VISUALIZATION OF MITOCHONDRIA WITH GREEN FLUORESCENT PROTEIN IN CULTURED FIBROBLASTS FROM PATIENTS WITH MITOCHONDRIAL DISEASES

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 d iffuse fluorescence throughout the cytoplasm and the nucleus, whereas pOTC-GFP cDNA gave mitochondria-associated fluorescence. Fluorescent m itochondrial structures could be classified into five patterns: thread -like mitochondria, fine thread-like ones, rod-like ones, granular one s, and granular ones with weak cytosolic fluorescence. pOTC-GFP mutant s resulted in a loss of mitochondrial fluorescence and an appearance o f weak fluorescence throughout the cytoplasm. pOTC-GFP cDNA was transf ected into fibroblasts from patients with various mitochondrial diseas es. Higher ratios of fibroblasts with granular mitochondria and those with fine thread-like ones were observed in a patient with Reye's synd rome and a patient with Kearns-Sayre syndrome. Weak cytosolic fluoresc ence was sometimes observed in fibroblasts from these patients. This m ethod will be useful to analyze mitochondrial structural alterations a nd disorders of mitochondrial protein import. (C) 1997 Academic Press.