Cell surface proteome characterization of the Candida albicans DSE1 mutant. (c2011)

Abstract

The diploid fungus Candida albicans is a common opportunistic pathogen that normally colonizes human mucosal surfaces but can cause a wide variety of diseases in an immunocompromised host. C. albicans infections, known as candidiasis, range from mild superficial to severe systemic candidiasis in case of C. albicans dissemination in the blood stream. In a pathogen, the cell wall and cell wall proteins are important virulence factors and antigenic determinants since they are the first elements to contact the host. Thus, an in depth investigation of the cell wall structure may help reveal novel characteristics behind Candida’s virulence. Dse1 is a cell wall protein that has been previously characterized in our lab by homologous recombination of marker cassettes creating a heterozygous stain. The strain was found to be attenuated in virulence, less resistant to cell surface disrupting agents such as calcofluour white, delayed in adhesion to human epithelial cells and deficient in biofilm formation. The current study aims to investigate the cell surface proteome to determine differences in protein expression patterns that might explain the above-mentioned phenotypes. As such the amount of total cell wall proteins in the mutant was found to be lower than in the wild type under filamentous conditions. Furthermore chitin content in the mutant was found to be reduced by 16%, possibly explaining the decreased resistance to calcofluour white, a cell wall disrupting agent that interferes with chitin microfibril assembly. Extracted proteins were then digested with trypsin and analyzed using MALDI-TOF MS, generating a mass spectrometric profile for each strain, each under different growth conditions. These different profiles were compared, and unique peaks for each strain were entered into MS-Fit search engine, compared against a Candida database, and identified by peptide mass fingerprinting (PMF). As such the mutant was shown to lack the chitin biosynthesis protein CHS5, possibly explaining the decrease in chitin biosynthesis. PMF analysis also suggested a mutant-specific expression of glucoamylase 1, a cell wall glycoprotein involved in carbohydrate metabolism and cell wall degradation, changing the cell wall organization and decreasing biofilm formation, and a decrease in lipase protein expression in the mutant, resulting in reduced virulence.