Molecular Plant-Microbe Interactions (Dec 2001)

Mutational Analysis of β-Glucanase Genes from the Plant-Pathogenic Fungus Cochliobolus carbonum

  • Hoon Kim,
  • Joong-Hoon Ahn,
  • Jenifer M. Görlach,
  • Claudio Caprari,
  • John S. Scott-Craig,
  • Jonathan D. Walton

DOI
https://doi.org/10.1094/MPMI.2001.14.12.1436
Journal volume & issue
Vol. 14, no. 12
pp. 1436 – 1443

Abstract

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Two new β-glucanase-encoding genes, EXG2 and MLG2, were isolated from the plant-pathogenic fungus Cochliobolus carbonum using polymerase chain reaction based on amino acid sequences from the purified proteins. EXG2 encodes a 46.6-kDa exo-β1,3-glucanase and is located on the same 3.5-Mb chromosome that contains the genes of HC-toxin biosynthesis. MLG2 encodes a 26.8-kDa mixed-linked (β1,3-β1,4) glucanase with low activity against β1,4-glucan and no activity against β1,3-glucan. Specific mutants of EXG2 and MLG2 were constructed by targeted gene replacement. Strains with multiple mutations (genotypes exg1/mlg1, exg2/mlg1, mlg1/mlg2, and exg1/exg2/mlg1/mlg2) were also constructed by sequential disruption and by crossing. Total mixed-linked glucanase activity in culture filtrates of mlg1/mlg2 and exg1/exg2/mlg1/mlg2 mutants was reduced by approximately 73%. Total β1,3-glucanase activity was reduced by 10, 54, and 96% in exg2, mlg1, and exg1/exg2/mlg1/mlg2 mutants, respectively. The quadruple mutant showed only a modest decrease in growth on β1,3-glucan or mixed-linked glucan. None of the mutants showed any decrease in virulence.

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