Lodge Lab Research
Cryptococcus neoformans is a significant fungal pathogen, particularly in immunocompromised patients, that causes pulmonary infections and meningoencephalitis. It has been estimated that over 1,000,000 new cases of Cryptococcus occur, with over 650,000 deaths per year. The majority of these cases are in parts of Africa.
The fungal cell wall is an essential organelle, vital for maintaining cell integrity against various chemical, physical and biological stressors. This complex structure, immediately adjacent to the plasma membrane, serves several biological functions. Its cohesive structure, linked by covalent and hydrogen bonding or other types of interactions, provides great strength and stability. In this way, the cell wall prevents bursting that would be caused by internal turgor pressure and protects against external mechanical injury. The cell wall is the primary determinant of cell morphogenesis, and is constantly being remodeled to allow for growth, cell division and transitioning from yeast to branching hyphal forms to structures of sporulation. Since it is present at the cell surface, the cell wall interacts with the environment. In the case of pathogenic fungi, host defense systems are often directed against cell wall components. The cell wall has long been recognized as an ideal target for antifungal therapies because it is essential for fungal cell viability and many of its components are absent from the mammalian host. Fungal walls are primarily composed of polysaccharides and proteins (principally mannoproteins), along with other components, including lipids, pigments and inorganic salts, and have largely been studied in the model system Saccharomyces cerevisiae. Cryptococcal walls are considerably more complex, containing 1,3-glucan, 1,6-glucan, a-1,3-glucan, chitin, chitosan, melanin, as well as mannoproteins and other proteins.
It is important to understand the biosynthesis and regulation of fungal cell wall, especially because they are essential organelles that have been successfully used as targets for antifungal therapy.
We have demonstrated that chitosan is an abundant component of Cryptococcus cell walls, both during vegetative growth and infection of mammalian lungs, and that it is a critical component of the wall during infection. Three independent lines of mutants that have defects in chitosan production are all completely avirulent, and rapidly cleared from mammalian lungs. This data suggests that chitosan biosynthesis is a highly attractive anticryptococcal target. We have explored genes that contribute to chitosan production, and shown that only one (CHS3) of eight chitin synthases, and one (CSR2) of three putative chitin synthase regulators substantially impact chitosan levels. In addition, we have demonstrated that three of four potential polysaccharide deacetylases produce chitosan, and are currently investigating the role of each of these in chitosan production. Our current models of chitosan biosynthesis suggest that a complex of a specific chitin synthase, a regulator and a chitin deacetylase are located in a membrane and together, produce chitosan.
Multiple signal transduction pathways detect the environment and are used by fungi to respond to stresses. We have shown that the PKC1 pathway (aka the cell integrity pathway) has a role in response to antimicrobial defenses that are found in the human host, including oxidative and nitrosative stresses and chitinases. Our lab is exploring the role of upstream signaling molecules that detect and transmit these stresses, and determining mechanisms of how C. neoformans protects itself from these stresses.