Konopka, J. B. | Search bioRxiv

bioRxiv2021

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

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.

microbiology↗

bioRxiv2021

A conserved machinery underlies the synthesis of a chitosan layer in the Candida chlamydospore cell wall

The polysaccharide chitosan is found in the cell wall of specific cell types in a variety of fungal species where it contributes to stress resistance, or in pathogenic fungi, virulence. Under certain growth conditions, the pathogenic yeast Candida dubliniensis forms a cell type termed a chlamydosospore, which has an additional internal layer in its cell wall as compared to hyphal or yeast cell types. We report that this internal layer of the chlamydospore wall is rich in chitosan. The ascospore wall of Saccharomyces cerevisiae also has a distinct chitosan layer. As in S. cerevisiae, formation of the chitosan layer in the C. dubliniensis wall requires the chitin synthase CHS3 and the chitin deacetylase CDA2. In addition, three lipid droplet-localized proteins Rrt8, Srt1, and Mum3, identified in S. cerevisiae as important for chitosan layer assembly in the ascospore wall, are required for the formation of the chitosan layer of the chlamydospore wall in C. dubliniensis. These results reveal that a conserved machinery is required for the synthesis of a distinct chitosan layer in the walls of these two yeasts and may be generally important for incorporation of chitosan into fungal walls. ImportanceThe cell wall is the interface between the fungal cell and its environment and disruption of cell wall assembly is an effective strategy for antifungal therapies. Therefore, a detailed understanding of how cell walls form is critical to identify potential drug targets and develop therapeutic strategies. This work shows that a set of genes required for assembly of a chitosan layer in the cell wall of S. cerevisiae is also necessary for chitosan formation in a different cell type in a different yeast, C. dubliniensis. Because chitosan incorporation into the cell wall can be important for virulence, the conservation of this pathway suggests possible new targets for antifungals aimed at disrupting cell wall function.

cell biology↗