Regulation of capsule biosynthesis in the fungal pathogen Cryptococcus neoformans

We are trying to understand how fungal pathogens of humans adapt to conditions inside the body. This knowledge may identify additional ways to diagnose and treat these important infections. The concentration of carbon dioxide is 150 fold higher inside our body than in the air surrounding us. When fungal pathogens of humans sense the higher CO2 inside the host, they initiate a series of development changes, which permits it to infect that host. Previously we identified the CO2 sensor responsible for these changes. Now we want to investigate precisely how this enzyme senses CO2. CO2 serves other functions important for the survival of fungal pathogens. For example it is a fundamental part of basic cellular metabolism. We also propose to study whether the CO2 sensing enzyme is related to fungal metabolism.

The ubiquitous fungal pathogen Cryptococcus neoformans infects the human central nervous system (CNS), causing meningoencephalitis that is usually fatal if untreated. AIDS is the predisposing factor in up to 90% of cryptococcal infections. Physiological concentrations of carbon dioxide have a strong impact on the expression of the C. neoformans capsule, the major virulence determinant of this fungus. CO2 also impacts on metabolism in this fungus. Capsule biosynthesis requires the enzyme adenylyl cyclase. The central hypothesis underpinning this work is that C. neoformans adenylyl cyclase (AC) is a CO2 chemosensor and amino acids of the catalytic core play an important role in sensing. Our overall aim is to understand the molecular basis of C. neoformans adenylyl cyclase activation by CO2/bicarbonate and determine how CO2 signalling and metabolism are integrated. In addition we will directly investigate the potential of the C. neoformans CO2/bicarbonate sensing pathway as a target for therapeutic intervention. To achieve our objectives we will mutagenize adenylyl cyclase to identify key amino acid residues required for CO2 chemosensing. We will also localize adenylyl cyclase inside the yeast cell, determine if it physically interacts with carbonic anhydrase and identify cellular carboxylases that depend on C. neoformans carbonic anhydrase activity.
The experiments proposed will provide new insights into the dynamics of CO2 sensing and may also translate into a larger drug-discovery programme in the future.