SummarySUT1 is a hypoxic gene encoding a nuclear protein that belongs to the Zn[II]2Cys-6 family. It has been shown that constitutive expression of SUT1 induces exogenous sterol uptake in aerobically growing Saccharomyces cerevisiae cells. A differential display approach was used to identify genes whose transcription is modified upon SUT1 induction. Within the promoter sequence of one of these genes, DAN1, we identified the region responsive to SUT1 and showed that it has a strong repressive activity when cloned in the vicinity of distinct promoters. Upon SUT1 constitutive expression in aerobiosis, the repression is released, allowing enhanced transcription of the reporter gene. We provide evidence that the repression is promoted by the Cyc8p(Ssn6p)±Tup1p co-repressor and that release of repression is the result of a physical interaction between Sut1p and Cyc8p. Moreover, genetic data suggest that complete derepression of the reporter gene requires a functional Cyc8p. In addition, we show that Sut1p is involved in the induction of hypoxic gene transcription when the cells are shifted from aerobiosis to anaerobiosis.

Summary.A novel aminopeptidase was purified by high performance liquid chromatography from a cytosoluble 100000 g extract of Candida albicans on the basis of its ability to cleave Larginine 7-amino-4-methylcoumarin. The purification factor was 36 and the yield was 20 %. The native enzyme had a mol. wt of 52 kDa as demonstrated by SDS-PAGE in the presence or absence of reducing conditions and exhibited an iso-electric point of 4.3. The aminopeptidase showed optimum activity at pH 7.2, a Michaelis constant of c. 50 p~ and a V, , , at 19 mM AMC released/min/mg of protein for L-Arg-AMC. This enzyme was shown to cleave at low affinity ~-1eucine-7-amino-4-methy1coumarin as demonstrated by the spectrofluorimetric method. The enzyme was strongly inhibited by specific metallo-enzyme inhibitors-EDTA and o-phenanthroline. Furthermore, there is evidence that a similar or identical enzyme occurs in other C. albicans clinical isolates and other Candidu spp.

Among potential virulence factors of Candida albicans, enzymes seem to play an important role. Many studies concern the secreted aspartic proteinases (saps), and the degradation of some components of the subendothelial extracellular matrix by the isoenzyme sap2 has been proved. Nevertheless, other proteolytic enzymes could be involved in the pathogenicity of the yeast. We studied the degradation of four constitutive proteins of the extracellular matrix: type I and IV collagens, laminin and fibronectin, by a 95-kDa metallopeptidase, localised in the cell wall of C. albicans. Each of these constituents was incubated with the purified enzyme and its degradation products analysed by an electrophoretic method. We observed that type I collagen and fibronectin were totally degraded by the enzyme whereas type IV collagen and laminin were only partially degraded. The C. albicans metallopeptidase may play a role in the degradation of the subendothelial extracellular matrix components. This enzyme could facilitate the migration of the yeast in the tissues after crossing the endothelial layer, allowing the fungal invasion of target organs.

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A myosin immunoanalogue was identified in conidia ofAspergillus fumigatus by Western blotting, indirect immunofluorescence assay, and gold immunoelectron microscopy with two different antimyosin antibodies. The distribution pattern of this protein was followed during the early stages of germination. A single 180-kDa polypeptide, detected predominantly in a cell envelope extract, was found to cross-react with monoclonal and polyclonal antibodies raised against vertebrate muscle myosin. Immunoelectron microscopy permitted precise localization of this polypeptide, indicating that myosin analogue was mainly distributed along the plasma membrane of resting and swollen conidia. In germinating conidia, indirect immunofluorescence microscopy revealed myosin analogue at the periphery of germ tubes, whereas actin appeared as dispersed punctate structures in the cytoplasm that were more concentrated at the site of germ tube emergence. A myosin ATPase inhibitor, butanedione monoxime, greatly reduced swelling and blocked germination. In contrast, when conidia were treated with cytochalasin B, an inhibitor of actin polymerization, swelling was not affected and germination was only partially reduced. Butanedione monoxime-treated conidia showed accumulation of cytoplasmic vesicles and did not achieve cell wall reorganization, unlike swollen conidia. Collectively, these results suggest an essential role for this myosin analogue in the deposition of cell wall components during germination of A. fumigatus conidia and therefore in host tissue colonization.