Inactivation of Kex2p Diminishes the Virulence of Candida albicans

Abstract

Deletion of the kexin gene (KEX2) in Candida albicanshas a pleiotropic effect on phenotype and virulence due partly to a defect in the expression of two major virulence factors: the secretion of active aspartyl proteinases and the formation of hyphae. kex2/kex2 mutants are highly attenuated in a mouse systemic infection model and persist within cultured macrophages for at least 24 h without causing damage. Pathology is modest, with little disruption of kidney matrix. The infecting mutant cells are largely confined to glomeruli, and are aberrant in morphology. The complex phenotype of the deletion mutants reflects a role for kexin in a wide range of cellular processes. Taking advantage of the specificity of Kex2p cleavage, an algorithm we developed to scan the 9168 open reading frames in Assembly 6 of the C. albicans genome identified 147 potential substrates of Kex2p. These include all previously identified substrates, including eight secreted aspartyl proteinases, the exoglucanase Xog1p, the immunodominant antigen Mp65, and the adhesin Hwp1p. Other putative Kex2p substrates identified include several adhesins, cell wall proteins, and hydrolases previously not implicated in pathogenesis. Kexins also process fungal mating pheromones; a modification of the algorithm identified a putative mating pheromone with structural similarities to Saccharomyces cerevisiae ␣-factor.Serine proteinases of the kexin/subtilisin superfamily activate precursor forms of many exported eukaryotic proteins by specific endoproteolytic cleavage adjacent and C-terminal to dibasic residues. Members of the superfamily play a crucial role in a large and varied set of biological processes in animals, plants, and fungi. In metazoan cells, kexins participate in both the constitutive and regulated secretory pathways, with substrates including hormones, serum proteins, neuropeptides, cell surface receptors, components of the extracellular matrix (ECM), 1 and proteinases, which modify the ECM (1). Furin, a member of the mammalian kexin family, processes several ECM metalloproteinases requisite for the dissemination of cancer cells (2). Kexins have also been suborned to mediate the maturation of viral proteins and bacterial toxins (3). In fungi, kexins cycle between the outer face of the Golgi and the prevacuolar compartment where they process proteins involved in maintenance and remodeling of the cell wall (4, 5), proteins associated with the formation of aerial hyphae (6), ␣-type mating prepropheromones (7), killer toxins (7) , zymogens of secreted proteinases (8 -10), lipases (11), polysaccharide-degrading enzymes (12, 13), and themselves (14). Deleting the kexin of the yeast Yarrowia lipolytica abolishes the formation of hyphae (15). kex2-null mutants of the yeast Saccharomyces cerevisiae are viable but exhibit conditional morphological abnormalities (16), defective vacuolar proton-translocating V-ATPase activity (17), cold-sensitive growth (16), and a partial defect in meiosis (genome-www.stanford.edu/Saccharomyces). Genetic data indicate that ...

Destructin-1 is a collagen-degrading endopeptidase secreted by Pseudogymnoascus destructans , the causative agent of white-nose syndrome

“…This peptidase was shown to cleave antimicrobial peptides produced by frog skin, and is thus implicated in fungal survival and pathogenesis (43). Furthermore, the kexin gene in C. albicans encodes a subtilisin-type protease that is necessary for virulence because of its role in processing of proproteins (44). These observations suggest that the family of subtilisin-type peptidases play diverse roles as fungal virulence factors.…”

Identification and Characterization of a Candida albicans Mating Pheromone

“…1), although there is no amino acid sequence similarity between the S. cerevisiae mature ␣-factor and the predicted C. albicans ␣-factor. Newport et al (34) had previously noted that ORF6.4306 may encode an ␣-factor.…”

Yeast wall protein 1 of Candida albicans

“…In our assay of blastoconidial adhesion, Kex2p 2 strain CNA3-1 showed no increase in adhesion or biofilm formation over Ywp1p-replete strains (data not shown). Additional Kex2p substrates, which include Hwp1p and other adhesin-like proteins, have been tentatively identified in C. albicans (Newport et al, 2003), raising the possibility of a more expansive role for propeptides in regulating cellular adhesion.…”

“…However, the 12 kDa species that would result from such cleavage was not evident in the MS analysis of the purified Ywp1 propeptide. To investigate this further, Ywp1p was examined in CNA3-1, a kex2/kex2 strain of C. albicans (Newport & Agabian, 1997;Newport et al, 2003). Surprisingly, this strain generated a Ywp1 propeptide that was about 12 kDa (Figs 3b and 4) and the 11 kDa form was not detected.…”