Background:Heterozygous TTN truncating variants cause 10% to 20% of idiopathic dilated cardiomyopathy (DCM). Although variants which disrupt canonical splice signals (ie, invariant dinucleotide of the splice donor site, invariant dinucleotide of the splice acceptor site) at exon-intron junctions are readily recognized as TTN truncating variants, the effects of other nearby sequence variations on splicing and their contribution to disease is uncertain.Methods:Rare variants of unknown significance located in the splice regions of highly expressed TTN exons from 203 DCM cases, 3329 normal subjects, and clinical variant databases were identified. The effects of these variants on splicing were assessed using an in vitro splice assay.Results:Splice-altering variants of unknown significance were enriched in DCM cases over controls and present in 2% of DCM patients (P=0.002). Application of this method to clinical variant databases demonstrated 20% of similar variants of unknown significance in TTN splice regions affect splicing. Noncanonical splice-altering variants were most frequently located at position +5 of the donor site (P=4.4×10⁷) and position -3 of the acceptor site (P=0.002). SpliceAI, an emerging in silico prediction tool, had a high positive predictive value (86%–95%) but poor sensitivity (15%–50%) for the detection of splice-altering variants. Alternate exons spliced out of most TTN transcripts frequently lacked the consensus base at +5 donor and −3 acceptor positions.Conclusions:Noncanonical splice-altering variants in TTN explain 1-2% of DCM and offer a 10-20% increase in the diagnostic power of TTN sequencing in this disease. These data suggest rules that may improve efforts to detect splice-altering variants in other genes and may explain the low percent splicing observed for many alternate TTN exons.

中文翻译：

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非典型剪接变异对 TTN 截短变异心肌病的贡献

背景：杂合TTN截短变异导致 10% 至 20% 的特发性扩张型心肌病 (DCM)。尽管在外显子-内含子连接处破坏规范剪接信号的变体（即剪接供体位点的不变二核苷酸，剪接受体位点的不变二核苷酸）很容易被识别为TTN截断变体，但其他附近序列变异对剪接的影响及其对疾病的贡献是不确定的。方法：位于高表达TTN的剪接区域的未知意义的稀有变体鉴定了来自 203 个 DCM 病例、3329 名正常受试者和临床变异数据库的外显子。使用体外剪接试验评估了这些变体对剪接的影响。结果：与对照组相比，DCM 病例中富含未知意义的剪接改变变体，并且存在于 2% 的 DCM 患者中（P = 0.002）。将该方法应用于临床变异数据库表明，TTN剪接区域中 20% 的未知意义的相似变异会影响剪接。非常规剪接改变变体最常位于供体位点的 +5 位 ( P =4.4×10 ⁷ ) 和受体位点的 -3 位 ( P=0.002)。SpliceAI 是一种新兴的计算机预测工具，具有较高的阳性预测值 (86%–95%)，但对于检测剪接改变变体的敏感性较差 (15%–50%)。从大多数TTN转录本剪接的替代外显子经常在 +5 供体和 -3 受体位置缺乏共有碱基。结论： TTN中的非规范剪接改变变体解释了 1-2% 的 DCM，并提供了 10-20% 的诊断增加TTN测序在这种疾病中的作用。这些数据表明了可以改进检测其他基因中剪接改变变体的努力的规则，并可以解释在许多替代TTN外显子中观察到的低剪接百分比。