Advertisement

Disruption of stcA blocks sterigmatocystin biosynthesis and improves echinocandin B production in Aspergillus delacroxii

  • Taoling MinEmail author
  • Lei Xiong
  • Yan Liang
  • Rui Xu
  • Chenchang Fa
  • Sheng Yang
  • Haifeng HuEmail author
Original Paper

Abstract

Echinocandin B (ECB) is an important lipohexapeptide used for chemical manufacture of the antifungal agent anidulafungin. Sterigmatocystin (ST) is a polyketide mycotoxin produced by certain species of Aspergillus such as Aspergillus delacroxii SIPIW15, which could produce both ECB and ST. However, the presence of the potent carcinogen ST will greatly affect the quality and safety of ECB production. Therefore, it is essential to eliminate the ST biosynthesis and increase ECB titers in Asp. delacroxii SIPIW15. In this study, the polyketide synthase gene (stcA) required for biosynthesis of ST and its flanking region in Asp. delacroxii SIPIW15 were cloned, sequenced and analyzed firstly. Based on Agrobacterium-mediated transformation, the ΔstcA mutant AMT-1 was obtained and its yield of ECB was increased by 40% without ST detected at the same time as compared to the original strain. The results of the fed-batch experiments showed that the ECB yield of the ΔstcA strain AMT-1 was increased to 2163 ± 31 mg/l and no ST was detected in the 50 l bioreactor. This work suggested that the ΔstcA strain AMT-1 has the potential for application in ECB production improvement, and more importantly, to eliminate ST-related environmental pollution in ECB fermentation industry.

Graphic abstract

Keywords

Echinocandin B Sterigmatocystin Agrobacterium-mediated transformation Aspergillus delacroxii 

Notes

Acknowledgements

This work was supported by Natural Science Foundation of Shanghai [2015ZR1440300] and Science and Technology Commission of Shanghai Municipality [11XD1423700]. We are grateful to Jun Lin and He Huang (Shanghai Institutes for Biological Sciences, Shanghai, China) for editing the English of the manuscript. We also thank Guihua Gong (Shanghai Institutes for Biological Sciences, Shanghai, China) for helpful advice in qRT-PCR.

