A Putative MAPK Kinase Kinase Gene<i>Ssos4</i>is Involved in Mycelial Growth, Virulence, Osmotic Adaptation, and Sensitivity to Fludioxonil and is Essential for SsHog1 Phosphorylation in<i>Sclerotinia sclerotiorum</i>

Abstract

The high osmolarity glycerol (HOG) pathway, comprising a two-component system and the Hog1 mitogen-activated protein kinase (MAPK) cascade, plays a pivotal role in eukaryotic organisms. Previous studies suggested that the biological functions of some key genes in the HOG pathway varied in filamentous fungi. In this study, we characterized a putative MAPK kinase kinase gene, <i>Ssos4</i>, in <i>Sclerotinia sclerotiorum</i>, which encoded a phosphotransferase in the MAPK cascade. Compared with the wild-type progenitor HA61, the deletion mutant ∆Ssos4-63 exhibited impaired mycelial growth, sclerotia formation, increased hyphal branches, and decreased virulence. The deficiencies of the deletion mutant ∆Ssos4-63 were recovered when the full-length <i>Ssos4</i> gene was complemented. Deletion of <i>Ssos4</i> increased the sensitivity to osmotic stresses and cell wall agents and the resistance to fludioxonil and dimethachlon. Intracellular glycerol accumulation was not induced in the deletion mutant ∆Ssos4-63 when treated with fludioxonil and NaCl and the phosphorylation of SsHog1 was also cancelled by the deletion of <i>Ssos4</i>. Consistent with the glycerol accumulation and increased expression levels of <i>SsglpA</i> and <i>Ssfps1</i>, controlling glycerol synthesis and close of glycerol channel under hyperosmotic stress, respectively, were detected in the wild-type strain HA61 but not in the deletion mutant ∆Ssos4-63. Moreover, the relative expression level of <i>Sshog1</i> significantly decreased, whereas the expression level of <i>Ssos5</i> increased in the deletion mutant ∆Ssos4-63. These results indicated that <i>Ssos4</i> played important roles in mycelial growth and differentiation, sclerotia formation, virulence, hyperosmotic adaptation, fungicide sensitivity, and the phosphorylation of SsHog1 in <i>S. sclerotiorum</i>.