Molecular Mechanisms Controlling the Disease Cycle in the Vascular Pathogen Characterized Through Forward Genetics and Transcriptomics.

The soil-borne pathogen has a worldwide distribution and a plethora of hosts of agronomic value. Molecular analysis of virulence processes can identify targets for disease control. In this work, we compared the global gene transcription profile of random T-DNA insertion mutant strain D-10-8F, which exhibits reduced virulence and alterations in microsclerotium formation and polar growth, with that of the wild-type strain. Three genes identified as differentially expressed were selected for functional characterization. To produce deletion mutants, we developed an updated version of one-step construction of -recombination-ready plasmids (OSCAR) that included the negative selection marker (herpes simplex virus thymidine kinase gene) to prevent ectopic integration of the deletion constructs. Deletion of (), encoding a regulator of G protein signaling (RGS) protein and highly upregulated in the wild type versus D-10-8F, resulted in phenotypic alterations in development and virulence that were indistinguishable from those of the random T-DNA insertion mutant. In contrast, deletion of the other two genes selected, () and (), showed that they do not play major roles in morphogenesis or virulence in . Taken together the results presented here on the transcriptomic analysis and phenotypic characterization of D-10-8F and ∆ strains provide evidence that variations in G protein signaling control the progression of the disease cycle in . We propose that G protein-mediated signals induce the expression of multiple virulence factors during biotrophic growth, whereas massive production of microsclerotia at late stages of infection requires repression of G protein signaling via upregulation of VdRGS1 activity.