References

  1. Abuodeh RO, Orbach MJ, Mandel MA, Das A, Galgiani JN (2000) Genetic transformation of Coccidioides immitis facilitated by Agrobacterium tumefaciens. J Infect Dis 181(6):2106–2110.  https://doi.org/10.1086/315525 CrossRefPubMedGoogle Scholar
  2. Adefarati AA, Hensens OD, Jones ET, Tkacz JS (1992) Pneumocandins from Zalerion arboricola V Glutamic acid- and leucine-derived amino acids in pneumocandin A0 (L-671,329) and distinct origins of the substituted proline residues in pneumocandins A0 and B0. J Antibiot (Tokyo) 45(12):1953–1957.  https://doi.org/10.7164/antibiotics.45.1953 CrossRefGoogle Scholar
  3. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W et al (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25(17):3389–3402.  https://doi.org/10.1093/nar/25.17.3389 CrossRefPubMedPubMedCentralGoogle Scholar
  4. Baltz RH (2014) Combinatorial biosynthesis of cyclic lipopeptide antibiotics: a model for synthetic biology to accelerate the evolution of secondary metabolite biosynthetic pathways. ACS Synth Biol 3(10):748–758.  https://doi.org/10.1021/sb3000673 CrossRefPubMedGoogle Scholar
  5. Beijersbergen A, Smith SJ, Hooykaas PJ (1994) Localization and topology of VirB proteins of Agrobacterium tumefaciens. Plasmid 32(2):212–218.  https://doi.org/10.1006/plas.1994.1057 CrossRefPubMedGoogle Scholar
  6. Bundock P, den Dulk-Ras A, Beijersbergen A, Hooykaas PJ (1995) Trans-kingdom T-DNA transfer from Agrobacterium tumefaciens to Saccharomyces cerevisiae. EMBO J 14(13):3206–3214.  https://doi.org/10.1002/j.1460-2075.1995.tb07323.x CrossRefPubMedPubMedCentralGoogle Scholar
  7. Bundock P, Mroczek K, Winkler AA, Steensma HY, Hooykaas PJ (1999) T-DNA from Agrobacterium tumefaciens as an efficient tool for gene targeting in Kluyveromyces lactis. Mol Gen Genet 261(1):115–121.  https://doi.org/10.1007/s004380050948 CrossRefPubMedGoogle Scholar
  8. Cacho RA, Jiang W, Chooi YH, Walsh CT, Tang Y (2012) Identification and characterization of the echinocandin B biosynthetic gene cluster from Emericella rugulosa NRRL 11440. J Am Chem Soc 134(40):16781–16790.  https://doi.org/10.1021/ja307220z CrossRefPubMedPubMedCentralGoogle Scholar
  9. Chen X, Stone M, Schlagnhaufer C, Romaine CP (2000) A fruiting body tissue method for efficient Agrobacterium-mediated transformation of Agaricus bisporus. Appl Environ Microbiol 66(10):4510–4513.  https://doi.org/10.1128/AEM.66.10.4510-4513.2000 CrossRefPubMedPubMedCentralGoogle Scholar
  10. Chen L, Yue Q, Zhang X, Xiang M, Wang C, Li S et al (2013) Genomics-driven discovery of the pneumocandin biosynthetic gene cluster in the fungus Glarea lozoyensis. BMC Genomics 14:339.  https://doi.org/10.1186/1471-2164-14-339 CrossRefPubMedPubMedCentralGoogle Scholar
  11. Emri T, Majoros L, Toth V, Pocsi I (2013) Echinocandins: production and applications. Appl Microbiol Biotechnol 97(8):3267–3284.  https://doi.org/10.1007/s00253-013-4761-9 CrossRefPubMedGoogle Scholar
  12. Frisch DA, Harris-Haller LW, Yokubaitis NT, Thomas TL, Hardin SH, Hall TC (1995) Complete sequence of the binary vector Bin 19. Plant Mol Biol 27(2):405–409.  https://doi.org/10.1007/BF00020193 CrossRefPubMedGoogle Scholar
  13. Gao W, Jiang L, Ge L, Chen M, Geng C, Yang G et al (2015) Sterigmatocystin-induced oxidative DNA damage in human liver-derived cell line through lysosomal damage. Toxicol In Vitro 29(1):1–7.  https://doi.org/10.1016/j.tiv.2014.08.007 CrossRefPubMedGoogle Scholar
