Cofilin-2: Molecular Function and it's Role in Myopathies

描述（由申请人提供）：本项目的长期目标是了解Cofilin-2 （CFL2）在骨骼肌中的作用及其在先天性肌病（CMs）中的突变。CMs的特点是相对非进行性骨骼肌无力，在生命早期出现张力低下。线状肌病（NM）是最常见的类型，是一种遗传疾病，肌肉中存在线状体，并且在构成细纤维的五种不同基因中发现突变。在最近的一项突破中，我发现了第六个基因，CFL2，在NM中发生了突变。两个兄弟姐妹的临床诊断为典型的NM和非特异性先天性肌病，由近亲父母出生，携带纯合错义突变c.103G> a (A35T)，并在他们的肌肉中表现出显著降低的cofilin-2蛋白水平。与正常对照相比，CFL2 mRNA的数量增加了约8倍。Cofilin-2属于AC蛋白，包括cofilin-1、Cofilin-2和肌动蛋白解聚因子。这些蛋白通过引起肌动蛋白分离和解聚而起作用，在包括运动、细胞分裂、内吞作用和肌肉发育在内的广泛细胞功能中都是必不可少的。这是首次发现人类AC组基因突变的病例。由这种突变引起的NM/先天性肌病的分子机制需要了解。此外，cofilin-2在骨骼肌中的功能也有待阐明。在目标1中，我们将筛选所有NM/其他CM病例的CFL2突变，并描述与这些突变相关的转录谱

DESCRIPTION (provided by applicant): The long-term goals of this project are to understand the role of Cofilin-2 (CFL2) in skeletal muscle and its mutation in congenital myopathies (CMs). CMs are characterized by relatively non-progressive skeletal muscle weakness, hypotonia presenting in early life. Nemaline Myopathy (NM), the most common type, is a genetic disorder with nemaline bodies present in the muscle and mutations identified in five different genes that constitute the thin filament. In a recent breakthrough, I have identified a sixth gene, CFL2, mutated in NM. Two siblings with clinical diagnosis of typical NM and non-specific congenital myopathy born to consanguineous parents carry a homozygous missense mutation c.103G>A (A35T), and exhibit drastically reduced levels of cofilin-2 protein in their muscles. In contrast, the amount of CFL2 mRNA is increased about 8 fold compared to normal controls. Cofilin-2 belongs to AC group of proteins that include cofilin-1, cofilin-2 and actin depolymerization factor. These proteins function by causing actin severance and depolymerization, and are essential in a wide range of cellular functions including locomotion, cytokinesis, endocytosis, and muscle development. This is the first identified case of a human mutation in the AC group of genes. The molecular mechanism of NM/congenital myopathy caused by this mutation needs to be understood. In addition, cofilin-2 function in skeletal muscle needs to be elucidated. In Aim 1, we will screen all NM/other CM cases for CFL2 mutations and characterize the transcriptional profile associated with these mutations to understand downstream effects. In Aim 2, we will evaluate the biochemical and functional consequences of the identified mutation. In Aim 3, CFL2 expression in C2C12 cells will be knocked down using shRNA techniques and functional consequences will be evaluated. In Aim 4, conventional Cfl2 knockout and c.103G>A mutation knockin mouse models will be generated. This project will introduce me to a variety of techniques including microarray analysis, gene-silencing and creating knockout/knockin mouse models. It willl give me the opportunity to work in an intellectually stimulating environment, where I will approach scientific questions in novel ways fostering my transition to independence. My sponsor Dr Beggs, collaborators/advisors Dr Darras, Dr Dormitzer, Dr Kohane, Dr Kunkel and Dr Maciver, and my Neonatology Division Chief Dr Kourembanas will help me in this process.