TheCryptococcus neoformansSTRIPAK complex controls genome stability, sexual development, and virulence
Authors: Peterson Patricia P. , Choi Jin-Tae , Fu Ci , Cowen Leah E. , Sun Sheng , Bahn Yong-Sun , Heitman Joseph 
Published: August 2024

AbstractThe eukaryotic serine/threonine protein phosphatase PP2A is a heterotrimeric enzyme composed of a scaffold A subunit, a regulatory B subunit, and a catalytic C subunit. Of the four known B subunits, the B’’’ subunit (known as striatin) interacts with the multi-protein striatin-interacting phosphatase and kinase (STRIPAK) complex. Orthologs of STRIPAK components were identified inCryptococcus neoformans, namely PP2AA/Tpd3, PP2AC/Pph22, PP2AB/Far8, STRIP/Far11, SLMAP/Far9, and Mob3. Structural modeling, protein domain analysis, and detected protein-protein interactions suggestC. neoformansSTRIPAK is assembled similarly to the human and fungal orthologs. Here, STRIPAK components Pph22, Far8, and Mob3 were functionally characterized. Whole-genome sequencing revealed that mutations in STRIPAK complex subunits lead to increased segmental and chromosomal aneuploidy, suggesting STRIPAK functions in maintaining genome stability. We demonstrate thatPPH22is a haploinsufficient gene: heterozygousPPH22/pph22Δ mutant diploid strains exhibit defects in hyphal growth and sporulation and have a significant fitness disadvantage when grown in competition against a wild-type diploid. Deletion mutantspph22Δ, far8Δ, andmob3Δ exhibit defects in mating and sexual differentiation, including impaired hyphae, basidia, and basidiospore production. Loss of eitherPPH22orFAR8in a haploid background leads to growth defects at 30⁰C, severely reduced growth at elevated temperature, abnormal cell morphology, and impaired virulence. Additionally,pph22Δ strains frequently accumulate suppressor mutations that result in overexpression of another putative PP2A catalytic subunit,PPG1.Thepph22Δ andfar8Δ mutants are also unable to grow in the presence of the calcineurin inhibitors cyclosporine A or FK506, and thus these mutations are synthetically lethal with loss of calcineurin activity. Conversely,mob3Δ mutants display increased thermotolerance, capsule production, and melanization, and are hypervirulent in a murine infection model. Taken together, these findings reveal that theC. neoformansSTRIPAK complex plays an important role in genome stability, vegetative growth, sexual development, and virulence in this prominent human fungal pathogen.Author summaryThis study focused on a highly conserved protein signaling complex known as STRIPAK, which is important for various developmental processes in fungi and humans. By investigating the functions of this complex inCryptococcus neoformans, it was discovered to play crucial roles in maintaining genome stability, sexual development, and pathogenesis. In particular, mutations in the genes encoding two subunits of the STRIPAK complex were found to lead to significant defects, including abnormal growth and cell morphology, compromised stress response, and impaired virulence. Interestingly, mutation of a third STRIPAK complex subunit resulted in hypervirulence, characterized by increased thermotolerance, enhanced production of melanin pigment and polysaccharide capsule, and reduced survival in infected animals. Our findings reveal that the STRIPAK complex is an important regulator of growth and virulence ofC. neoformans, highlighting its potential as a target for therapies aimed at combating fungal infections. This work furthers understanding of how the STRIPAK complex functions inCryptococcus, and also in other organisms including humans, where related protein phosphatase complexes govern key cellular processes.