Protein phosphatase 2C domain contributes to the pathobiological function of adenylyl cyclase in Cryptococcus neoformans

Adenylyl cyclase produces cyclic adenosine monophosphate, a signalling molecule that controls fungal development and virulence. Fungal adenylyl cyclases are distinguished by the presence of a conserved protein phosphatase 2C domain, whose function remains unknown. Here we show that the protein phosphatase 2C domain of Cac1, the adenylyl cyclase of the human pathogen Cryptococcus neoformans, has an unusual structure but functions as a metal-dependent serine/threonine phosphatase. Deletion of this domain or the adenylyl cyclase catalytic domain demonstrated that the protein phosphatase 2C domain is required for full Cac1 activity, influencing melanin and capsule synthesis, sexual differentiation, titan cell formation, and cell wall integrity. Notably, loss of the protein phosphatase 2C domain induces type 2 helper T-cell-biased immunity and extensive pulmonary damage yet does not cause mortality in mice. Integrated transcriptomic and phosphoproteomic analyses further revealed shared and domain-specific signalling outputs. Together, these findings define a pivotal role for the adenylyl cyclase-linked protein phosphatase 2C domain in C. neoformans pathobiology. Using biochemical, structural, transcriptomic and phosphoproteomic analyses, this study characterises the PP2C domain of Cac1 in Cryptococcus neoformans. The domain is an active Ser/Thr phosphatase required for Cac1 signalling and pathobiology.