RhoA和血清反应因子（SRF）介导E1A激活基因阻遏子诱导人血管平滑肌细胞分化
The Cellular Repressor of E1A-stimulated Genes Promotes Human VSMCs Differentiation In vitro Mediated by RhoA and SRF
查看参考文献12篇
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文摘
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为探讨E1A激活基因阻遏子（CREG）对人血管平滑肌细胞（VSMCs）分化的调控机制，应用重组逆转录病毒表达载体pLNCX2（＋）／CREG及pLXSN（-）／CREG制备稳定感染HITASY细胞模型，观察CREG蛋白过表达及表达抑制对人VSMCs分化的影响并探讨其调控机制．结果显示：pLNCX2（＋）／CREG稳定感染细胞呈分化表型，细胞细长变成组织样聚集生长趋势，细胞中CREG蛋白和平滑肌分化标志蛋白平滑肌α-肌动蛋白（SM α-actin）表达显著增加，同时SM α-aetin相关调控因子——血清反应因子（SRF）入核转位，RhoA总蛋白表达上调，以Rho激酶特异性抑制剂Y-27632作用后，CREG诱导的SM α-actin表达下调的同时SRF出核转位；pLXSN（-）／CREG稳定感染的细胞体积变大，细胞极性消失，呈无序生长，细胞中CREG和SM α-actin蛋白表达显著降低，同时伴有SRF出核转位及RhoA总蛋白表达下调．免疫共沉淀分析发现，CREG蛋白能被分泌到VSMCs培养基中表达，并可与细胞膜受体6-磷酸甘露糖／胰岛素样生长因子Ⅱ型受体（M6P／IGF2R）发生直接相互作用．用蛋白磷酸酶PP2A特异性抑制剂okadaie acid减少M6P／IGF2R在细胞膜表面分布，可明显抑制CREG过表达引起的RhoA、SRF和SM α-aetin表达．上述结果提示，在体外培养的人VSMCs中，CREG可能作为一种分泌型蛋白质通过与细胞膜受体M6P／IGF2R相互作用，依次激活SM α-actin蛋白相关调控因子RhoA和SRF引起SM α-actin表达增加，促进VSMCs向分化表型转换． |
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其他语种文摘
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To investigate the mechanism of the cellular repressor of E1A-stimulated genes （CREG） on the differentiation of vascular smooth muscle cells （VSMCs）, the human internal thoracic artery cells （named HITASY cells） were stably infected with sense-CREG [pLNCX2 （＋）/CREG] and antisense-CREG [pLXSN （-）/CREG] retroviral vectors and positive clone was abtained by G418 selection. The cells infected with pLNCX2（＋）/CREG take on the differentiated phenotype and overexpression of CREG can enhance SM α-actin expression accompanied with increase of the total RhoA and nuclear serum response factor （SRF） expression detected by immunofluorescence and Western blot. And treated with a selective Rho kinase inhibitor Y-27632, the expression of SM α-actin and nuclear SRF protein induced by CREG was effectively reduced. Meanwhile, the opposite results were observed in the CREG-depression HITASY cells. And immunoprecipitation assays indicate that as a secreted protein, CREG can be detected in the media of pLNCX2（＋）/CREG cells and can interact with insulin-like growth factor Ⅱ/mannose 6-phosphate receptor （M6P/IGF2R）. Furthermore, that using okadaic acid （an inhibitor of the protein phosphatases 2A） to decrease the number of M6P/IGF2R at the cell surface can significantly inhibit the expression of total RhoA, nuclear SRF and SM α-actin induced by CREG-overexpression. Taken together, as a secreted protein, CREG can enhance the expression of SM α-actin by interaction with M6P/IGF2R and perform a pivotal role in the process of VSMCs differentiation and phenotype modulation. |
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来源
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生物化学与生物物理进展
,2006,33(5):438-445 【核心库】
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关键词
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阻遏蛋白
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E1A
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表型
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细胞
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血管平滑肌
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人
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地址
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1.
沈阳军区总医院全军心血管病研究所心内科, 沈阳, 110016
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大连市中心医院急诊科, 大连, 116033
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语种
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中文 |
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文献类型
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研究性论文 |
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ISSN
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1000-3282 |
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学科
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生物化学 |
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基金
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国家自然科学基金资助项目
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文献收藏号
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CSCD:2543256
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参考文献 共
12
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