  14. George J, Reboli AC (2012) Anidulafungin: when and how? The clinician's view. Mycoses 55(1):36–44.  https://doi.org/10.1111/j.1439-0507.2011.02052.x CrossRefPubMedGoogle Scholar
  15. Gouka RJ, Gerk C, Hooykaas PJ, Bundock P, Musters W, Verrips CT et al (1999) Transformation of Aspergillus awamori by Agrobacterium tumefaciens-mediated homologous recombination. Nat Biotechnol 17(6):598–601.  https://doi.org/10.1038/9915 CrossRefPubMedGoogle Scholar
  16. Gravelat FN, Askew DS, Sheppard DC (2012) Targeted gene deletion in Aspergillus fumigatus using the hygromycin-resistance split-marker approach. In: Brand AC, MacCallum DM (eds) Host-fungus interactions: methods and protocols. Humana Press, Totowa, pp 119–130.  https://doi.org/10.1007/978-1-61779-539-8_8 CrossRefGoogle Scholar
  17. Green MR, Sambrook J (2012) Molecular cloning: a laboratory manual, 4th edn. Cold Spring Harbor Laboratory Press, New YorkGoogle Scholar
  18. Guo M, Ye J, Gao D, Xu N, Yang J (2019) Agrobacterium-mediated horizontal gene transfer: Mechanism, biotechnological application, potential risk and forestalling strategy. Biotechnol Adv 37(1):259–270.  https://doi.org/10.1016/j.biotechadv.2018.12.008 CrossRefPubMedGoogle Scholar
  19. Higgens CE, Michel KH (1977) Antibiotic A-22082 and process for production thereof. USA Patent 4024246Google Scholar
  20. Hodges RL, Hodges DW, Goggans K, Xuei X, Skatrud P, McGilvray D (1994) Genetic modification of an echinocandin B-producing strain of Aspergillus nidulans to produce mutants blocked in sterigmatocystin biosynthesis. J Ind Microbiol 13(6):372–381.  https://doi.org/10.1007/BF01577222 CrossRefPubMedGoogle Scholar
  21. Hodges RL, Kelkar HS, Xuei X, Skatrud PL, Keller NP, Adams TH et al (2000) Characterization of an echinocandin B-producing strain blocked for sterigmatocystin biosynthesis reveals a translocation in the stcW gene of the aflatoxin biosynthetic pathway. J Ind Microbiol Biotechnol 25(6):333–341.  https://doi.org/10.1038/sj/jim/7000076 CrossRefPubMedGoogle Scholar
  22. Jiang W, Cacho RA, Chiou G, Garg NK, Tang Y, Walsh CT (2013) EcdGHK are three tailoring iron oxygenases for amino acid building blocks of the echinocandin scaffold. J Am Chem Soc 135(11):4457–4466.  https://doi.org/10.1021/ja312572v CrossRefPubMedPubMedCentralGoogle Scholar
  23. Jurjevic Z, Peterson SW, Solfrizzo M, Peraica M (2013) Sterigmatocystin production by nine newly described Aspergillus species in section Versicolores grown on two different media. Mycotoxin Res 29(3):141–145.  https://doi.org/10.1007/s12550-013-0160-4 CrossRefPubMedGoogle Scholar
  24. Keller-Schierlein W, Widmer J (1976) The aromatic amino acid constituent of echinocandin B: 3, 4-dihydroxyhomotyrosine (author's transl). Helv Chim Acta 59(6):2021–2031.  https://doi.org/10.1002/hlca.19760590615 CrossRefPubMedGoogle Scholar
  25. Klich M, Mendoza C, Mullaney E, Keller N, Bennett JW (2001) A new sterigmatocystin-producing Emericella variant from agricultural desert soils. Syst Appl Microbiol 24(1):131–138.  https://doi.org/10.1078/0723-2020-00007 CrossRefPubMedGoogle Scholar
  26. Lacroix B, Citovsky V (2013) The roles of bacterial and host plant factors in Agrobacterium-mediated genetic transformation. Int J Dev Biol 57(6–8):467–481.  https://doi.org/10.1387/ijdb.130199bl CrossRefPubMedGoogle Scholar
