SUMMARY The Candida albicans cell wall maintains the structural integrity of the organism in addtion to providing a physical contact interface with the environment. The major components of the cell wall are fibrillar polysaccharides and proteins. The proteins of the cell wall are the focus of this review. Three classes of proteins are present in the candidal cell wall. One group of proteins attach to the cell wall via a glycophosphatidylinositol remnant or by an alkali-labile linkage. A second group of proteins with N-terminal signal sequences but no covalent attachment sequences are secreted by the classical secretory pathway. These proteins may end up in the cell wall or in the extracellular space. The third group of proteins lack a secretory signal, and the pathway(s) by which they become associated with the surface is unknown. Potential constituents of the first two classes have been predicted from analysis of genome sequences. Experimental analyses have identified members of all three classes. Some members of each class selected for consideration of confirmed or proposed function, phenotypic analysis of a mutant, and regulation by growth conditions and transcription factors are discussed in more detail.

SUMMARY The cell wall is essential to nearly every aspect of the biology and pathogenicity of Candida albicans. Although it was intially considered an almost inert cellular structure that protected the protoplast against osmotic offense, more recent studies have demonstrated that it is a dynamic organelle. The major components of the cell wall are glucan and chitin, which are associated with structural rigidity, and mannoproteins. The protein component, including both mannoprotein and nonmannoproteins, comprises some 40 or more moieties. Wall proteins may differ in their expression, secretion, or topological location within the wall structure. Proteins may be modified by glycosylation (primarily addition of mannose residues), phosphorylation, and ubiquitination. Among the secreted enzymes are those that are postulated to have substrates within the cell wall and those that find substrates in the extracellular environment. Cell wall proteins have been implicated in adhesion to host tissues and ligands. Fibrinogen, complement fragments, and several extracellular matrix components are among the host proteins bound by cell wall proteins. Proteins related to the hsp70 and hsp90 families of conserved stress proteins and some glycolytic enzyme proteins are also found in the cell wall, apparently as bona fide components. In addition, the expression of some proteins is associated with the morphological growth form of the fungus and may play a role in morphogenesis. Finally, surface mannoproteins are strong immunogens that trigger and modulate the host immune response during candidiasis.

Candida albicans is a commensal yeast normally present in small numbers in the oral flora of a large proportion of humans. Colonization of the oral cavity by C. albicans involves the acquisition and maintenance of a stable yeast population. Micro-organisms are continually being removed from the oral cavity by host clearance mechanisms, and so, in order to survive and inhabit this eco-system, C. albicans cells have to adhere and replicate. The oral cavity presents many niches for C. albicans colonization, and the yeast is able to adhere to a plethora of ligands. These include epithelial and bacterial cell-surface molecules, extracellular matrix proteins, and dental acrylic. In addition, saliva molecules, including basic proline-rich proteins, adsorbed to many oral surfaces promote C. albicans adherence. Several adhesins present in the C. albicans cell wall have now been partially characterized. Adherence involves lectin, protein-protein, and hydrophobic interactions. As C. albicans cells evade host defenses and colonize new environments by penetrating tissues, they are exposed to new adherence receptors and respond by expressing alternative adhesins. The relatively small number of commensal Candida cells in the oral flora raises the possibility that strategies can be devised to prevent oral colonization and infection. However, the variety of oral niches and the complex adherence mechanisms of the yeast mean that such a goal will remain elusive until more is known about the contribution of each mechanism to colonization.

No abstract

SUMMARY The cell wall of Candida albicans not only is the structure in which many biological functions essential for the fungal cells reside but also is a significant source of candidal antigens. The major cell wall components that elicit a response from the host immune system are proteins and glycoproteins, the latter being predominantly mannoproteins. Both the carbohydrate and protein moieties are able to trigger immune responses. Although cell-mediated immunity is often considered to be the most important line of defense against candidiasis, cell wall protein and glycoprotein components also elicit a potent humoral response from the host that may include some protective antibodies. Proteins and glycoproteins exposed at the most external layers of the wall structure are involved in several types of interactions of fungal cells with the exocellular environment. Thus, coating of fungal cells with host antibodies has the potential to influence profoundly the host-parasite interaction by affecting antibody-mediated functions such as opsonin-enhanced phagocytosis and blocking the binding activity of fungal adhesins for host ligands. In this review, the various members of the protein and glycoprotein fraction of the C. albicans cell wall that elicit an antibody response in vivo are examined. Although a number of proteins have been shown to stimulate an antibody response, for some of these species the response is not universal. On the other hand, some of the studies demonstrate that certain cell wall antigens and anti-cell wall antibodies may be the basis for developing specific and sensitive serologic tests for the diagnosis of candidasis, particularly the disseminated form. In addition, recent studies have focused on the potential for antibodies to cell wall protein determinants to protect the host against infection. Hence, a better understanding of the humoral response to cell wall antigens of C. albicans may provide the basis for the development of (i) effective procedures for the serodiagnosis of disseminated candidiasis and (ii) novel prophylactic (vaccination) and therapeutic strategies for the management of this type of infection.

