Bodnar A G, Cooper J M, Holt I J, Leonard J V, Schapira A H
Department of Neurological Science, Royal Free Hospital School of Medicine, London, England.
Am J Hum Genet. 1993 Sep;53(3):663-9.
We have studied cultured skin fibroblasts from a patient with a fatal mitochondrial disease manifesting soon after birth. These fibroblasts were found to grow only in the presence of pyruvate and uridine, a characteristic of cells lacking mtDNA (rho0 cells). Southern blot and PCR analyses confirmed that the patient's fibroblasts contained less than 2% of control levels of mtDNA. Biochemical analyses indicated that the activities of all the respiratory-chain enzymes were severely decreased in mitochondria isolated from these fibroblasts. In order to elucidate the underlying molecular defect, cell fusions were performed between enucleated fibroblasts from this patient and a human-derived rho0 cell line (rho0 A549.B2). The resulting cybrids were plated in medium lacking pyruvate and uridine, to select for the restoration of respiratory-chain function. Complementation was observed between the nuclear genome of the rho0 A549.B2 cells and the mtDNA of the patient's cells, restoring mtDNA levels and respiratory-chain function in the cybrid cells. These results indicate that mtDNA depletion in our patient is under the control of the nuclear genome.
我们研究了一名患有致命线粒体疾病的患者的培养皮肤成纤维细胞，该疾病在出生后不久就会显现。这些成纤维细胞被发现仅在丙酮酸和尿苷存在的情况下才能生长，这是缺乏线粒体DNA的细胞（ρ0细胞）的特征。Southern印迹和PCR分析证实，患者的成纤维细胞所含线粒体DNA水平不到对照水平的2%。生化分析表明，从这些成纤维细胞分离出的线粒体中，所有呼吸链酶的活性均严重降低。为了阐明潜在的分子缺陷，我们将该患者去核的成纤维细胞与一种人源ρ0细胞系（ρ0 A549.B2）进行了细胞融合。将所得的胞质杂种接种在缺乏丙酮酸和尿苷的培养基中，以选择恢复呼吸链功能的细胞。在ρ0 A549.B2细胞的核基因组与患者细胞的线粒体DNA之间观察到了互补作用，恢复了胞质杂种细胞中的线粒体DNA水平和呼吸链功能。这些结果表明，我们患者中线粒体DNA的耗竭受核基因组的控制。
