Project Portugal 2030

Summary

All living organisms face challenging environments and rely on adaptation mechanisms to survive and thrive. Immediate adaptation relies on signalling pathways that trigger transcriptional responses modulating the organism proteome.In turn, long-term adaptations include advantageous genetic mutations that can be passed to subsequent generations, altering the proteome quantitatively and heritably, often leading to visible phenotypic changes.Protein biosynthesis errors during mRNA translation (mistranslation) have emerged as alternative adaptive mechanisms. Unlike genetic mutations, translational mutations occur randomly in a small percentage of polypeptides, affecting numerous proteins or the entire proteome often in a transient manner and without heritability. Mistranslation yields statistical proteins, comprising both wild type and mutated polypeptides, with the latter representing a small fraction of the total polypeptides for each protein. We developed a new proteogenomic pipeline for mistranslation analysis in C.albicans. Unlike previous reporter systems that are limited to one protein and to a single codon, our approach analyses translation errors at the proteomic scale and without the need of reporter integration, facilitating the analysis of clinical strains for the first time.Having this tool, we will push the analysis of translation fidelity further, to achieve our main goal: ASSESS THE CLINICAL IMPLICATIONS OF PROTEIN BIOSYNTHESIS ERRORS IN C.albicans AND IDENTIFY MISTRANSLATION MARKERS TO PREDICT THE PATHOGENIC POTENTIAL OF CLINICAL STRAINS. The CANDIDATE project aims to accomplish the following objectives: A) DETERMINE THE CORRELATION BETWEEN TRANSLATION FIDELITY AND NICHE LOCALIZATION OF C.albicans. Task 1: Proteogenomic analysis of C.albicans clinical strains across diverse human body niches. B) INVESTIGATE WHETHER STRESS-INDUCED MISTRANSLATION IS DIRECTED TOWARDS SPECIFIC PROTEINS, PROVIDING POTENTIAL MARKERS FOR PATHOGENICITY PREDICTION. Task 2: Identification and validation of mistranslation markers in clinical isolates of C.albicans using targeted-MS: new insights on stress responses. C) EXAMINE THE INTERPLAY BETWEEN TRANSLATION FIDELITY AND HOST-PATHOGEN INTERACTIONS. Task 3: Assessing the impact of host-pathogen interactions on translation fidelity: in vitro assays. Task4: Exploring mistranslation dynamics in murine commensalism and Galleria mellonella infection: in vivo assays. The transcriptome and proteome of C.albicans varies strongly with the environment suggesting a correlation with the infection source. While mistranslation can modulate the structure, function, and activity of proteins, which may impact the fitness and virulence of C.albicans, its role in niche localization remains unclear.This project integrates proteomic analysis (Task1) with genomic and phenotypic data (Supplemental figures), to accurately assess translation-derived mutations and their putative correlation with infection sources. Proteins with misincorporations will serve as mistranslation markers for stress response analysis (Task2). New MS methods will be optimized, facilitating screening of clinical isolates in several conditions. Finally, the project investigates C.albicans´ use of mistranslation in establishing as a gut commensal or causing disseminated infections (Tasks3-4). Overall, this project offers insights into C.albicans' proteomic landscape and provides tools to evaluate protein biosynthesis errors in various microorganisms.