Applied Microbiology and Biotechnology

, Volume 103, Issue 1, pp 327–337 | Cite as

Isopropylmalate isomerase MoLeu1 orchestrates leucine biosynthesis, fungal development, and pathogenicity in Magnaporthe oryzae

  • Wei TangEmail author
  • Haolang Jiang
  • Qiaojia Zheng
  • Xuehang Chen
  • Rufeng Wang
  • Shuai Yang
  • Guiyuan Zhao
  • Jiao Liu
  • Justice Norvienyeku
  • Zonghua WangEmail author
Genomics, transcriptomics, proteomics


The biosynthesis of branched-chain amino acids (BCAAs) is conserved in fungi and plants, but not in animals. The Leu1 gene encodes isopropylmalate isomerase that catalyzes the conversion of α-isopropylmalate into β-isopropylmalate in the second step of leucine biosynthesis in yeast. Here, we identified and characterized the functions of MoLeu1, an ortholog of yeast Leu1 in the rice blast fungus Magnaporthe oryzae. The transcriptional level of MoLEU1 was increased during conidiation and in infectious stages. Cellular localization analysis indicated that MoLeu1 localizes to the cytoplasm at all stages of fungal development. Targeted gene deletion of MoLEU1 led to leucine auxotrophy, and phenotypic analysis of the generated ∆Moleu1 strain revealed that MoLeu1-mediated leucine biosynthesis was required for vegetative growth, asexual development, and pathogenesis of M. oryzae. We further observed that invasive hyphae produced by the ∆Moleu1 strain were mainly limited to the primary infected host cells. The application of exogenous leucine fully restored vegetative growth and partially restored conidiation as well as pathogenicity defects in the ∆Moleu1 strain. In summary, our results suggested that MoLeu1-mediated leucine biosynthesis crucially promotes vegetative growth, conidiogenesis, and pathogenicity of M. oryzae. This study helps unveil the regulatory mechanisms that are essential for infection-related morphogenesis and pathogenicity of the rice blast fungus.


Magnaporthe oryzae Isopropylmalate isomerase MoLeu1 Asexual development Pathogenicity Leucine biosynthesis 


Funding information

This research was supported by the Natural Science Foundation of China (31601584), Natural Science Foundation of Fujian Province (2016J05070), Science Fund for Distinguished Young Scholars of Fujian Agriculture and Forestry University to W. T. (KXJQ17020), and China Scholarship Council.

Compliance with ethical standards

This article does not contain any studies with human participants or animals performed by any of the authors.

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

253_2018_9456_MOESM1_ESM.pdf (849 kb)
ESM 1 (PDF 848 kb)


