Visualization of mitochondria with green fluorescent protein in cultured fibroblasts from patients with mitochondrial diseases
Visualization of mitochondria with green fluorescent protein in cultured fibroblasts from patients with mitochondrial diseases
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DOI：
10.1006/bbrc.1997.7448
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发表时间：
1997-10-20
影响因子：
3.1
通讯作者：
Niimi, H
中科院分区：
文献类型：
--
作者：
Kanazawa, M;Yano, M;Niimi, H
将绿色荧光蛋白（GFP）和含有人鸟氨酸转氨甲酰酶前序列的GFP融合蛋白（pOTC-GFP）的cDNA转染到培养的人成纤维细胞中。GFP cDNA在整个细胞质和细胞核中产生弥散荧光，而pOTC-GFP cDNA产生与细胞核相关的荧光。荧光线粒体结构可分为五种类型：线状线粒体、细线状线粒体、杆状线粒体、颗粒状线粒体和胞质荧光较弱的颗粒状线粒体。pOTC-GFP突变体导致线粒体荧光的丧失和整个细胞质中弱荧光的出现。将pOTC-GFP cDNA转染到来自患有各种线粒体疾病的患者的成纤维细胞中。在Reye综合征和Kearns-Sayre综合征患者中观察到颗粒状线粒体和细线状线粒体的成纤维细胞比例较高。在这些患者的成纤维细胞中有时观察到弱的胞质荧光。该方法将有助于分析线粒体结构改变和线粒体蛋白质输入障碍。(C)北京：科学出版社.
cDNAs for green fluorescent protein (GFP) and for a GFP fusion protein containing the presequence of human ornithine transcarbamylase (pOTC-GFP) were transfected into cultured human fibroblasts. GFP cDNA gave diffuse fluorescence throughout the cytoplasm and the nucleus, whereas pOTC-GFP cDNA gave mitochondria-associated fluorescence. Fluorescent mitochondrial structures could be classified into five patterns: thread-like mitochondria, fine thread-like ones, rod-like ones, granular ones, and granular ones with weak cytosolic fluorescence. pOTC-GFP mutants resulted in a loss of mitochondrial fluorescence and an appearance of weak fluorescence throughout the cytoplasm. pOTC-GFP cDNA was transfected into fibroblasts from patients with various mitochondrial diseases. Higher ratios of fibroblasts with granular mitochondria and those with fine thread-like ones were observed in a patient with Reye's syndrome and a patient with Kearns-Sayre syndrome. Weak cytosolic fluorescence was sometimes observed in fibroblasts from these patients. This method will be useful to analyze mitochondrial structural alterations and disorders of mitochondrial protein import. (C) 1997 Academic Press.