A mutation in the glutamate-rich region of RNA-binding motif protein 20 causes dilated cardiomyopathy through missplicing of titin and impaired Frank-Starling mechanism
A mutation in the glutamate-rich region of RNA-binding motif protein 20 causes dilated cardiomyopathy through missplicing of titin and impaired Frank-Starling mechanism
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DOI：
10.1093/cvr/cvw192
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发表时间：
2016-10-01
影响因子：
10.8
通讯作者：
Pinto, Yigal M.
中科院分区：
文献类型：
--
作者：
Beqqali, Abdelaziz;Bollen, Ilse A. E.;Pinto, Yigal M.
RNA结合基序蛋白20（RBM 20）RS结构域的突变最近被鉴定为与家族性扩张型心肌病（DCM）的侵袭性形式分离。大鼠RBM 20的缺失导致肌节基因肌联蛋白（TTN）的错误剪接。RBM 20突变热点以外的RBM 20突变的功能和生理后果迄今尚未探索。在本研究中，我们在人心肌细胞中发现了一个新的RBM 20突变引起的DCM的病理机制，我们鉴定了一个携带RBM 20（E913 K/+）突变的DCM家族，该突变位于RBM 20的谷氨酸丰富区。内源性RBM 20蛋白质的Western印迹分析显示RBM 20（E913 K/+）携带者的心脏中蛋白质水平强烈降低。RNA深度测序表明RBM 20（E913 K/+）载体中大量包含编码肌联蛋白弹簧区域的外显子。肌联蛋白同种型分析揭示了RBM 20（E913 K/+）心脏中从较低顺应性N2 B向高度顺应性N2 BA同种型的显著转变。此外，在单个心肌细胞中观察到肌节静止长度增加，等长收缩力测量显示Frank-Starling机制（FSM）减弱，蛋白激酶A治疗可挽救该机制; RBM 20突变热点外的突变导致RBM 20单倍不足。这会导致TTN的选择性剪接受到干扰，从而导致向高度顺应性肌联蛋白同种型的急剧转变和FSM受损。这些作用可能有助于由RBM 20突变引起的DCM的早期发作和恶性过程。总之，我们的研究结果表明，RBM 20的杂合丢失足以通过其TTN剪接的干扰而深刻地损害肌细胞生物力学。
Mutations in the RS-domain of RNA-binding motif protein 20 (RBM20) have recently been identified to segregate with aggressive forms of familial dilated cardiomyopathy (DCM). Loss of RBM20 in rats results in missplicing of the sarcomeric gene titin (TTN). The functional and physiological consequences of RBM20 mutations outside the mutational hotspot of RBM20 have not been explored to date. In this study, we investigated the pathomechanism of DCM caused by a novel RBM20 mutation in human cardiomyocytes.We identified a family with DCM carrying a mutation (RBM20(E913K/+)) in a glutamate-rich region of RBM20. Western blot analysis of endogenous RBM20 protein revealed strongly reduced protein levels in the heart of an RBM20(E913K/+) carrier. RNA deep-sequencing demonstrated massive inclusion of exons coding for the spring region of titin in the RBM20(E913K/+) carrier. Titin isoform analysis revealed a dramatic shift from the less compliant N2B towards the highly compliant N2BA isoforms in RBM20(E913K/+) heart. Moreover, an increased sarcomere resting-length was observed in single cardiomyocytes and isometric force measurements revealed an attenuated Frank-Starling mechanism (FSM), which was rescued by protein kinase A treatment.A mutation outside the mutational hotspot of RBM20 results in haploinsufficiency of RBM20. This leads to disturbed alternative splicing of TTN, resulting in a dramatic shift to highly compliant titin isoforms and an impaired FSM. These effects may contribute to the early onset, and malignant course of DCM caused by RBM20 mutations. Altogether, our results demonstrate that heterozygous loss of RBM20 suffices to profoundly impair myocyte biomechanics by its disturbance of TTN splicing.