Systematic capsule gene disruption reveals the central role of galactose metabolism on Cryptococcus neoformans virulence

Abstract

The polysaccharidic capsule is the main virulence factor of Cryptococcus neoformans. It primarily comprised of two polysaccharides: glucuronoxylomannan (GXM, 88% of the capsule mass) and galactoxylomannan (GalXM, 7% of the capsule mass). We constructed a large collection of mutant strains in which genes potentially involved in capsule biosynthesis were deleted. We used a new post-genomic approach to study the virulence of the strains. Primers specific for unique tags associated with the disruption cassette were used in a real-time PCR virulence assay to measure the fungal burden of each strain in different organs of mice in multi-infection experiments. With this very sensitive assay, we identified a putative UDP-glucose epimerase (Uge1p) and a putative UDP-galactose transporter (Ugt1p) essential for C. neoformans virulence. The uge1Delta and ugt1Delta strains are temperature sensitive and do not produce GalXM but synthesize a larger capsule. These mutant strains (GalXM negative, GXM positive) are not able to colonize the brain even at the first day of infection whereas GXM-negative strains (GalXM positive) can still colonize the brain, although less efficiently than the wild-type strain.