A fungal lytic polysaccharide monooxygenase is required for cell wall integrity, thermotolerance, and virulence of the fungal human pathogen Cryptococcus neoformans

Abstract

Fungi often adapt to environmental stress by altering their size, shape, or rate of cell division. These morphological changes require reorganization of the cell wall, a structural feature external to the cell membrane composed of highly interconnected polysaccharides and glycoproteins. Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes that are typically secreted into the extracellular space to catalyze initial oxidative steps in the degradation of complex biopolymers such as chitin and cellulose. However, their roles in modifying endogenous microbial carbohydrates are poorly characterized. The CEL1 gene in the human fungal pathogen Cryptococcus neoformans (Cn) is predicted by sequence homology to encode an LPMO of the AA9 enzyme family. The CEL1 gene is induced by host physiological pH and temperature, and it is primarily localized to the fungal cell wall. Targeted mutation of the CEL1 gene revealed that it is required for the expression of stress response phenotypes, including thermotolerance, cell wall integrity, and efficient cell cycle progression. Accordingly, a cel1{Delta} deletion mutant was avirulent in two models of C. neoformans infection. Therefore, in contrast to LPMO activity in other microorganisms that primarily targets exogenous polysaccharides, these data suggest that CnCel1 promotes intrinsic fungal cell wall remodeling events required for efficient adaptation to the host environment. Author summaryFungi need to adapt quickly to environmental stresses to thrive. The fungal cell wall, which supplies support and integrity to the cell, is an essential compartment to react and interact with the surrounding environment. Rapid changes within the carbohydrate composition and architecture occur in response to environmental stresses. Lytic polysaccharide monooxygenases (LPMOs) are mononuclear copper-enzymes, secreted by microbes to assist in the first steps of remodeling and degrading complex and recalcitrant carbohydrates. In this study we explore the role of the putative AA9 family LPMO CnCel1 for growth and virulence of the fungal human pathogen Cryptococcus neoformans. The CEL1 gene is highly up-regulated in presence of host stresses and a cel1{Delta} mutant strain is avirulent in a murine model of infection. Downstream analysis of virulence-associated phenotypes identified the CEL1 gene to be required for thermotolerance as well as cell wall integrity, and efficient cell cycle progression in presence of host-mimicking stresses. Based upon those findings we propose that CnCel1 likely promotes intrinsic fungal cell wall remodeling events essential for adaptation to the host environment.