  27. Liu Y, Du M, Zhang G (2014) Proapoptotic activity of aflatoxin B1 and sterigmatocystin in HepG2 cells. Toxicol Rep 1:1076–1086.  https://doi.org/10.1016/j.toxrep.2014.10.016 CrossRefPubMedPubMedCentralGoogle Scholar
  28. Minto RE, Townsend CA (1997) Enzymology and molecular biology of aflatoxin biosynthesis. Chem Rev 97(7):2537–2556.  https://doi.org/10.1021/cr960032y CrossRefPubMedGoogle Scholar
  29. Mora-Lugo R, Zimmermann J, Rizk AM, Fernandez-Lahore M (2014) Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach. BMC Microbiol 14:247.  https://doi.org/10.1186/s12866-014-0247-x CrossRefPubMedPubMedCentralGoogle Scholar
  30. Nguyen KT, Ho QN, Pham TH, Phan T-N, Tran V-T (2016) The construction and use of versatile binary vectors carrying pyrG auxotrophic marker and fluorescent reporter genes for Agrobacterium-mediated transformation of Aspergillus oryzae (journal article). World J Microbiol Biotechnol 32(12):204.  https://doi.org/10.1007/s11274-016-2168-3 CrossRefPubMedGoogle Scholar
  31. Nyfeler R, Keller-Schierlein W (1974) Metabolites of microorganisms 143 Echinocandin B, a novel polypeptide-antibiotic from Aspergillus nidulans var. echinulatus: isolation and structural components. Helv Chim Acta 57(8):2459–2477.  https://doi.org/10.1002/hlca.19740570818 CrossRefPubMedGoogle Scholar
  32. Oakley CE, Edgerton-Morgan H, Oakley BR (2012) Tools for manipulation of secondary metabolism pathways: rapid promoter replacements and gene deletions in Aspergillus nidulans. Methods Mol Biol 944:143–161.  https://doi.org/10.1007/978-1-62703-122-6_10 CrossRefPubMedGoogle Scholar
  33. Park SY, Jeong MH, Wang HY, Kim JA, Yu NH, Kim S et al (2013) Agrobacterium tumefaciens-mediated transformation of the lichen fungus Umbilicaria muehlenbergii. PLoS ONE 8(12):e83896.  https://doi.org/10.1371/journal.pone.0083896 CrossRefPubMedPubMedCentralGoogle Scholar
  34. Rank C, Nielsen KF, Larsen TO, Varga J, Samson RA, Frisvad JC (2011) Distribution of sterigmatocystin in filamentous fungi. Fungal Biol 115(4–5):406–420.  https://doi.org/10.1016/j.funbio.2011.02.013 CrossRefPubMedGoogle Scholar
  35. Ream W (1989) Agrobacterium tumefaciens and Interkingdom Genetic Exchange. Annu Rev Phytopathol 27:583–618.  https://doi.org/10.1146/annurev.py.27.090189.003055 CrossRefPubMedGoogle Scholar
  36. Rho HS, Kang S, Lee YH (2001) Agrobacterium tumefaciens-mediated transformation of the plant pathogenic fungus Magnaporthe grisea. Mol Cells 12(3):407–411.  https://doi.org/10.1016/S1011-1344(01)00259-7 CrossRefPubMedGoogle Scholar
  37. Samson RA, Visagie CM, Houbraken J, Hong SB, Hubka V, Klaassen CH et al (2014) Phylogeny, identification and nomenclature of the genus Aspergillus. Stud Mycol 78:141–173.  https://doi.org/10.1016/j.simyco.2014.07.004 CrossRefPubMedPubMedCentralGoogle Scholar
  38. Song P, Yuan K, Qin T, Zhang K, Ji XJ, Ren L et al (2018) Metabolomics profiling reveals the mechanism of increased pneumocandin B0 production by comparing mutant and parent strains. J Ind Microbiol Biotechnol 45(9):767–780.  https://doi.org/10.1007/s10295-018-2047-z CrossRefPubMedGoogle Scholar
  39. Sun J, Xu R, Xiao S, Lu Y, Zhang Q, Xue C (2018) Agrobacterium tumefaciens-mediated transformation as an efficient tool for insertional mutagenesis of Kabatiella zeae. J Microbiol Methods 149:96–100.  https://doi.org/10.1016/j.mimet.2018.05.004 CrossRefPubMedGoogle Scholar
