The Candida albicans Cdk8-dependent phosphoproteome reveals repression of hyphal growth through a Flo8-dependent pathway
| Autor: | Zhongle Liu, Lawrence C. Myers, Jeffrey M. Hollomon, Smith Ak, Deborah A. Hogan, Nicole P. Jenkins, Scott F. Rusin, Katja Koeppen, Arminja N. Kettenbach |
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| Rok vydání: | 2021 |
| Předmět: |
Proteomics
Hyphal growth Cancer Research Mutant Gene Expression Yeast and Fungal Models QH426-470 Pathology and Laboratory Medicine Biochemistry Loss of Function Mutation Gene Expression Regulation Fungal Candida albicans Medicine and Health Sciences Protein phosphorylation Post-Translational Modification Phosphorylation Genetics (clinical) Candida Fungal Pathogens biology Organic Compounds Chemistry Kinase Transcriptional Control Monosaccharides Eukaryota Cell biology Phenotypes Experimental Organism Systems Medical Microbiology Physical Sciences Saccharomyces Cerevisiae Pathogens Research Article Carbohydrates Hyphae Mycology Research and Analysis Methods Microbiology Fungal Proteins Saccharomyces Model Organisms Gene Types DNA-binding proteins Genetics Gene Regulation Kinase activity Molecular Biology Microbial Pathogens Transcription factor Ecology Evolution Behavior and Systematics Gene Expression Profiling Organic Chemistry Organisms Fungi Chemical Compounds Biology and Life Sciences Proteins Cyclin-Dependent Kinase 8 biology.organism_classification Yeast Regulatory Proteins Glucose Purines Animal Studies Regulator Genes Cyclin-dependent kinase 8 Transcription Factors |
| Zdroj: | PLoS Genetics PLoS Genetics, Vol 18, Iss 1, p e1009622 (2022) |
| Popis: | Ssn3, also known as Cdk8, is a member of the four protein Cdk8 submodule within the multi-subunit Mediator complex involved in the co-regulation of transcription. In Candida albicans, the loss of Ssn3 kinase activity affects multiple phenotypes including cellular morphology, metabolism, nutrient acquisition, immune cell interactions, and drug resistance. In these studies, we generated a strain in which Ssn3 was replaced with a functional variant of Ssn3 that can be rapidly and selectively inhibited by the ATP analog 3-MB-PP1. Consistent with ssn3 null mutant and kinase dead phenotypes, inhibition of Ssn3 kinase activity promoted hypha formation. Furthermore, the increased expression of hypha-specific genes was the strongest transcriptional signal upon inhibition of Ssn3 in transcriptomics analyses. Rapid inactivation of Ssn3 was used for phosphoproteomic studies performed to identify Ssn3 kinase substrates associated with filamentation potential. Both previously validated and novel Ssn3 targets were identified. Protein phosphorylation sites that were reduced specifically upon Ssn3 inhibition included two sites in Flo8 which is a transcription factor known to positively regulate C. albicans morphology. Mutation of the two Flo8 phosphosites (threonine 589 and serine 620) was sufficient to increase Flo8-HA levels and Flo8 dependent transcriptional and morphological changes, suggesting that Ssn3 kinase activity negatively regulates Flo8.Under embedded conditions, when ssn3Δ/Δ and efg1Δ/Δ mutants were hyperfilamentous, FLO8 was essential for hypha formation. Previous work has also shown that loss of Ssn3 activity leads to increased alkalinization of medium with amino acids. Here, we show that the ssn3Δ/Δ medium alkalinization phenotype, which is dependent on STP2, a transcription factor involved in amino acid utilization, also requires FLO8 and EFG1. Together, these data show that Ssn3 activity can modulate Flo8 and its direct and indirect interactions in different ways, and underscores the potential importance of considering Ssn3 function in the control of transcription factor activities. Author summary In Candida albicans, Ssn3 kinase activity co-regulates the transcription of numerous genes involved in hyphal growth, metabolism and nutrient acquisition, immune cell interactions, and drug resistance. Using a strain in which Ssn3 could be rapidly and selectively inhibited, we identified both known and novel Ssn3 targets. We identified two phosphosites in Flo8, a regulator of morphology and virulence, that were shown to negatively regulate Flo8 levels and activity. The data and tools presented here will enable a better understanding of how Ssn3 impacts transcriptional and post-transcriptional regulation in order to coordinate processes during physiological and morphological transitions as well as during steady state growth. |
| Databáze: | OpenAIRE |
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