What is the evidence of a role for host genetics in susceptibility to influenza A/H5N1?

The apparent family clustering of avian influenza A/H5N1 has led several groups to postulate the existence of a host genetic influence on susceptibility to A/H5N1, yet the role of host factors on the risk of A/H5N1 disease has received remarkably little attention compared to the efforts focused on viral factors. We examined the epidemiological patterns of human A/H5N1 cases, their possible explanations, and the plausibility of a host genetic effect on susceptibility to A/H5N1 infection. The preponderance of familial clustering of cases and the relative lack of non-familial clusters, the occurrence of related cases separated by time and place, and the paucity of cases in some highly exposed groups such as poultry cullers, are consistent with a host genetic effect. Animal models support the biological plausibility of genetic susceptibility to A/H5N1. Although the evidence is circumstantial, host genetic factors are a parsimonious explanation for the unusual epidemiology of human A/H5N1 cases and warrant further investigation.

The evolutionary origins of Southeast Asian Ovalocytosis.

Southeast Asian Ovalocytosis (SAO) is a common red blood cell disorder that is maintained as a balanced polymorphism in human populations. In individuals heterozygous for the SAO-causing mutation there are minimal detrimental effects and well-documented protection from severe malaria caused by Plasmodium vivax and Plasmodium falciparum; however, the SAO-causing mutation is fully lethal in utero when homozygous. The present-day high frequency of SAO in Island Southeast Asia indicates the trait is maintained by strong heterozygote advantage. Our study elucidates the evolutionary origin of SAO by characterizing DNA sequence variation in a 9.5 kilobase region surrounding the causal mutation in the SLC4A1 gene. We find substantial haplotype diversity among SAO chromosomes and estimate the age of the trait to be approximately 10,005 years (95% CI: 4930-23,200 years). This date is far older than any other human malaria-resistance trait examined previously in Southeast Asia, and considerably pre-dates the widespread adoption of agriculture associated with the spread of speakers of Austronesian languages some 4000 years ago. Using a genealogy-based method we find no evidence of historical positive selection acting on SAO (s=0.0, 95% CI: 0.0-0.03), in sharp contrast to the strong present-day selection coefficient (e.g., 0.09) estimated from the frequency of this recessively lethal trait. This discrepancy may be due to a recent increase in malaria-driven selection pressure following the spread of agriculture, with SAO targeted as a standing variant by positive selection in malarial populations.

Antimitochondrial autoantibodies of primary biliary cirrhosis as a novel probe in the study of 2-oxo acid dehydrogenases in patients with mitochondrial myopathies.

Autoantibodies present in the autoimmune disease primary biliary cirrhosis react by immunoblotting with four human skeletal muscle mitochondrial antigens of 70 kDa, 52 kDa, 50 kDa and 45 kDa, identified as the lipoate acetyl transferases (E2) of the pyruvate dehydrogenase, component X of E2 pyruvate dehydrogenase, E2 of 2-oxo glutarate dehydrogenase and E2 of branched-chain 2-oxo acid dehydrogenase complexes respectively. These autoantibodies have been employed as a novel probe to study whether there is a defect in the synthesis of the 2-oxo acid dehydrogenase complexes in patients with mitochondrial respiratory chain disorders. The reactive antigens are present normally in four patients with oculomyopathy in whom partial deletions of the mtDNA have been detected, and in two patients with MERRF and MELAS encephalomyopathy. Thus, unlike in the yeast Saccharomyces cerevisiae, there appear to be no regulatory interactions which coordinate the assembly of the mitochondrial respiratory chain with the development of the pyruvate dehydrogenase complex, which plays an important role in regulating the flow of metabolic intermediates to oxidative energy metabolism.

Phenotypic expression of mtDNA heteroplasmy in the skeletal muscle of patients with oculomyopathy: defect in mitochondrial protein synthesis.

The biochemical consequences of mtDNA heteroplasmy, observed in patients with a range of diseases associated with the mitochondrial respiratory enzymes deficiency is of particular interest, as they might provide information with regard to the regulatory interactions which govern the expression of the human mitochondrial genome. Three patients with chronic progressive external ophthalmoplegia (CPEO) were investigated to study the consequences of mtDNA heteroplasmy on mitochondrial protein synthesis. All 3 patients exhibited partially deleted mtDNA species (varying in size from 10.5 to 14 kb) in their skeletal muscle, which co-existed with the normal 16.5 kb mtDNA. The examination of mitochondrial translation products following the incorporation of [35S]methionine by isolated mitochondria, showed grossly abnormal patterns of mitochondrial translation products, suggesting a major disturbance in the regulation of mitochondrial protein synthesis.

