ENDOPLASMIC RETICULUM STRESS-INDUCED CALCIUM INCREASE ACTIVATES PERK PATHWAY

JUAN VILCHEZ ARUANI; MACARENA FERNANDEZ; JUAN C. DE BATTISTA; MARIANA BOLLO

Congreso; Congreso SAIB SAMIGE 2021; 2021

The Endoplasmic Reticulum (ER) is a dynamic organelle where are performed numerous functions, such as: storage and release of Ca2+ , lipid synthesis, folding and post-transductional modifications of proteins. All these processes are interconnected and can be performed only if the Ca2+ concentration in the lumen is optimal, this Ca2+ acts as a key messenger.When the loaded of newly synthesized proteins exceeds the folding and/or processing capacity in the organelle, the ER enters into stress condition. To restore the homeostasis, the organelle activates a signaling transduction pathway collectively termed the Unfolded Protein Response (UPR). (PKR)-like-ER kinase (PERK) is an early stress response ER-transmembrane proteinthat is generally inactive due to its association with the chaperone BiP. During ER stress, BiP is tritrated by the unfolded protein, leading PERK activation and phosphorylation of eukaryotic initiation factor-2 alpha (eIF2 ), which attenuates protein synthesis. We demonstrated that calcineurin-A/B(CN-A/B), an heterotrimeric Ca2+ protein, directly associates withPERK, increasing its auto-phosphorylation and significantly enhancing inhibition of protein translation. It has also been observed that the β isoform of subunit A of CN (CN-Aβ) in astrocytes has an important PERK-dependent cytoprotective effect.Although the involvement of Ca2+ signaling in a multitude of cellular functions has been well documented, little is known about its role in restoring homeostasis, once UPR is activated. Recently, we described an active ER Ca2+ release through the translocon during acute phase of UPR. The translocon is a protein complex formed by a heterotrimeric core (Sec61α, β, γ).Sec61α, extends on the ER lipid bilayer and forms the channel pore. Here, we evaluated, in astrocytes, the dependence of Ca2+ on PERK activation by immunocytochemistry as well PERK/CN interaction and eIF2α phosphorylation, after induces stress and pharmacologically modify translocon activity. Moreover, we demonstrated that, using a cell line deficient in all isoformsof IP3 receptor and by performing blue native PAGE followed by two-dimensional gel, PERK forms a macromolecular complex with the translocon (Sec61α) and CN, under stress condition. Overall, these data strongly suggest that PERK is activated by cytosolic Ca2+ increase originated through the translocon during acute phase of UPR.