The oligosaccharyl transferase subunit STT3 mediates fungal development and is required for virulence in Verticillium dahliae

Verticillium dahliae is the most overwhelming plant pathogen, causing Verticillium wilt in a number of economic crops. The molecular mechanism is still unclear and identification of the genes involved in the pathogenicity or virulence of this fungus would benefit to uncover such mechanism. STT3 is a catalytic subunit of the multi-subunit oligosaccharyl transferase (OST) and plays an essential role in glycoprotein modification. Here, we characterized STT3 gene (VDAG_03232.1) of V. dahliae to explore its regulatory role in the development and virulence by deletion and complementation of this gene, as well as its silence in transgenic plants. The expression of the STT3 gene increased at the stage of conidia germination and reached its peak level with germ tube formation and elongation. We generated the knockout mutants (Delta STT3) using protoplast transformation. Mycelial growth, sporulation ability and glycoprotein secretion were impaired when Delta STT3 mutants were grown on media supplemented with different carbon sources. Moreover, Delta STT3 mutants exhibited distinctly decreased germination ratio and reduction in virulence compared with the wild type (Vd wt) and complementary (Delta STT3-C) strains. We also generated transgenic Nicotiana benthamiana (Trans-1 and -2) plants by expressing dsRNA against the STT3 gene. Transgenic plants showed significant reduction in the disease index and fungal biomass resulting in elevated resistance to V. dahliae compared with the wild-type plants when inoculated with Vd wt. Our results indicated that STT3 mediates the full virulence through the regulation in fungal development, hyphal growth, glycoprotein secretion of V. dahliae and merits further study as a potential RNAi target to control this fungus.