Insights into phosphorylation post-translational modifications in the proteome of C. albicans during oxidative stress response

Candida albicans is a common fungus of the human microbiota that can turn into an opportunistic pathogen under specific circumstances, particularly in individuals with a weakened immune system, leading to invasive candidiasis. During infection, C. albicans interacts with phagocytes, initiating the respiratory burst which produces reactive oxidant species (ROS) such as hydrogen peroxide (H₂O₂) in the intend to destroy the fungus. C. albicans survival depends on its ability to respond to this oxidative stress. Several well-known pathways have been involved in the response to the oxidative stress, such as the high osmolarity glycerol (HOG) MAPK pathway, that stands out as a primary mechanism for signalling the oxidative stress response (1), and Cap1 as the main transcription factor involved (2), finally promoting the expression of antioxidant proteins. To dig in other proteins and routes contributing, our group has done several proteomic studies, that have unveiled important proteins for C. albicans survival such as Prn1 and Oye32 (3, 4).

The present work describes a quantitative proteomic assay (DDA-MS) conducted with the wild-type strain SC5314 to characterize the changes in protein abundance and PTMs in response to a 200-min 10 mM H₂O₂ treatment. The assay identified approximately 1,900 phosphopeptides from 895 proteins that contained 2421 phosphosites. Gene Ontology (GO) term enrichment analysis of the proteins with significantly altered phosphorylation revealed an enrichment in proteins implicated in chromatin remodelling and in nucleotide binding proteins. Several protein kinases also exhibited significant changes in phosphorylation and, in concordance, npr1∆, kis1∆, and kin2∆ mutant strains displayed increased susceptibility to H₂O₂. However, Hog1 and Mkc1 MAPK signalling pathways, which sense H₂O₂, were deactivated after 200 min of H₂O₂ treatment. Among the nucleotide-binding GO Term enriched proteins we identified several transcription factors, such as Frc1 and Gzf3. As in the case of MAPK mutants, frz1∆ and gzf3∆ strains presented increased susceptibility to H₂O₂.

An integrative analysis of significant changes in both phosphopeptide and peptide abundances revealed that autophagy and cell cycle are strongly regulated after H₂O₂ treatment. Specifically, the decreased phosphorylation of Rim15 between Ser1183-Ser1190 might suggest that autophagy signalling pathway might be activated. In addition, cells undergo a G2 cell cycle arrest probably mediated by decreased phosphorylation of Cdc5 at Thr25.

Alonso-Monge R, et al. Eukaryot Cell. 2003Dai B Di, et al. FEBS. 2013Amador-García A, et al. mSystems. 2021Arribas V, et al. Antioxidants. 2024