Unfolded Protein Response governs an Alternative Splicing program conserved from healthy to malignant cells

The unfolded protein response (UPR) is a critical adaptive program triggered upon cellular stresses that profoundly reshapes the transcriptome and translatome. In the very first minutes of cellular stress, translation blockage, RNA decay and RNA granules formation prompt the synthesis of proteins essential to the stress response. Due to the dynamic nature of these processes and ribosome stalling, investigating translation upon stress has proven to be challenging; therefore, our understanding of these mechanisms and translatome rewiring upon stress remains limited. Here, we exploited O-Propargyl-puromycin (OPP) labelling of de novo peptides followed by LC-MS/MS to identify de novo proteins translated upon endoplasmic reticulum stress. Our approach combined to transcriptomic analyses revealed the synthesis of core splicing factor proteins and a profound reshaping of the splicing landscape upon ER stress. We identified a signature of 8 splicing events systematically occurring in eukaryotic cells exposed to ER stress. Using pharmacological, genetic, phosphoproteomic and sequencing approaches, we demonstrated that this specific ERsplice signature is driven by PERK activation and is dependent on splicing factors’ phosphorylation. Our study unveils a new role for ER stress and PERK in splicing regulation, and provide a molecular signature of ER stress through an ERsplice signature conserved from healthy to malignant tissues.