Integrative multi-omics profiling reveals cAMP-independent mechanisms regulating hyphal morphogenesis in Candida albicans

Abstract

Microbial pathogens grow in a wide range of different morphologies that provide distinct advantages for virulence. In the fungal pathogen Candida albicans, adenylyl cyclase (Cyr1) is thought to be a master regulator of the switch to invasive hyphal morphogenesis and biofilm formation. However, faster growing cyr1{Delta}/{Delta} pseudorevertant (PR) mutants were identified that form hyphae in the absence of cAMP. Isolation of additional PR mutants revealed that their improved growth was due to loss of one copy of BCY1, the negative regulatory subunit of protein kinase (A) from the left arm of chromosome 2. Furthermore, hyphal morphogenesis was improved in some of PR mutants by multigenic haploinsufficiency resulting from loss of large regions of the left arm of chromosome 2, including global transcriptional regulators. Interestingly, hyphal-associated genes were also induced in a manner that was independent of cAMP. This indicates that basal protein kinase A activity is an important prerequisite to induce hyphae, but activation of the cAMP pathway is not needed. Instead, phosphoproteomic analysis indicated that the Cdc28 cyclin-dependent kinase and the casein kinase 1 family member Yck2 play key roles in promoting polarized growth. In addition, integrating transcriptomic and proteomic data reveals hyphal stimuli induce increased production of key transcription factors that contribute to polarized morphogenesis. AUTHOR SUMMARYThe human fungal pathogen Candida albicans switches between budding and filamentous hyphal morphologies to gain advantages for virulence and survival in the host. Although adenylyl cyclase has been thought to be a master regulator that controls this switch, we identified C. albicans pseudorevertant mutants that grow better and form hyphae in the absence of adenylyl cyclase and cAMP. The mutant cells were also able to induce hyphal-associated genes in the absence of cAMP that are needed for virulence. Integrating information from different omics approaches identified cAMP-independent mechanisms that promote hyphal growth. This includes phosphoproteomic studies that revealed key roles for the Cdc28 cyclin-dependent kinase and casein kinase 1 in promoting hyphal growth. In addition, integrating transcriptomic and proteomic data revealed that post-transcriptional mechanisms regulate the levels of a set of key transcription factors that are important for hyphal induction, suggesting a novel form of translational regulation. These studies better define the pathways that stimulate C. albicans to switch from budding to hyphal growth, which is important for invasion into tissues, escape from the immune system, and biofilm formation.