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Random Insertional Mutagenesis Identifies Genes Associated with Virulence in the Wheat Scab Fungus Fusarium graminearum

First, second, third, and fifth authors: Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907; and fourth author: U.S. Department of Agriculture-Agricultural Research Service, Cereal Disease Laboratory, Department of Plant Pathology, University of Minnesota, St. Paul 55108

Fusarium graminearum is an important pathogen of small grains and maize in many areas of the world. To better understand the molecular mechanisms of F. graminearum pathogenesis, we used the restriction enzyme-mediated integration (REMI) approach to generate random insertional mutants. Eleven pathogenicity mutants were identified by screening 6,500 hygromycin-resistant transformants. Genetic analyses indicated that the defects in plant infection were tagged by the transforming vector in six of these mutants. In mutant M8, the transforming plasmid was integrated 110-bp upstream from the start codon of the cystathionine betalyase gene (CBL1). Gene replacement mutants deleted for CBL1 and the methionine synthase gene MSY1 were also obtained. Both the cbl1 and msy1 deletion mutants were methionine auxotrophic and significantly reduced in virulence on corn silks and wheat heads. We also identified genes disrupted by the transforming DNA in three other REMI mutants exhibiting reduced virulence. In mutants M68, the transforming vectors were inserted in the NADH: ubiquinone oxidoreductase. The putative b-ZIP transcription factor gene and the transducin beta-subunit-like gene disrupted in mutants M7 and M75, respectively, had no known homologs in filamentous fungi and were likely to be novel fungal virulence factors.

Additional keywords: auxotrophic mutant , cystathionine , Gibberella zeae , methionine .