Abstract 16941: The Role of the PERK-Mediated Unfolded Protein Response Pathway in the Regulation of Cardiac Sodium Channel During Human Heart Failure

Abstract

Background: The unfolded protein response (UPR) is a series of interrelated signaling pathways that occur when the endoplasmic reticulum (ER) experiences excess secretory load, accumulates misfolded proteins, or is subject to other pathological conditions. UPR acts to attenuating general protein synthesis, induces the expression of ER chaperone proteins, and enhances the degradation of misfolded proteins. In published work, we have shown that heart failure (HF) increases alternative splicing of the SCN5A gene (encoding cardiac Na + channel), generating mRNA variants E28C and E28D encoding truncated, nonfunctional Na + channel. The presence of these variants causes a dominant negative downregulation of the wild-type SCN5A mRNA and Na + current to a sufficient extent to be arrhythmogenic. We tested whether PERK-mediated UPR contributed to the dominant negative effect on Na + current when truncated Na + channel mRNA variants are present in HF. Methods: The correlation of expression changes among PERK, major chaperon proteins (calnexin, CHOP) and SCN5A were analyzed by both qPCR and Western Blot in human HF tissue samples and human embryonic stem cell-derived cardiomyocytes (hESC-CMs). Hypoxia and Ang II were used as inducers of abnormal splicing since they have been shown to mediate some of the pathological consequences of HF. hESC-CMs were divided into six experimental groups: normoxic, 1% O 2 hypoxia-treated, 100 nmol/L Ang II-treated, E28C overexpressed, E28D overexpressed, and empty vector control. E28C and E28D constructs were transduced to overexpress the truncated proteins. Results: The expressions of major UPR components (PERK, calnexin, CHOP) were increased in HF tissues, and cardiac Na + channels were downregulated. In hESC-CMs, induction of SCN5A variants E28C and E28D with Ang II or hypoxia as well as expression of exogenous variants could induce major UPR components (PERK, calnexin, CHOP). Finally, downregulation of PERK prevented the loss of full-length SCN5A mRNA abundance with these stimuli. Conclusions: SCN5A variants could induce the expression of major UPR components and could induce PERK-mediated Na + channel downregulation. The results indicate that the UPR contributes to Na + channel regulation during human HF.