Adam M Noyes1, Anyu Zhou1, Ge Gao2, Lianzhi Gu2, Sharlene Day3, J Andrew Wasserstrom4, Samuel C Dudley5. 1. Department of Cardiology, Warren Alpert School of Medicine at Brown University, Providence, RI, United States. 2. Cardiology Section, University of Illinois at Chicago, Chicago, IL, United States. 3. Department of Internal Medicine, University of Michigan Medical Center, Ann Arbor, MI, United States. 4. Feinberg Cardiovascular Research Institute, Feinberg School of Medicine, Northwestern University, Chicago, IL, United States. 5. Department of Cardiology, Warren Alpert School of Medicine at Brown University, Providence, RI, United States. Electronic address: samuel_dudley@brown.edu.
Abstract
BACKGROUND: Our previous studies showed that in ischemic and nonischemic heart failure (HF), the voltage-gated cardiac Na+ channel α subunit (SCN5A) mRNA is abnormally spliced to produce two truncated transcript variants (E28C and D) that activate the unfolded protein response (UPR). We tested whether SCN5A post-transcriptional regulation was abnormal in hypertrophic cardiomyopathy (HCM). MATERIAL AND METHODS: Human heart tissue was obtained from HCM patients. The changes in relative abundances of SCN5A, its variants, splicing factors RBM25 and LUC7A, and PERK, a major effector of the UPR, were analyzed by real time RT-PCR and the expression changes were confirmed by Western Blot. RESULTS: We found reduced full-length transcript, increased SCN5A truncation variants and activation of UPR in HCM when compared to control hearts. In these patients, real time RT-PCR revealed that HCM patients had decreased SCN5A mRNA to 27.8±4.07% of control (P<0.01) and an increased abundance of E28C and E28D (3.4±0.3 and 2.8±0.3-fold, respectively, P<0.05). PERK mRNA increased 8.2±3.1 fold (P<0.01) in HCM patients. Western blot confirmed a significant increase of PERK. CONCLUSIONS: These data suggested that the full-length SCN5A was reduced in patients with HCM. This reduction was accompanied by abnormal SCN5A pre-mRNA splicing and UPR activation. These changes may contribute to the arrhythmic risk in HCM.
BACKGROUND: Our previous studies showed that in ischemic and nonischemic heart failure (HF), the voltage-gated cardiac Na+ channel α subunit (SCN5A) mRNA is abnormally spliced to produce two truncated transcript variants (E28C and D) that activate the unfolded protein response (UPR). We tested whether SCN5A post-transcriptional regulation was abnormal in hypertrophic cardiomyopathy (HCM). MATERIAL AND METHODS:Human heart tissue was obtained from HCM patients. The changes in relative abundances of SCN5A, its variants, splicing factors RBM25 and LUC7A, and PERK, a major effector of the UPR, were analyzed by real time RT-PCR and the expression changes were confirmed by Western Blot. RESULTS: We found reduced full-length transcript, increased SCN5A truncation variants and activation of UPR in HCM when compared to control hearts. In these patients, real time RT-PCR revealed that HCM patients had decreased SCN5A mRNA to 27.8±4.07% of control (P<0.01) and an increased abundance of E28C and E28D (3.4±0.3 and 2.8±0.3-fold, respectively, P<0.05). PERK mRNA increased 8.2±3.1 fold (P<0.01) in HCM patients. Western blot confirmed a significant increase of PERK. CONCLUSIONS: These data suggested that the full-length SCN5A was reduced in patients with HCM. This reduction was accompanied by abnormal SCN5A pre-mRNA splicing and UPR activation. These changes may contribute to the arrhythmic risk in HCM.
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