Tandem Mass Spectrometric Cell Wall Proteome Profiling of a Candida albicans hwp2 Mutant Strain and Fluconazole Resistant Strains. (c2017)

Abstract

Candida albicans, often recognized as a harmless opportunistic yeast, is ordinarily present as part of the normal gut flora and detected in the oral cavities as well as the GI tracts of around forty to sixty percent of human adults. However benign C. albicans colonization can turn pathogenic and cause a variety of infections ranging from mild to severe in immune-compromised individuals. In addition and due to improper use of antifungal drug therapy, resistance rates to antifungals specifically fluconazole is on the rise. In a pathogen the cell wall and cell surface proteins are major antigenic determinants as they are the primary structure that contacts the host. Cell surface proteins perform a variety of functions that are necessary for virulence such as adhesion, host degradation, resistance to oxidative stress, and drug resistance amongst others. Our lab had previously characterized Hwp2, a Candida albicans cell wall adhesin that was shown to play a major role in the cell wall architecture and function since cells lacking Hwp2 were found to be deficient in chitin deposition, filamentation, adhesion and invasive growth, virulence, and resistance to oxidative stress. In this study, we utilized tandem mass spectrometry coupled with a bioinformatics approach to differentially profile the cell wall proteome of a wild type strain compared to an hwp2 null mutant to determine key differentially expressed proteins. Many such proteins identified exclusively in the wild type go a long way in explaining the above mentioned phenotypes. These proteins include known virulence factors such as members of the SAP (secreted aspartyl proteases) family of proteins that include Sap4, Sap5, and Sap10 as well as several lipases involved in host degradation. We also identified members of the PGA Protein family (Pga28, Pga32, Pga41 and Pga50) that function in adhesion, Cht2 a chitinase involved in chitin remodeling, and several proteins that function in promoting filamentation (Phr1, Mts1, Rbr1). In parallel, we applied a similar approach to generate a differential profile of C. albicans fluconazole resistant strains and successfully identified proteins such as Sec4, a protein involved in resistance to drugs, as well as Pil1, a protein known to be involved in echinocandin resistance, suggesting a novel mechanism whereby resistance to fluconazole and caspofungin may be linked.