REGULATION OF PROTEIN KINASE A SUBUNITS EXPRESSION DURING THERMOTOLERANCE IN SACCHAROMYCES CEREVISIAE

GALELLO F; PORTELA P; CAÑONERO L; ROSSI S; ORTOLÁ MARTÍNEZ MC; OJEDA L

Modalidad virtual

Congreso; SAIB-SAIGE 2021; 2021

Organisms are constantly exposed to environmental changes. Variations in external conditions directly affect cellular homeostasis so that organisms have developed different strategies to overcome those situations. During stress a reprogramming of gene expression occurs, which involves not only the global inhibition of translation initiation but also the large-scale induction of stress‐responsive mRNAs through both transcriptional and translational regulation. Several mechanisms allow organisms to be prepared for recurring stressors. One of them is the anticipatory response or cellular memory, through which a current environment acts as a signal or input resulting in adaptation to future challenges. This response is known as ?acquired stress resistance?. S. cerevisiae PKA is composed of two catalytic subunits encoded by TPK1, TPK2 and TPK3 genes and two regulatory subunits encoded by BCY1 gene. The specificity of the PKA pathway depends on several factors as substrates specificity and interaction with anchoring proteins (AKAPs). Moreover, transcriptional regulation and PKA subunits expression level are also events involved in maintaining specificity. We have previously demonstrated that all PKA subunits share a negative autoregulatory mechanism mediated by PKA activity. TPK1 is the only PKA subunit that is transcriptionally upregulated during heat shock. To further understand the molecular process involved in regulating PKA subunits expression, the existence of a memory mechanism was evaluated. To this aim, cells were exposed to a scheme of two consecutive heat shocks: a 30-minute heat treatment at 37°C followed by a second 10-minute heat treatment at 45°C. TPKs and BCY1 promoter activities, mRNA and protein levels were assessed in mild log cells exposed to the thermotolerance scheme. We demonstrated that only TPK1 expression increases during thermotolerance. In yeast cells the final protein output of a genetic program is determined not only by transcription control and mRNA translation, but also by regulating mRNA localization and turnover rates. As part of this dynamic process, it has been proposed that components of the mRNA decay machinery can directly regulate transcription. The 5?-3? exonuclease Xrn1 is known as a key regulator of general mRNA decay pathways which also participates in transcription activation. We have previously demonstrated that Xrn1 affect TPK1 mRNA half-life. In order to evaluate the role of Xrn1 in TPK1 expression during thermotolerance we assessed TPK1 promoter activity and measured mRNA and protein levels in a mutant Δxrn1 strain. Our results showed that TPK1 expression is regulated during thermotolerance and that Xrn1 has an important role in this regulation.