Abstract
Candida albicans is a prevalent fungal resident and opportunistic pathogen of humans, exhibiting a variety of ovoid and filamentous morphologies. Anchored within the cell wall of the ovoid yeast form of C. albicans is an abundant glycoprotein termed yeast wall protein 1 (Ywp1). Ywp1 has an antiadhesive effect that may facilitate yeast cell dispersal; it also contributes to the masking of the glucan matrix of the yeast cell wall, potentially providing shielding from recognition by the human immune system. Mature Ywp1 consists of an O-glycosylated core of 378 amino acids associated with an N-glycosylated propeptide that originates from an N-terminal segment of Ywp1. A tribasic (-RRR-) sequence in the immature Ywp1 polypeptide is separated by 8 amino acids from a dibasic (-KR-) sequence that is a canonical site for cleavage by the intracellular endopeptidase Kex2, and cleavage occurs at both of these sites to generate an 11 kilodalton (kDa) propeptide that remains strongly associated with the mature core of Ywp1. Previous studies demonstrated an absence of the 11 kDa propeptide in strains lacking Kex2, but the presence of lesser amounts of a 12 kDa propeptide ostensibly (and paradoxically) arising from cleavage at the dibasic site. Subsequent studies of wild type strains, however, suggested that post-secretion cleavages were carried out in vitro by acid proteases in unbuffered cultures to generate the 12 kDa propeptide. Here, intact and Gfp-tagged Ywp1 are utilized to show that neither of the two multibasic sites is normally cleaved in the absence of Kex2, but that uncleaved Ywp1 is still N-glycosylated and subsequently anchored to the cell wall. This furthers our understanding of the multistep cleavage of this highly conserved sequence, as well as the possible contributions of the cleaved propeptide to the maturation and functioning of Ywp1.