Stress responsive glycosylphosphatidylinositol-anchored protein SsGSP1 contributes to <i>Sclerotinia sclerotiorum</i> virulence

Fungal cell wall acts as a defense barrier, shielding the cell from varying environmental stresses. Cell wall proteins, such as glycosylphosphatidylinositol (GPI)-anchored proteins, are involved in swift and appropriate responses to minor environmental changes in fungi. However, the roles of these proteins in the pathogenic <i>Sclerotinia sclerotiorum</i> remain largely unexplored. Here, we identified a novel GPI-anchored protein in <i>S. sclerotiorum</i>, SsGSP1, comprising a Kre9_KNH domain. <i>SsGSP1</i> was upregulated during infection, and the loss-of-function mutants of <i>SsGSP1</i> exhibited the compromised cell wall integrity and reduced β-glucan content. During inoculation on <i>Arabidopsis thaliana</i>, <i>Nicotiana benthamiana</i>, and <i>Brassica napus</i>, the <i>SsGSP1</i>-deletion strains demonstrated the decreased virulence. The transgenic <i>A</i>. <i>thaliana</i> line carrying the sRNA targeting <i>SsGSP1</i> enhanced resistance to <i>S. sclerotiorum</i> via Host-Induced Gene Silencing (HIGS). The <i>SsGSP1</i>-deficient strains displayed the heightened sensitivity to various stresses, including osmotic pressure, oxidative stress, and heat shock. The yeast two-hybrid and BiFC assays confirmed that SsGSP1 interacted with the key stress-related proteins catalase SsCat2, heat shock protein Sshsp60, and ABC transporter SsBMR1. Accordingly, transcriptome analysis revealed that the disruption of <i>SsGSP1</i> downregulated the expression of genes involved in oxidative stress response, heat shock response, and chemical agent resistance. These results collectively delineate the intricate role of GPI-anchored protein SsGSP1 in β-glucan, cell wall integrity, and virulence and may act as a potential surface sensor to elicit signal transduction in response to environmental stresses in <i>S. sclerotiorum</i>.