Background:Abnormalities in Ca²⁺ homeostasis are associated with cardiac arrhythmias and heart failure. Triadin plays an important role in Ca²⁺ homeostasis in cardiomyocytes. Alternative splicing of a single triadin gene produces multiple triadin isoforms. The cardiac-predominant isoform, mouse MT-1 or human Trisk32, is encoded by triadin exons 1 to 8. In humans, mutations in the triadin gene that lead to a reduction in Trisk32 levels in the heart can cause cardiac dysfunction and arrhythmias. Decreased levels of Trisk32 in the heart are also common in patients with heart failure. However, mechanisms that maintain triadin isoform composition in the heart remain elusive.Methods:We analyzed triadin expression in heart explants from patients with heart failure and cardiac arrhythmias and in hearts from mice carrying a knockout allele for Trdn-as, a cardiomyocyte-specific long noncoding RNA encoded by the antisense strand of the triadin gene, between exons 9 and 11. Catecholamine challenge with isoproterenol was performed on Trdn-as knockout mice to assess the role of Trdn-as in cardiac arrhythmogenesis, as assessed by ECG. Ca²⁺ transients in adult mouse cardiomyocytes were measured with the IonOptix platform or the GCaMP system. Biochemistry assays, single-molecule fluorescence in situ hybridization, subcellular localization imaging, RNA sequencing, and molecular rescue assays were used to investigate the mechanisms by which Trdn-as regulates cardiac function and triadin levels in the heart.Results:We report that Trdn-as maintains cardiac function, at least in part, by regulating alternative splicing of the triadin gene. Knockout of Trdn-as in mice downregulates cardiac triadin, impairs Ca²⁺ handling, and causes premature death. Trdn-as knockout mice are susceptible to cardiac arrhythmias in response to catecholamine challenge. Normalization of cardiac triadin levels in Trdn-as knockout cardiomyocytes is sufficient to restore Ca²⁺ handling. Last, Trdn-as colocalizes and interacts with serine/arginine splicing factors in cardiomyocyte nuclei and is essential for efficient recruitment of splicing factors to triadin precursor mRNA.Conclusions:These findings reveal regulation of alternative splicing as a novel mechanism by which a long noncoding RNA controls cardiac function. This study indicates potential therapeutics for heart disease by targeting the long noncoding RNA or pathways regulating alternative splicing.

中文翻译：

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心肌细胞特异性长非编码 RNA 调节心脏中 Triadin 基因的选择性剪接

背景：Ca ²⁺稳态异常与心律失常和心力衰竭有关。^{Triadin 在心肌细胞 Ca 2+}稳态中发挥重要作用。单个三联蛋白基因的选择性剪接产生多种三联蛋白亚型。心脏主要同种型小鼠 MT-1 或人类 Trisk32，由三联蛋白外显子 1 至 8 编码。在人类中，三联蛋白基因突变导致心脏中 Trisk32 水平降低，可导致心脏功能障碍和心律失常。心脏中 Trisk32 水平降低在心力衰竭患者中也很常见。然而，维持心脏中三联蛋白异构体组成的机制仍然难以捉摸。方法：我们分析了心力衰竭和心律失常患者的心脏外植体以及携带Trdn-as（一种心肌细胞特异性长链）敲除等位基因的小鼠心脏中三联蛋白的表达。由triadin基因反义链编码的非编码 RNA ，位于外显子 9 和 11 之间。对Trdn-as敲除小鼠进行儿茶酚胺攻击，以评估Trdn-as在心律失常发生中的作用（通过心电图评估）。使用 IonOptix 平台或 GCaMP 系统测量成年小鼠心肌细胞中的Ca ^2+瞬变。采用生物化学测定、单分子荧光原位杂交、亚细胞定位成像、RNA测序和分子救援测定来研究Trdn-as调节心脏功能和心脏三联蛋白水平的机制。结果：我们报告Trdn - as as至少部分通过调节三联蛋白基因的选择性剪接来维持心脏功能。敲除小鼠中的Trdn-as会下调心脏三联蛋白，损害 Ca ²⁺处理，并导致过早死亡。Trdn-as基因敲除小鼠在儿茶酚胺激发后容易出现心律失常。Trdn-as敲除心肌细胞中心脏三联蛋白水平的正常化足以恢复 Ca ²⁺处理。最后，Trdn-as与心肌细胞核中的丝氨酸/精氨酸剪接因子共定位并相互作用，对于有效地将剪接因子募集到三联蛋白前体 mRNA 中至关重要。结论：这些发现揭示了选择性剪接的调节是长非编码 RNA 的一种新机制。控制心脏功能。这项研究表明，通过靶向长非编码 RNA 或调节选择性剪接的途径，可以治疗心脏病。