Complete sequencing of 62 mitochondrial DNAs (mtDNAs) belonging (or very closely related) to haplogroup H revealed that this mtDNA haplogroup--by far the most common in Europe--is subdivided into numerous subhaplogroups, with at least 15 of them (H1-H15) identifiable by characteristic mutations. All the haplogroup H mtDNAs found in 5,743 subjects from 43 populations were then screened for diagnostic markers of subhaplogroups H1 and H3. This survey showed that both subhaplogroups display frequency peaks, centered in Iberia and surrounding areas, with distributions declining toward the northeast and southeast--a pattern extremely similar to that previously reported for mtDNA haplogroup V. Furthermore, the coalescence ages of H1 and H3 (~11,000 years) are close to that previously reported for V. These findings have major implications for the origin of Europeans, since they attest that the Franco-Cantabrian refuge area was indeed the source of late-glacial expansions of hunter-gatherers that repopulated much of Central and Northern Europe from ~15,000 years ago. This has also some implications for disease studies. For instance, the high occurrence of H1 and H3 in Iberia led us to re-evaluate the haplogroup distribution in 50 Spanish families affected by nonsyndromic sensorineural deafness due to the A1555G mutation. The survey revealed that the previously reported excess of H among these families is caused entirely by H3 and is due to a major, probably nonrecent, founder event.

Ethylmalonic encephalopathy (EE) is a devastating infantile metabolic disorder affecting the brain, gastrointestinal tract, and peripheral vessels. High levels of ethylmalonic acid are detected in the body fluids, and cytochrome c oxidase activity is decreased in skeletal muscle. By use of a combination of homozygosity mapping, integration of physical and functional genomic data sets, and mutational screening, we identified GenBank D83198 as the gene responsible for EE. We also demonstrated that the D83198 protein product is targeted to mitochondria and internalized into the matrix after energy-dependent cleavage of a short leader peptide. The gene had previously been known as "HSCO" (for hepatoma subtracted clone one). However, given its role in EE, the name of the gene has been changed to "ETHE1." The severe consequences of its malfunctioning indicate an important role of the ETHE1 gene product in mitochondrial homeostasis and energy metabolism.

We studied nine infant patients with a combination of progressive neurological and hepatic failure. Eight children, including two sibling pairs and four singletons, were affected by Alpers' hepatopathic poliodystrophy. A ninth baby patient suffered of a severe floppy infant syndrome associated with liver failure. Analysis of POLG1, the gene encoding the catalytic subunit of mitochondrial DNA polymerase, revealed that all the patients carried different allelic mutations in this gene. POLG1 is a major disease gene in mitochondrial disorders. Mutations in this gene can be associated with multiple deletions, depletion or point mutations of mitochondrial DNA (mtDNA). In turn, these different molecular phenotypes dictate an extremely heterogeneous spectrum of clinical outcomes, ranging from adult-onset progressive ophthalmoplegia to juvenile ataxic syndromes with epilepsy, to rapidly fatal hepatocerebral presentations, including Alpers' syndrome.

Isolated complex I deficiency, the most frequent OXPHOS disorder in infants and children, is genetically heterogeneous. Mutations have been found in seven mitochondrial DNA (mtDNA) and eight nuclear DNA encoded subunits, respectively, but in most of the cases the genetic basis of the biochemical defect is unknown. We analyzed the entire mtDNA and 11 nuclear encoded complex I subunits in 23 isolated complex I-deficient children, classified into five clinical groups: Leigh syndrome, progressive leukoencephalopathy, neonatal cardiomyopathy, severe infantile lactic acidosis, and a miscellaneous group of unspecified encephalomyopathies. A genetic definition was reached in eight patients (35%). Mutations in mtDNA were found in six out of eight children with Leigh syndrome, indicating a prevalent association between this phenotype and abnormalities in ND genes. In two patients with leukoencephalopathy, homozygous mutations were detected in two different nuclear-encoded complex I genes, including a novel transition in NDUFS1 subunit. In addition to these, a child affected by mitochondrial encephalomyopathy had heterozygous mutations in NDUFA8 and NDUFS2 genes, while another child with neonatal cardiomyopathy had a complex rearrangement in a single NDUFS7 allele. The latter cases suggest the possibility of unconventional patterns of inheritance in complex I defects.

Mutations in ETHE1, a gene located at chromosome 19q13, have recently been identified in patients affected by ethylmalonic encephalopathy (EE). EE is a devastating infantile metabolic disorder, characterised by widespread lesions in the brain, hyperlactic acidaemia, petechiae, orthostatic acrocyanosis, and high levels of ethylmalonic acid in body fluids. To investigate to what extent ETHE1 is responsible for EE, we analysed this gene in 29 patients with typical EE and in 11 patients presenting with early onset progressive encephalopathy with ethylmalonic aciduria (non-EE EMA). Frameshift, stop, splice site, and missense mutations of ETHE1 were detected in all the typical EE patients analysed. Western blot analysis of the ETHE1 protein indicated that some of the missense mutations are associated with the presence of the protein, suggesting that the corresponding wild type amino acid residues have a catalytic function. No ETHE1 mutations were identified in non-EE EMA patients. Experiments based on two dimensional blue native electrophoresis indicated that ETHE1 protein works as a supramolecular, presumably homodimeric, complex, and a three dimensional model of the protein suggests that it is likely to be a mitochondrial matrix thioesterase acting on a still unknown substrate. Finally, the 625GRA single nucleotide polymorphism in the gene encoding the short chain acyl-coenzyme A dehydrogenase (SCAD) was previously proposed as a co-factor in the aetiology of EE and other EMA syndromes. SNP analysis in our patients ruled out a pathogenic role of SCAD variants in EE, but did show a highly significant prevalence of the 625A alleles in non-EE EMA patients.