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.