  40. Tokashiki J, Hayashi R, Yano S, Watanabe T, Yamada O, Toyama H et al (2019) Influence of alpha-1,3-glucan synthase gene agsE on protoplast formation for transformation of Aspergillus luchuensis. J Biosci Bioeng. 5:4.  https://doi.org/10.1016/j.jbiosc.2019.01.018 CrossRefGoogle Scholar
  41. Tóth V, Nagy CT, Miskei M, Pócsi I, Emri T (2011) Polyphasic characterization of “Aspergillus nidulans var. roseus” ATCC 58397. Folia Microbiol 56(5):381–388.  https://doi.org/10.1007/s12223-011-0059-4 CrossRefGoogle Scholar
  42. Vazquez JA, Sobel JD (2006) Anidulafungin: a novel echinocandin. Clin Infect Dis 43(2):215–222.  https://doi.org/10.1086/505204 CrossRefPubMedGoogle Scholar
  43. Versilovskis A, De Saeger S (2010) Sterigmatocystin: occurrence in foodstuffs and analytical methods–an overview. Mol Nutr Food Res 54(1):136–147.  https://doi.org/10.1002/mnfr.200900345 CrossRefPubMedGoogle Scholar
  44. Wang Y (2016) Screening the mutant with over-producing echinocandin B and improving its fermentation and extraction technology. Master, China State Institute of Pharmaceutical IndustryGoogle Scholar
  45. Wang JS, Groopman JD (1999) DNA damage by mycotoxins. Mutat Res 424(1–2):167–181.  https://doi.org/10.1016/S0027-5107(99)00017-2 CrossRefPubMedGoogle Scholar
  46. Weyda I, Yang L, Vang J, Ahring BK, Lubeck M, Lubeck PS (2017) A comparison of Agrobacterium-mediated transformation and protoplast-mediated transformation with CRISPR-Cas9 and bipartite gene targeting substrates, as effective gene targeting tools for Aspergillus carbonarius. J Microbiol Methods 135:26–34.  https://doi.org/10.1016/j.mimet.2017.01.015 CrossRefPubMedGoogle Scholar
  47. Wohlleben W, Mast Y, Muth G, Rottgen M, Stegmann E, Weber T (2012) Synthetic biology of secondary metabolite biosynthesis in actinomycetes: engineering precursor supply as a way to optimize antibiotic production. FEBS Lett 586(15):2171–2176.  https://doi.org/10.1016/j.febslet.2012.04.025 CrossRefPubMedGoogle Scholar
  48. Yu JH, Leonard TJ (1995) Sterigmatocystin biosynthesis in Aspergillus nidulans requires a novel type I polyketide synthase. J Bacteriol 177(16):4792–4800.  https://doi.org/10.1007/BF00003802 CrossRefPubMedPubMedCentralGoogle Scholar
  49. Zhang J, Liu C, Xie Y, Li N, Ning Z, Du N et al (2017) Enhancing fructooligosaccharides production by genetic improvement of the industrial fungus Aspergillus niger ATCC 20611. J Biotechnol 249:25–33.  https://doi.org/10.1016/j.jbiotec.2017.03.021 CrossRefPubMedGoogle Scholar
  50. Zouaoui N, Mallebrera B, Berrada H, Abid-Essefi S, Bacha H, Ruiz MJ (2016) Cytotoxic effects induced by patulin, sterigmatocystin and beauvericin on CHO-K1 cells. Food Chem Toxicol 89:92–103.  https://doi.org/10.1016/j.fct.2016.01.010 CrossRefPubMedGoogle Scholar
  51. Zupan JR, Zambryski P (1995) Transfer of T-DNA from Agrobacterium to the plant cell. Plant Physiol 107(4):1041–1047.  https://doi.org/10.1104/pp.107.4.1041 CrossRefPubMedPubMedCentralGoogle Scholar
  52. Zwiers LH, De Waard MA (2001) Efficient Agrobacterium tumefaciens-mediated gene disruption in the phytopathogen Mycosphaerella graminicola. Curr Genet 39(5–6):388–393.  https://doi.org/10.1007/s002940100216 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.State Key Lab of New Drugs and Pharmaceutical Process, Shanghai Institute of Pharmaceutical IndustryChina State Institute of Pharmaceutical IndustryShanghaiChina
  2. 2.Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological SciencesChinese Academy of SciencesShanghaiChina

Personalised recommendations