Cryptococcus neoformans: synthesis of the polysaccharide capsule

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Cryptococcus is a fungus having a thick polysaccharide capsule, and infections (cryptococcosis) are an emerging problem in Canada. Especially individuals with a suppressed immune system are at risk. An outbreak of cryptococcosis in British Columbia demonstrated that strains of this fungus can be transferred world-wide and also infect individuals with a normal immune system. The capsule surrounding this fungus is a major virulence factor and can circumvent immune defenses and phagocytosis. The structures of C. neoformans capsule polysaccharides have been analyzed and the genome of several Cryptococcus species has been sequenced. However, the enzymes and mechanisms to assemble capsule polysaccharides remain to be elucidated. A number of capsule-associated genes are known that are required for the proper structure or size of the capsule but their specific functions are still unclear. This is a major knowledge gap in Canada and world-wide and we hope to fill this gap. The short term goal of this proposal is to use information from the capsule-associated genes and the capsule structures to identify the enzymes involved in capsule synthesis, to characterize these proteins and to determine the pathways of capsule synthesis in Cryptococcus. The sequence and control mechanisms of glycosyltransferases and O-acetyltransferases involved in capsule assembly will be studied in vitro and in fungal cells. This includes studies of protein structures and functions of specific amino acids in substrate binding and catalysis. The long term goal of this program is to acquire the knowledge and basic understanding of fungal polysaccharide synthesis and to train scientists in the field of fungal glycobiology and glycomics. Our previous expertise in the glycosyltransferases and glycosylation of mammalian cells and in bacteria is the basis for a successful outcome of this project. In addition, the P.I. has assembled a team of experts in Cryptococcus genetics and biology, synthesis of enzyme substrate analogs, protein structure determination and fungal biochemistry. These ground breaking discovery studies will establish a unique, needed expertise in fungal research in Canada. The research team will include at least two graduate and ten undergraduate students and three postdoctoral fellows. Our trainees will acquire a unique multidisciplinary expertise that can find applications in basic research and discovery, in biotechnology and drug development. The benefits are highly valuable for research laboratories and industry. It is essential to have this basic expertise in Canada.