Smooth muscle calponin: an unconventional CArG-dependent gene that antagonizes neointimal formation.
Smooth muscle calponin: an unconventional CArG-dependent gene that antagonizes neointimal formation.
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DOI：
10.1161/atvbaha.111.232785
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发表时间：
2011-10
期刊：
影响因子：
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通讯作者：
Miano JM
中科院分区：
文献类型：
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作者：
Long X;Slivano OJ;Cowan SL;Georger MA;Lee TH;Miano JM
平滑肌钙调蛋白（CNN 1）含有多个保守的内含子CArG元件，其结合血清反应因子（SRF）并在体外显示增强子活性。本文的目的是评估这些CArG元件在转基因小鼠中的活性，并确定人CNN 1对损伤诱导的血管重塑的影响。在人CNN 1基因内含子CArG元件控制下携带lacZ报告基因的小鼠未能显示出平滑肌细胞（SMC）限制性活性。然而，在小鼠中的orthopathic序列的删除废除内源性Cnn 1启动子的活性，表明它们的必要性在体内Cnn 1表达。通过免疫组织化学和蛋白质印迹法测量，携带含有人CNN 1基因的38-腺苷酸酶细菌人工染色体（BAC）的小鼠显示相应CNN 1蛋白的SMC限制性表达。广泛的BAC重组工程研究揭示了一个单一的内含子CArG元件的绝对必要性，正确的SMC限制性表达的人CNN 1。过表达人CNN 1抑制动脉损伤后新生内膜形成。具有相同BAC的小鼠在CArG元件中携带抑制人CNN 1表达的突变，表现出向外重塑和新生内膜形成。一个单一的内含子CArG元件是必要的，但不足以适当的CNN 1在体内表达。CNN 1过表达拮抗动脉损伤诱导的新生内膜形成
Smooth muscle calponin (CNN1) contains multiple, conserved intronic CArG elements that bind serum response factor (SRF) and display enhancer activity in vitro. The objectives here were to evaluate these CArG elements for activity in transgenic mice and determine the effect of human CNN1 on injury-induced vascular remodeling. Mice carrying a lacZ reporter under control of intronic CArG elements in the human CNN1 gene failed to show smooth muscle cell (SMC)-restricted activity. However, deletion of the orthologous sequences in mice abolished endogenous Cnn1 promoter activity suggesting their necessity for in vivo Cnn1 expression. Mice carrying a 38-kilobase bacterial artificial chromosome (BAC) harboring the human CNN1 gene displayed SMC- restricted expression of the corresponding CNN1 protein as measured by immunohistochemistry and Western blotting. Extensive BAC recombineering studies revealed the absolute necessity of a single intronic CArG element for correct SMC-restricted expression of human CNN1. Over-expressing human CNN1 suppressed neointimal formation following arterial injury. Mice with an identical BAC carrying mutations in CArG elements that inhibit human CNN1 expression, showed outward remodeling and neointimal formation. A single intronic CArG element is necessary but insufficient for proper CNN1 expression in vivo. CNN1 over-expression antagonizes arterial injury-induced neointimal formation.