PERK REGULATION VIA ALTERNATIVE SPLICING PARTICIPATES IN UPR REGULATION IN NEURONS FOLLOWING ER STRESS

One of the main hallmarks of most neurodegenerative disorders is the presence of misfolded proteins, leading to activation of the unfolded protein response (UPR). While UPR activation is essential to restore cellular proteostasis, its dysregulation leads to cellular dysfunction and has been reported in most neurodegenerative disorders. PERK is one of the ER-resident kinases comprising the UPR and, it is well-characterized that, in response to ER stress, it activates via autophosphorylation leading to downregulation of translation through phosphorylation of eIF2α. However, little is known about regulation of Eif2ak3, PERK coding gene, at the RNA level. Therefore, this project aims to investigate whether RNA regulation plays a role in PERK function during ER stress using murine primary neurons as an in vitro model. Our results show that PERK is indeed regulated at a transcriptional level, both in a development- and stressor-dependent manner. Moreover, we identify several previously unknown Eif2ak3 mRNA variants generated via alternative splicing. Sequence analysis predicts that the new identified Eif2ak3 mRNA variants lead to the activation of the nonsense-mediated decay pathway due to the presence of premature stop codons. Importantly, we describe that antisense oligonucleotide-mediated knockdown of one Eif2ak3 alternative transcript is enough to partially prevent ER stress-induced eIF2α phosphorylation, without altering PERK protein levels. Overall, our results suggest that alternative splicing contributes to the fine-tunned PERK regulation during ER stress and UPR activation in neurons with potential implications for most neurodegeneration disorders involving UPR dysfunction.