Identification of wall-specific antigens synthesized during germ tube formation by Candida albicans

Abstract

Walls of the two cellular forms (blastoconidia and mycelia) of Candida albicans ATCC 26555 were obtained from cells metabolically labeled (6-h pulse) with 'C-protein hydrolysate and [3H]threonine. Walls were purified by thorough washings with buffered and sodium dodecyl sulfate solutions and digested with Zymolyase 20T. The enzymatic treatment released four major high-molecular-weight mannoproteins (HMWM), with apparent molecular masses of 650, 500, 340, and 200 kilodaltons (HMWM-650, HMWM-500, HMWM-340, and HMWM-200, respectively), from yeast cells, whereas two high-molecular-mass mannoproteins (HMWM-260 and HMWM-180) were solubilized from mycelial cells. Some additional minor low-molecular-weight species were also detected in the enzymatic digests of walls from both types of cell. Single and dual pulse-chase experiments indicated that the HMWM-260 and HMWM-180 species reflect de novo synthesis of new proteins specific for the mycelia and do not represent a topological rearrangement of blastoconidium wall components. Monoclonal antibodies were raised against the HMWM-260 species (quantitatively the predominant component in the mycelial walls), and polyclonal rabbit antibodies were obtained against yeast or mycelial cell walls. Anti-mycelial cell wall polyclonal antibodies were adsorbed to whole killed blastoconidia to remove antibodies against common blastoconidium and mycelial wall antigens. Titration by enzyme-linked immunosorbent assay revealed that the monoclonal antibodies could recognize an epitope of the protein moiety of the HMWM-260 mannoprotein. Immunoblotting and immunofluorescence techniques using these monoclonal and polyclonal antibodies confirmed that the HMWM-260 and HMWM-180 species are specific components of the envelope of the mycelial cell walls.

Molecular organization of the cell wall ofCandida albicansand its relation to pathogenicity

“…A 47-kDa protein component of different strains of C. albicans was identified as an immunodominant antigen present in the cytoplasm and cell wall of both yeast and mycelial cells, but mainly exposed on the outer surface and unable to bind to Concanavalin A (Matthews et al, 1988). An interesting wall glycoprotein of 260 kDa, specific to the mycelial form of C. albicans, was isolated and monoclonal antibodies raised against its polypeptide moiety (Casanova et al, 1989). (It is relevant to recall that antigenicity of different wall glycoproteins has been associated not with the protein but with their mannan moiety.)…”

Identification and immunochemical characterization of a germ tube specific antigen of Candida albicans

“…Many kinds of mAb that recognize cell surface antigens of Candida species have previously been described. However, only a few of them have been shown to exhibit specificity to the hyphal phase of C. albicans [7,16].…”

“…Other specific epitopes of the mycelial phase have already been observed using rabbit immune sera that were pre-adsorbed with blastospores [8,10,11] and more recently by the use of metabolic radiolabelling or monoclonal antibodies [7,12]. Nevertheless, the distribution of these antigens among members of the Candida genera has not been fully investigated.…”

“…Evidence has been presented to support the pathogenic involvement of germ tubes [2,3], although the precise relationship between dimorphism and pathogenicity is not understood. Surface-specific molecules of C. albicans germ tubes have been identified using biochemical [4,5] and immunological approaches [6][7][8][9][10][11][12], some of which are involved in the adherence of the fungus to host cells or to prosthetic devices [13][14][15].…”

“…Monoclonal antibodies (mAbs) are powerful tools in the analysis of structural and functional 176 organization of cell surfaces; however to date few mAbs have been found that are strictly specific to the C. albicans mycelial phase [7,16]. It is interesting to note that the expression of some cell surface antigens of C. albicans was variable during morphogenesis of the cells [17][18][19][20].…”