DEFECTS IN MITOCHONDRIAL PROTEIN-SYNTHESIS AND RESPIRATORY-CHAIN ACTIVITY SEGREGATE WITH THE TRANSFER RNA(LEU)(UUR) MUTATION ASSOCIATED WITH MITOCHONDRIAL MYOPATHY, ENCEPHALOPATHY, LACTIC-ACIDOSIS, AND STROKE-LIKE EPISODES

Cytoplasts from two unrelated patients with MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes) harboring an A --> G transition at nucleotide position 3243 in the tRNA(Leu(UUR)) gene of the mitochondrial genome were fused with human cells lacking endogenous mitochondrial DNA (mtDNA) (rho(0) cells). Selected cybrid lines, containing < 15 or greater-than-or-equal-to 95% mutated genomes, were examined for differences in genetic, biochemical, and morphological characteristics. Cybrids containing greater-than-or-equal-to 95% mutant mtDNA, but not those containing normal mtDNA, exhibited decreases in the rats of synthesis and in the steady-state levels of the mitochondrial translation products. In addition, NADH dehydrogenase subunit 1 (ND 1) exhibited a slightly altered mobility on polyacrylamide gel electrophoresis. The mutation also correlated with a severe respiratory chain deficiency. A small but consistent increase in the steady-state levels of an RNA transcript corresponding to 16S rRNA + tRNA(Leu(UUR)) + ND 1 genes was detected. However, there was no evidence of major errors in processing of the heavy-strand-encoded transcripts or of altered steady-state levels or ratios of mitochondrial rRNAs or mRNAs. These results provide evidence for a direct relationship between the tRNA(Leu(UUR)) mutation and the pathogenesis of this mitochondrial disease.