  1. Binder S (2010) Branched-chain amino acid metabolism in Arabidopsis thaliana. Arabidopsis Book 8:e0137. CrossRefPubMedPubMedCentralGoogle Scholar
  2. Bruno KS, Tenjo F, Li L, Hamer JE, Xu JR (2004) Cellular localization and role of kinase activity of PMK1 in Magnaporthe grisea. Eukaryot Cell 3(6):1525–1532. CrossRefPubMedPubMedCentralGoogle Scholar
  3. Chen Y, Zuo RF, Zhu Q, Sun Y, Li MY, Dong YH, Ru YY, Zhang HF, Zheng XB, Zhang ZG (2014) MoLys2 is necessary for growth, conidiogenesis, lysine biosynthesis, and pathogenicity in Magnaporthe oryzae. Fungal Genet Biol 67:51–57. CrossRefPubMedGoogle Scholar
  4. Do E, Hu G, Caza M, Oliveira D, Kronstad JW, Jung WH (2015) Leu1 plays a role in iron metabolism and is required for virulence in Cryptococcus neoformans. Fungal Genet Biol 75:11–19. CrossRefPubMedGoogle Scholar
  5. Du Y, Zhang H, Hong L, Wang J, Zheng X, Zhang Z (2013) Acetolactate synthases MoIlv2 and MoIlv6 are required for infection-related morphogenesis in Magnaporthe oryzae. Mol Plant Pathol 14(9):870–884. CrossRefPubMedGoogle Scholar
  6. Ebbole DJ (2007) Magnaporthe as a model for understanding host-pathogen interactions. Annu Rev Phytopathol 45(1):437–456. CrossRefPubMedGoogle Scholar
  7. Foster AJ, Ryder LS, Kershaw MJ, Talbot NJ (2017) The role of glycerol in the pathogenic lifestyle of the rice blast fungus Magnaporthe oryzae. Environ Microbiol 19(3):1008–1016. CrossRefPubMedGoogle Scholar
  8. Guo M, Gao F, Zhu X, Nie X, Pan Y, Gao Z (2015) MoGrr1, a novel F-box protein, is involved in conidiogenesis and cell wall integrity and is critical for the full virulence of Magnaporthe oryzae. Appl Microbiol Biotechnol 99(19):8075–8088. CrossRefPubMedGoogle Scholar
  9. Hamer L, Adachi K, Dezwaan T, Lo S, Montenegro-Chamorro MV, Frank S, Darveaux B, Mahanty S, Heiniger R, Skalchunes A, Pan H, Tarpey R, Shuster J, Tanzer MM (2004) Methods for the identification of inhibitors of 3-isopropylmalate dehydratase as antibiotics. United States Patent 6733963Google Scholar
  10. Howard RJ, Ferrari MA, Roach DH, Money NP (1991) Penetration of hard substrates by a fungus employing enormous turgor pressures. Proc Natl Acad Sci U S A 88(24):11281–11284CrossRefGoogle Scholar
  11. Hsu YP, Schimmel P (1984) Yeast LEU1. Repression of mRNA levels by leucine and relationship of 5′-noncoding region to that of LEU2. J Biol Chem 259(6):3714–3719PubMedGoogle Scholar
  12. Kimball SR, Jefferson LS (2006) Signaling pathways and molecular mechanisms through which branched-chain amino acids mediate translational control of protein synthesis. J Nutr 136(1 Suppl):227S–231S. CrossRefPubMedGoogle Scholar
  13. Kohlhaw GB (2003) Leucine biosynthesis in fungi: entering metabolism through the back door. Microbiol Mol Biol Rev 67(1):1–15.–15.2003Google Scholar
  14. Kong LA, Li GT, Liu Y, Liu MG, Zhang SJ, Yang J, Zhou XY, Peng YL, Xu JR (2013) Differences between appressoria formed by germ tubes and appressorium-like structures developed by hyphal tips in Magnaporthe oryzae. Fungal Genet Biol 56:33–41. CrossRefPubMedGoogle Scholar
  15. Liu WD, Xie SY, Zhao XH, Chen X, Zheng WH, Lu GD, Xu JR, Wang ZH (2010) A homeobox gene is essential for conidiogenesis of the rice blast fungus Magnaporthe oryzae. Mol Plant-Microbe Interact 23(4):366–375. CrossRefPubMedGoogle Scholar
  16. Liu X, Han Q, Wang J, Wang X, Xu J, Shi J (2016) Two FgLEU2 genes with different roles in leucine biosynthesis and infection-related morphogenesis in Fusarium graminearum. PLoS One 11(11):e0165927.
  17. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔC T method. Methods 25(4):402–408. CrossRefPubMedPubMedCentralGoogle Scholar
  18. McCourt JA, Duggleby RG (2006) Acetohydroxyacid synthase and its role in the biosynthetic pathway for branched-chain amino acids. Amino Acids 31(2):173–210. CrossRefPubMedGoogle Scholar
  19. Monirujjaman M, Ferdouse A (2014) Metabolic and physiological roles of branched-chain amino acids. Adv Mol Biol 2014:1–6. CrossRefGoogle Scholar
  20. Norvienyeku J, Zhong Z, Lin L, Dang X, Chen M, Lin X, Zhang H, Anjago WM, Abdul W, Wang Z (2017) Methylmalonate-semialdehyde dehydrogenase mediated metabolite homeostasis essentially regulate conidiation, polarized germination and pathogenesis in Magnaporthe oryzae. Environ Microbiol 19(10):4256–4277. CrossRefPubMedGoogle Scholar