Biofilms of Candida albicans are less susceptible to many antifungal drugs than are planktonic yeast cells. We investigated the contribution of cell density to biofilm phenotypic resistance. Planktonic yeast cells in RPMI 1640 were susceptible to azole-class drugs, amphotericin B, and caspofungin at 1 ؋ 10 3 cells/ml (standard conditions) using the XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide sodium salt] assay. As reported by others, as the cell concentration increased to 1 ؋ 10 8 cells/ml, resistance was observed with 10-to 20-fold-greater MICs. Biofilms that formed in microtiter plate wells, like high-density planktonic organisms, were resistant to drugs. When biofilms were resuspended before testing, phenotypic resistance remained, but organisms, when diluted to 1 ؋ 10 3 cells/ml, were susceptible. Drug-containing medium recovered from high-cell-density tests inhibited low-cell-density organisms. A fluconazole-resistant strain showed greater resistance at high planktonic cell density, in biofilm, and in resuspended biofilm than did low-density planktonic or biofilm organisms. A strain lacking drug efflux pumps CDR1, CDR2, and MDR1, while susceptible at a low azole concentration, was resistant at high cell density and in biofilm. A strain lacking CHK1 that fails to respond to the quorum-sensing molecule farnesol had the same response as did the wild type. FK506, reported to abrogate tolerance to azole drugs at low cell density, had no effect on tolerance at high cell density and in biofilm. These observations suggested that cell density has a role in the phenotypic resistance of biofilm, that neither the drug efflux pumps tested nor quorum sensing through Chk1p contributes to resistance, and that azole drug tolerance at high cell density differs mechanistically from tolerance at low cell density.

We have used a polyclonal antiserum t o cell wall proteins of Candida albicans to isolate several clones from a cDNA i l g t l l expression library. Affinitypurified antibody prepared to the fusion protein of one clone identified a 40 kDa moiety present in cell wall extracts from both morphologies of the organism. Indirect immunof luorescence demonstrated expression of this moiety at the C. albicans cell surface. Sequencing of a pBluescript II genomic clone identified with the cDNA clone revealed an open reading frame for a 417 amino acid protein. The nucleotide sequence showed significant homology with 3-phosphoglycerate kinase (PGK) genes, with 88%, 77% and 76% nucleotide homology with the PGK genes from Candida maltosa, Sacchammyces cere visiae and Kluyveromyces lactis, respectively. The deduced amino acid sequence was consistent with this identification of the sequence as PGKI of C. albicans. This finding was confirmed by a positive immunological response of a commercially available purified PGK from S. cerevisiae with the affinity-purified antibody against the fusion protein of the cDNA clone. The presence of PGK in the cell wall was confirmed by two additional methods.Cell wall proteins were biotinylated with a derivative that does not permeate the cell membrane to distinguish extracellular from cytosolic proteins. Biotinylated PGK was detected among the biotinylated proteins obtained following streptavidin affinity chromatography. lmmunoelectron microscopy revealed that the protein was present at the outer surface of the cell membrane and cell wall as well as expected in the cytoplasm. Northern blot analysis revealed that the gene transcript was present in C. albicans cells growing under different conditions, including different media, temperatures and morphologies. Most of the enzyme activity was found in the cytosol. Low enzymic activity was detected in intact cells but not in culture filtrates. These observations confirmed that PGK is a bona fide cell wall protein of C. albicans.

Candida albicans is a human commensal and opportunistic pathogen that participates in biofilm formation on host surfaces and also on medical devices. We used DIGE analysis to assess the cytoplasmic and non-covalently attached cell surface proteins in biofilm formed on polymethylmethacrylate and planktonic yeast cells and hyphae. Of the 1490 proteins spots from cytoplasmic and 580 protein spots from the surface extracts analyzed, 265 and 108 were differentially abundant respectively (≥1.5 fold, p<0.05). Differences of both greater and lesser abundance were found between biofilms and both planktonic conditions as well as between yeast cells and hyphae. The identity of 114 cytoplasmic and 80 surface protein spots determined represented 73 and 25 unique proteins respectively. Analyses showed that yeast cells differed most in cytoplasmic profiling while biofilms differed most in surface profiling. Several processes and functions were significantly affected by the differentially abundant cytoplasmic proteins. Particularly noted were many of the enzymes of respiratory and fermentative pentose and glucose metabolism, folate interconversions and proteins associated with oxidative and stress response functions, host response, and multi-organism interaction. The differential abundance of cytoplasmic and surface proteins demonstrated that sessile and planktonic organisms have a unique profile.