<scp> <i>Ustilago maydis</i> </scp> Phosphatidic Acid and Phosphatidylserine‐Targeting Secreted Lipase Lip3 Is Involved in Pathogenesis

Lipases constitute important virulence factors. By targeting specific lipids involved in various cellular processes, lipases regulate growth, development, and pathogenic mechanisms in many organisms. The Ustilago maydis genome codes for a set of secreted lipases that exhibit differential expression during infection. In this study, the biological function of one of the secreted lipases, Lip3, during pathogenesis of U. maydis was investigated. Pathogenicity assays with a deletion mutant of the gene showed slow progress of infection with much reduced sporulation. Through the analysis of the total lipids isolated from plants infected with either SG200 or SG200Δlip3, the substrate preference of Lip3 towards different phospholipids, including phosphatidic acid (PA) and phosphatidylserine (PS), was shown. The interaction of Lip3 with PA and PS was further confirmed through in vitro lipid binding assays. Recombinant Lip3 showed lipolytic activity against purified PA and PS from sunflower and Glycine max, respectively. Using a PS targeting lactadherin C2 domain-based biosensor, the differential distribution of PS within the biotrophic interfacial membrane of SG200 and SG200Δlip3 infected Zea mays plants was demonstrated. We also detected apoplast alkalinisation in the case of SG200 infection that was absent in the case of SG200Δlip3 infection. It is possible that Lip3 contributes to the pathogenesis of U. maydis by regulating apoplastic pH. However, the precise mechanism of pH regulation requires further investigation.