  21. Radford JA, Lieberman BA, Brison DR, Smith AR, Critchlow JD, Russell SA, Watson AJ, Clayton JA, Harris M, Gosden RG, Shalet SM (2001) Orthotopic reimplantation of cryopreserved ovarian cortical strips after high-dose chemotherapy for Hodgkin’s lymphoma. Lancet 357(9263):1172–1175CrossRefGoogle Scholar
  22. Ryan ED, Tracy JW, Kohlhaw GB (1973) Subcellular localization of the leucine biosynthetic enzymes in yeast. J Bacteriol 116(1):222–225PubMedPubMedCentralGoogle Scholar
  23. Solomon PS, Oliver RP (2001) The nitrogen content of the tomato leaf apoplast increases during infection by Cladosporium fulvum. Planta 213(2):241–249. CrossRefPubMedGoogle Scholar
  24. Solomon PS, Tan KC, Oliver RP (2003) The nutrient supply of pathogenic fungi; a fertile field for study. Mol Plant Pathol 4(3):203–210. CrossRefPubMedGoogle Scholar
  25. Sweigard JA, Chumley FG, Valent B (1992) Disruption of a Magnaporthe grisea cutinase gene. Mol Gen Genet 232(2):183–190PubMedGoogle Scholar
  26. Talbot NJ (2003) On the trail of a cereal killer: exploring the biology of Magnaporthe grisea. Annu Rev Microbiol 57:177–202. CrossRefPubMedGoogle Scholar
  27. Talbot NJ, Ebbole DJ, Hamer JE (1993) Identification and characterization of MPG1, a gene involved in pathogenicity from the rice blast fungus Magnaporthe grisea. Plant Cell 5(11):1575–1590. CrossRefPubMedPubMedCentralGoogle Scholar
  28. Tang QY, Zhang CX (2013) Data Processing System (DPS) software with experimental design, statistical analysis and data mining developed for use in entomological research. Insect Sci 20(2):254–260. CrossRefPubMedGoogle Scholar
  29. Tang W, Ru Y, Hong L, Zhu Q, Zuo R, Guo X, Wang J, Zhang H, Zheng X, Wang P, Zhang Z (2015) System-wide characterization of bZIP transcription factor proteins involved in infection-related morphogenesis of Magnaporthe oryzae. Environ Microbiol 17(4):1377–1396. CrossRefPubMedGoogle Scholar
  30. Wilson RA, Fernandez J, Quispe CF, Gradnigo J, Seng A, Moriyama E, Wright JD (2012) Towards defining nutrient conditions encountered by the rice blast fungus during host infection. PLoS One 7(10):e47392.
  31. Wilson RA, Talbot NJ (2009) Under pressure: investigating the biology of plant infection by Magnaporthe oryzae. Nat Rev Microbiol 7(3):185–195. CrossRefPubMedGoogle Scholar
  32. Yoshizawa F (2004) Regulation of protein synthesis by branched-chain amino acids in vivo. Biochem Biophys Res Commun 313(2):417–422CrossRefGoogle Scholar
  33. Yu JH, Hamari Z, Han KH, Seo JA, Reyes-Dominguez Y, Scazzocchio C (2004) Double-joint PCR: a PCR-based molecular tool for gene manipulations in filamentous fungi. Fungal Genet Biol 41(11):973–981. CrossRefPubMedGoogle Scholar
  34. Zhang H, Liu K, Zhang X, Tang W, Wang J, Guo M, Zhao Q, Zheng X, Wang P, Zhang Z (2011a) Two phosphodiesterase genes, PDEL and PDEH, regulate development and pathogenicity by modulating intracellular cyclic AMP levels in Magnaporthe oryzae. PLoS One 6(2):e17241. CrossRefPubMedPubMedCentralGoogle Scholar
  35. Zhang H, Tang W, Liu K, Huang Q, Zhang X, Yan X, Chen Y, Wang J, Qi Z, Wang Z, Zheng X, Wang P, Zhang Z (2011b) Eight RGS and RGS-like proteins orchestrate growth, differentiation, and pathogenicity of Magnaporthe oryzae. PLoS Pathog 7(12):e1002450. CrossRefPubMedPubMedCentralGoogle Scholar
  36. Zhang Y, Shi H, Liang S, Ning G, Xu N, Lu J, Liu X, Lin F (2015) MoARG1, MoARG5,6 and MoARG7 involved in arginine biosynthesis are essential for growth, conidiogenesis, sexual reproduction, and pathogenicity in Magnaporthe oryzae. Microbiol Res 180:11–22. CrossRefPubMedGoogle Scholar
  37. Zhao X, Xue C, Kim Y, Xu JR (2004) A ligation-PCR approach for generating gene replacement constructs in Magnaporthe grisea. Fungal Genet Newsl 51(1):17–18. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Wei Tang
    • 1
    Email author
  • Haolang Jiang
    • 1
  • Qiaojia Zheng
    • 1
  • Xuehang Chen
    • 1
  • Rufeng Wang
    • 1
  • Shuai Yang
    • 2
  • Guiyuan Zhao
    • 1
  • Jiao Liu
    • 1
  • Justice Norvienyeku
    • 1
    • 2
  • Zonghua Wang
    • 1
    • 3
    Email author
  1. 1.State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops, College of Plant ProtectionFujian Agriculture and Forestry UniversityFuzhouChina
  2. 2.State Key Laboratory for Ecological Pest Control of Fujian and Taiwan Crops, College of Life ScienceFujian Agriculture and Forestry UniversityFuzhouChina
  3. 3.Institute of Ocean ScienceMinjiang UniversityFuzhouChina

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