Functional characterization of the novel DES mutation p.L136P associated with dilated cardiomyopathy reveals a dominant filament assembly defect
Functional characterization of the novel DES mutation p.L136P associated with dilated cardiomyopathy reveals a dominant filament assembly defect
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DOI：
10.1016/j.yjmcc.2015.12.015
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发表时间：
2016-02-01
影响因子：
5
通讯作者：
Milting, Hendrik
中科院分区：
文献类型：
--
作者：
Brodehl, Andreas;Dieding, Mareike;Milting, Hendrik
背景：扩张型心肌病(DCM)可能由40多个不同基因的突变引起。然而，在大多数情况下，特定突变的致病影响尚不清楚，这将使受影响家庭的遗传咨询复杂化。因此，功能研究有助于区分致病突变和良性变异。在这里，我们提出了一种新的杂合子DES错义变体(c.407C>；T；p.L136P)，通过下一代测序在DCM患者中鉴定。DES编码心脏中间丝蛋白结蛋白，在心肌细胞的机械稳定和细胞结构连接中具有重要作用。方法与结果：通过细胞转染实验和结合原子力显微镜的重组结蛋白组装实验，研究了这种新的DES变异体对微丝形成的影响。Desmin-p.L136P形成细胞质聚集体，表明该突变体存在严重的内在细丝组装缺陷。野生型和突变型结蛋白与不同荧光蛋白的共转染实验表明，突变型结蛋白对微丝组装具有显性影响。这些实验还得到了无孔扫描近场光学显微镜的补充。结论：体外分析表明，desmin-p.L136P不能形成规则的细丝，而是聚集在细胞质中。因此，我们将Des-p.L136P归类为可能的致病突变。综上所述，Des-p.L136P的功能特征可能与DES类似突变家系的遗传咨询有关，并有助于区分致病突变和良性罕见变异。(C)2016爱思唯尔有限公司。保留所有权利。
Background: Dilated cardiomyopathy (DCM) could be caused by mutations in more than 40 different genes. However, the pathogenic impact of specific mutations is in most cases unknown complicating the genetic counseling of affected families. Therefore, functional studies could contribute to distinguish pathogenic mutations and benign variants. Here, we present a novel heterozygous DES missense variant (c.407C > T; p.L136P) identified by next generation sequencing in a DCM patient. DES encodes the cardiac intermediate filament protein desmin, which has important functions in mechanical stabilization and linkage of the cell structures in cardiomyocytes.Methods and results: Cell transfection experiments and assembly assays of recombinant desmin in combination with atomic force microscopy were used to investigate the impact of this novel DES variant on filament formation. Desmin-p.L136P forms cytoplasmic aggregates indicating a severe intrinsic filament assembly defect of this mutant. Co-transfection experiments of wild-type and mutant desmin conjugated to different fluorescence proteins revealed a dominant affect of this mutant on filament assembly. These experiments were complemented by apertureless scanning near-field optical microscopy.Conclusion: In vitro analysis demonstrated that desmin-p.L136P is unable to form regular filaments and accumulate instead within the cytoplasm. Therefore, we classified DES-p.L136P as a likely pathogenic mutation. In conclusion, the functional characterization of DES-p.L136P might have relevance for the genetic counseling of affected families with similar DES mutations and could contribute to distinguish pathogenic mutations from benign rare variants. (C) 2016 Elsevier Ltd. All rights reserved.