters. Nat Chem Biol 13:607–609

    CAS  Google Scholar 

  • Zhao Y, Song Z, Ma Z, Bechthold A, Yu X (2019) Sequential improvement of rimocidin production in Streptomyces rimosus M527 by introduction of cumulative drug-resistance mutations. J Ind Microbiol Biotechnol 46(5):697–708

    CAS  PubMed  Google Scholar 

  • Zhao YF, Lu DD, Bechthold A, Ma Z, Yu XP (2018) Impact of otrA expression on morphological differentiation, actinorhodin production, and resistance to aminoglycosides in Streptomyces coelicolor M145. J Zhejiang Univ Sci B 19(9):708–717

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgments

The authors are grateful to Prof. Yuhui Sun (Wuhan University) for kindly providing the pWHU2653. This work was supported by the National Natural Science Foundation of China (31772213, 31972320), and the excellent youth fund of Zhejiang province, China (LR17C140002).

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Authors and Affiliations

  1. Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, College of Life Sciences, China Jiliang University, Xueyuan Street, Xiasha Higher Education District, Hangzhou, 310018, Zhejiang Province, China

    Zhijun Liao, Zhangqing Song, Jie Xu, Zheng Ma & Xiaoping Yu

  2. Institute for Pharmaceutical Sciences, Pharmaceutical Biology and Biotechnology, University of Freiburg, 79104, Freiburg, Germany

    Andreas Bechthold

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  1. Zhijun Liao
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  2. Zhangqing Song
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  4. Zheng Ma
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  5. Andreas Bechthold
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  6. Xiaoping Yu
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Contributions

Z Liao, Z Song, and J Xu conducted experiments. Z Ma designed research and wrote this article. A Bechthold revised this article. XP Yu checked the final version. All authors read and approved the manuscript.

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Correspondence to Zheng Ma or Xiaoping Yu.

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Liao, Z., Song, Z., Xu, J. et al. Identification of a gene from Streptomyces rimosus M527 negatively affecting rimocidin biosynthesis and morphological differentiation. Appl Microbiol Biotechnol 104, 10191–10202 (2020). https://doi.org/10.1007/s00253-020-10955-8

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  • DOI: https://doi.org/10.1007/s00253-020-10955-8

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