The cell surface of Candida albicans is enriched in highly glycosylated mannoproteins that are involved in the interaction with the host tissues. N glycosylation is a posttranslational modification that is initiated in the endoplasmic reticulum (ER), where the Glc 3 Man 9 GlcNAc 2 N-glycan is processed by ␣-glucosidases I and II and ␣1,2-mannosidase to generate Man 8 GlcNAc 2 . This N-oligosaccharide is then elaborated in the Golgi to form N-glycans with highly branched outer chains rich in mannose. In Saccharomyces cerevisiae, CWH41, ROT2, and MNS1 encode for ␣-glucosidase I, ␣-glucosidase II catalytic subunit, and ␣1,2-mannosidase, respectively. We disrupted the C. albicans CWH41, ROT2, and MNS1 homologs to determine the importance of N-oligosaccharide processing on the N-glycan outer-chain elongation and the host-fungus interaction. Yeast cells of Cacwh41⌬, Carot2⌬, and Camns1⌬ null mutants tended to aggregate, displayed reduced growth rates, had a lower content of cell wall phosphomannan and other changes in cell wall composition, underglycosylated ␤-N-acetylhexosaminidase, and had a constitutively activated PKC-Mkc1 cell wall integrity pathway. They were also attenuated in virulence in a murine model of systemic infection and stimulated an altered pro-and anti-inflammatory cytokine profile from human monocytes. Therefore, N-oligosaccharide processing by ER glycosidases is required for cell wall integrity and for host-fungus interactions.Candida albicans is an opportunistic fungal pathogen of humans that can cause superficial infections of the mucosa and, in the immunocompromised host, life-threatening systemic infections (10,52,53,61). The cell wall of C. albicans is the immediate point of contact between the fungus and host and therefore plays a key role in the host-fungus interaction. The cell wall is composed of an inner layer of chitin and ␤1,3-and ␤1,6-glucans and an outer layer that is rich in mannoproteins that accounts for 40% of the yeast form cell wall mass (39).

In fungi, phototropism, the induction of carotenogenesis and reproductive structures, and resetting of the circadian rhythm are controlled by blue light. Trichoderma atroviride, a fungus used in biological control, sporulates in a synchronized manner following a brief pulse of blue light. Due to its apparent simplicity, this response was chosen for pursuing photoreceptor isolation. Two genes were cloned, blue-light regulators 1 and 2 (blr-1 and blr-2), similar to the Neurospora crassa white-collar 1 and 2, respectively. The BLR-1 protein has all the characteristics of a blue-light photoreceptor, whereas the structure of the deduced BLR-2 protein suggests that it interacts with BLR-1 through PAS domains to form a complex. Disruption of the corresponding genes demonstrated that they are essential for blue-light-induced conidiation. blr-1 and blr-2 were also shown to be essential for the light-induced expression of the photolyase-encoding gene (phr-1). Mechanical injury of mycelia was found to trigger conidiation of T. atroviride, a response not described previously. This response was not altered in the mutants. A novel effect of both red and blue light on mycelial growth was found involving another light receptor, which is compensated by the BLR proteins.

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Candidiasis is a significant cause of invasive human mycosis with associated mortality rates that are equivalent to, or worse than, those cited for most cases of bacterial septicemia. As a result, considerable efforts are being made to understand how the fungus invades host cells and to identify new targets for fungal chemotherapy. This has led to an increasing interest in Candida glycobiology, with an emphasis on the identification of enzymes essential for glycoprotein and adhesion metabolism, and the role of N- and O-linked glycans in host recognition and virulence. Here, we refer to studies dealing with the identification and characterization of enzymes such as dolichol phosphate mannose synthase, dolichol phosphate glucose synthase and processing glycosidases and synthesis, structure and recognition of mannans and discuss recent findings in the context of Candida albicans pathogenesis.

A soluble alpha-mannosidase from Candida albicans was purified to homogeneity by sequential size exclusion, ion exchange, and affinity chromatographies in columns of Sepharose CL6B, DEAE Bio-Gel A, and Concanavalin A Sepharose 4B, respectively. Analytical electrophoresis of the purified preparation in 10% SDS-polyacrylamide gels stained with Coomassie blue revealed a single polypeptide of 43 kDa that was responsible for enzyme activity. The purified enzyme primarily trimmed Man(9)GlcNAc(2) to produce Man(8)GlcNAc(2) isomer B and mannose as a function of time of incubation up to 12 h at 37 degrees C. Prolonged incubation with the enzyme resulted in the accumulation after 24 h of other oligosaccharides corresponding to Man(7)GlcNAc(2) and probably Man(6)GlcNAc(2). These two products were also observed when Man(8)GlcNAc(2) isomer B instead of Man(9)GlcNAc(2) was used as substrate. Other oligosaccharides, such as Man(6)GlcNAc(2)-Asn, Man(5)GlcNAc(2)-Asn, and the alpha1,3- and alpha1,6-linked mannobiosides, were not hydrolyzed at all. These properties are consistent with an alpha1,2-mannosidase that may represent a new member of the glycosylhydrolase family 47.

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A soluble alpha-mannosidase from Candida albicans CAI-4 was purified by conventional methods of protein isolation. Analytical electrophoresis of the purified preparation revealed two polypeptides of 52 and 27 kDa, the former being responsible for enzyme activity. The purified, 52 kDa enzyme trimmed Man9GlcNAc2, producing Man8GlcNAc2 isomer B and mannose, and was inhibited preferentially by 1-deoxymannojirimycin. These properties are consistent with an endoplasmic reticulum-resident alpha1,2-mannosidase of the glycosyl hydrolase family 47. Moreover, a proteolytic activity responsible for converting the 52 kDa alpha-mannosidase into a polypeptide of 43 kDa retaining full enzyme activity, was demonstrated in membranes of ATCC 26555, but not in CAI-4 strain.