Sap6, a secreted aspartyl proteinase, participates in maintenance the cell surface integrity of Candida albicans

Background: The polymorphic species Candida albicans is the major cause of candidiasis in humans. The secreted aspartyl proteinases (Saps) of C. albicans, encoded by a family of 10 SAP genes, have been investigated as the virulent factors during candidiasis. However, the biological functions of most Sap proteins are still uncertain. In this study, we applied co-culture system of C. albicans and THP-1 human monocytes to explore the pathogenic roles and biological functions of Sap proteinases. Results: After 1 hr of co-culture of C. albicans strains and THP-1 human monocytes at 37 degrees C, more than 60% of the THP-1-engulfed wild type and Delta sap5 Candida cells were developing long hyphae. However, about 50% of THP-1-engulfed Delta sap6 Candida cells were generating short hyphae, and more dead Candida cells were found in Delta sap6 strain that was ingested by THP-1 cells (about 15% in Delta sap6 strain vs. 2 similar to 2.5% in SC5314 and Delta sap5 strains). The immunofluorescence staining demonstrated that the Sap6 is the major hyphal tip located Sap protein under THP-1 phagocytosis. The sap6-deleted strains (Delta sap6, Delta sap4/6, and Delta sap5/6) appeared slower growth on Congo red containing solid medium at 25 degrees C, and the growth defect was exacerbated when cultured at 37 degrees C in Congo red or SDS containing medium. In addition, more proteins were secreted from Delta sap6 strain and the beta-mercaptoethanol (beta-ME) extractable surface proteins from Delta sap6 mutant were more abundant than that of extracted from wild type strain, which included the plasma membrane protein (Pma1p), the ER-chaperone protein (Kar2p), the protein transport-related protein (Arf1p), the cytoskeleton protein (Act1), and the mitochondrial outer membrane protein (porin 1). Moreover, the cell surface accessibility was increased in sap6-deleted strains. Conclusion: From these results, we speculated that the cell surface constitution of C. albicans Delta sap6 strain was defect. This may cause the more accessible of beta-ME to disulfide-bridged cell surface components and may weaken the resistance of Delta sap6 strain encountering phagocytosis of THP-1 cells. Sap6 protein displays a significant function involving in maintenance the cell surface integrity.