Morphological analysis of 13 LMNA variants identified in a cohort of 324 unrelated patients with idiopathic or familial dilated cardiomyopathy.
Morphological analysis of 13 LMNA variants identified in a cohort of 324 unrelated patients with idiopathic or familial dilated cardiomyopathy.
复制标题
DOI：
10.1161/circgenetics.109.905422
复制
发表时间：
2010-02
期刊：
影响因子：
--
通讯作者：
Hershberger RE
中科院分区：
文献类型：
--
作者：
Cowan J;Li D;Gonzalez-Quintana J;Morales A;Hershberger RE
编码核纤层蛋白A/C的LMNA基因突变是扩张型心肌病（DCM）的重要原因。我们最近在324名无关的DCM患者队列中发现了18种改变蛋白质的LMNA变体。然而，在6个家系中，每个家系至少有一个DCM家族成员缺乏LMNA突变（非分离），而另外5个家族的小规模排除了分离的明确确定，提出了关于这些变异对疾病的贡献的问题。因此，我们在COS 7细胞中表达了GFP-前层蛋白A（GFPLaA）融合构建体，该融合构建体在非分离的家系中并入了六种变体。（R101 P，A318 T，R388 H，R399 C，S437 Hfsx 1，and R654 X），家系中的4个变异体，分离未知[R89 L，R166 P（在2个家族中），I210 S，R471 H]和三个与疾病分离的额外错义变体（R190 Q，E203 K，L215 P）。共聚焦免疫荧光显微镜用于表征GFP-核纤层蛋白A的定位和核形态。在10/13（77%）个变异体（R89 L、R101 P、R166 P、R190 Q、E203 K、I210 S、L215 P、R388 H、S437 Hfsx 1、R654 X）中观察到异常表型，包括4/6个证明未分离和3/4个分离不确定。所有七个影响线圈1B的变异，和核纤层蛋白A-唯一的突变，R654 X，表现出膜结合的GFP-核纤层蛋白A聚集体和核形状异常。出乎意料的是，R388 H在很大程度上将GFP-核纤层蛋白A限制在细胞质中。同样出乎意料的是具有S437 Hfsx 1的独特条纹聚集体，以及具有S437 Hfsx 1和R654 X的巨大聚集体。这项工作扩大了核纤层蛋白A定位异常的DCM的公认光谱。它还提供了证据，支持13个测试的LMNA变体中的10个的致病性，包括一些不确定或非分离的变体。
Mutations in the LMNA gene, encoding lamins A/C, represent a significant cause of dilated cardiomyopathy (DCM). We recently identified 18 protein-altering LMNA variants in a cohort of 324 unrelated patients with DCM. However, at least one family member with DCM in each of six pedigrees lacked the LMNA mutation (nonsegregation), while small sizes of five additional families precluded definitive determinations of segregation, raising questions regarding contributions by those variants to disease. We have, consequently, expressed, in COS7 cells, GFP-prelamin A (GFPLaA) fusion constructs incorporating the six variants in pedigrees with nonsegregation (R101P, A318T, R388H, R399C, S437Hfsx1, and R654X), the four variants in pedigrees with unknown segregation [R89L, R166P (in 2 families), I210S, R471H], and three additional missense variants (R190Q, E203K, L215P) that segregated with disease. Confocal immunofluorescence microscopy was used to characterize GFP-lamin A localization and nuclear morphology. Abnormal phenotypes were observed for 10/13 (77%) variants (R89L, R101P, R166P, R190Q, E203K, I210S, L215P, R388H, S437Hfsx1, R654X), including 4/6 demonstrating nonsegregation and 3/4 with uncertain segregation. All seven variants affecting coil 1B, and the lamin A-only mutation, R654X, exhibited membrane-bound GFP-lamin A aggregates and nuclear shape abnormalities. Unexpectedly, R388H largely restricted GFP-lamin A to the cytoplasm. Equally unexpected were unique streaked aggregates with S437Hfsx1, and giant aggregates with both S437Hfsx1 and R654X. This work expands the recognized spectrum of lamin A localization abnormalities in DCM. It also provides evidence supporting pathogenicity of 10 of 13 tested LMNA variants, including some with uncertain or nonsegregation.