Identification and characterization of the Cryptococcus neoformans phosphomannose isomerase-encoding gene, MAN1, and its impact on pathogenicity

The polysaccharide capsule surrounding Cryptococcus neoformans comprises ma nose, xylose and glucuronic acid, of which mannose is the major constituent . The GDP-mannose biosynthesis pathway is highly conserved in fungi and con sists of three key enzymes: phosphomannose isomerase (PMI), phosphomannomut ase (PMM) and GDP-mannose pyrophosphorylase (GMP). The MAN1 gene, encoding for the PMI enzyme, was isolated and sequenced from C. neoformans, and a di sruption of the MAN1 gene was generated. One MAN1 disruption mutant, man1, which showed poor capsule formation, reduced polysaccharide secretion and m orphological abnormalities, was chosen for virulence studies. In both the r abbit and the mouse models of invasive cryptococcosis, man1 was shown to be severely impaired in its virulence, with complete elimination of the yeast from the host. A reconstituted strain of man1 was constructed using gene r eplacement at the native locus. The wild-type and reconstituted strains wer e significantly more virulent than the knock-out mutant in both animal mode ls. Our findings reveal that PMI activity is essential for the survival of C. neoformans in the host. The fact that the man1 mutant was not pathogenic suggests that blocking mannose synthesis could be fungicidal in the mammal ian host and thus an excellent target for antifungal drug development.