We purified CALLA from human kidney and isolated a cDNA clone reactive with two oligonucleotide probes corresponding to two distinct peptides. The amino acid sequence translated from the CALLA cDNA revealed 100% identity with that of human neutral endopeptidase (NEP, enkephalinase). The distribution of CALLA antigen and NEP in normal tissues are similar.

Mice homozygous for a defect in the PTCD2 (pentatricopeptide repeat domain protein 2) gene were generated in order to study the role of this protein in mitochondrial RNA metabolism. These mice displayed specific but variable reduction of ubiquinol-cytochrome c reductase complex activity in mitochondria of heart, liver and skeletal muscle due to a decrease in the expression of mitochondrial DNA-encoded cytochrome b, the catalytic core of the complex. This reduction in mitochondrial function has a profound effect on the myocardium, with replacement of ventricular cardiomyocytes by fibro-fatty tissue. Northern blotting showed a reduction in the mRNA for the mitochondrial DNA encoded proteins cytochrome b (cytb) and ND5 (NADH dehydrogenase subunit 5) and an elevation in a combined pre-processed ND5-CYTB transcript. This suggests that the PTCD2 protein is involved in processing RNA transcripts involving cytochrome b derived from mitochondrial DNA. This defines the site for PTCD2 action in mammalian mitochondria and suggests a possible role for dysfunction of this protein in the aetiology of heart failure.

A T-to-C missense mutation at nucleotide position 9,185 in the protein-coding ATP6 gene of the mitochondrial genome was present at high heteroplasmy in members of a Canadian family with Leigh syndrome with predominant ataxia and peripheral neuropathy. This mutation results in the substitution of a proline residue for an evolutionary-conserved leucine at position of amino acid 220 near the carboxyl terminus of the mitochondrial protein. The index patient and brother, who had an identical clinical presentation, had >90% mutant mtDNA in cultured skin fibroblasts, lymphocytes, and whole blood. Their mother and a maternal uncle, symptomatic with a peripheral neuropathy alone, had 86% and 85% heteroplasmy, respectively. Symptomatic maternal cousins with early onset revealed 90% and 91% mutant mtDNA in all tissues analyzed. Studies of lymphoblasts from the asymptomatic maternal grandmother and eldest brother of the proband were heteroplasmic for mutant mtDNA with 56% and 17%, respectively. Biochemical analysis demonstrated normal respiratory chain enzyme activity in muscle and fibroblasts, normal ATP synthesis, but reduced oligomycin-sensitive H(+)ATPase in cultured lymphoblast mitochondria. We propose that the 9,185T > C mtDNA mutation is pathogenic even though the initial phenotype is mild and the biochemical phenotype not easily detectable.