Molecular genetics of Leber's hereditary optic neuropathy: study of a six-generation family from Western Australia.

Molecular genetic studies were carried out on a 6-generation family from Western Australia with Leber's hereditary optic neuropathy. Pedigree analysis confirms the maternal inheritance of the genetic lesion underlying the disorder in this family. The presence of a recently reported disease-associated mutation at nucleotide 11778 of the mtDNA was established in one clinically affected family member by the sequencing of an appropriate 1.6 kb PCR-amplified fragment of the mtDNA; this mutation leads to an Arg340----His amino acid replacement in the ND4 subunit of respiratory complex I. The 11778 G to A base substitution is associated with the loss of an SfaNI restriction site. Examination of the representative members for this site revealed that while only mtDNA carrying this substitution could be detected in the leukocytes of 4 family members of the sixth generation, the mutated mtDNA was found to co-exist with the normal mtDNA population (heteroplasmy) in a clinically unaffected member from the fifth generation. This observation suggests that the nt 11778 mutation observed in this LHON family is relatively new; the observation of both heteroplasmy and apparent homoplasmy of the mtDNA in different family members might reflect the normal progression in the establishment of a mitochondrially inherited mutation.

Functional respiratory chain studies in subjects with chronic progressive external ophthalmoplegia and large heteroplasmic mitochondrial DNA deletions.

The functional consequences of large heteroplasmic mtDNA deletions were investigated in a group of 6 patients with chronic progressive external ophthalmoplegia (CPEO) syndromes. State III respiration rates corrected for age were low with site I and II substrates in all cases and cytochrome oxidase activity was depressed. The severity of impairment varied and is consistent with inclusion of a variable percentage of non-functioning mitochondria (with deleted mtDNA) in the pellet. Western blot studies with a holocomplex antibody battery revealed no abnormalities in subunit content of complexes III and IV. A deficiency of several complex I subunits in 3 cases suggests that abnormal nuclear-mitochondrial regulation of complex I assembly may follow large mtDNA deletions.

Developmental genetics of deleted mtDNA in mitochondrial oculomyopathy.

Heteroplasmic populations of mtDNA, consisting of normal mtDNA and mtDNA with large deletions, are found in the skeletal muscle and other tissues of certain patients with mitochondrial respiratory chain deficiencies, particularly in those with the CPEO (chronic progressive external ophthalmoplegia) phenotype. To study the developmental genetics of this mitochondrial disorder, the distribution of the deleted mtDNA in a wide range of tissues of different embryonic origins (total 34 samples from 27 tissues obtained at autopsy) was investigated in a patient with the CPEO syndrome. Three species of partially deleted mtDNA were observed, with deletions of 2.3 kb, 5.0 kb and 6.4 kb. Their tissue distribution suggests that the mtDNA deletions have occurred very early during embryonic development, prior to the differentiation events that lead to the formation of the three primary embryonic germ layers, and that the partially deleted mtDNA species were segregated during development mainly to the skeletal muscle and to tissues of the central nervous system.

Anti-mitochondrial autoantibodies of primary biliary cirrhosis as a novel probe in the study of the biosynthetic regulation of the yeast 2-oxo acid dehydrogenase complexes.

Autoantibodies present in the disease primary biliary cirrhosis react by immunoblotting with four major yeast mitochondrial antigens of 58 kDa, 55 kDa, 52 kDa and 45 kDa, tentatively identified as the lipoate acetyl transferases (E2) of the pyruvate dehydrogenase, component X of E2 pyruvate dehydrogenase, E2 of 2-oxo glutarate dehydrogenase and E2 of branched-chain 2-oxo acid dehydrogenase complexes respectively. The synthesis of these antigens is sensitive to catabolite repression. The reactive antigens are present in mit- mutants of yeast which have specific defects in the mitochondrial apocytochrome b, cytochrome oxidase subunit II and H+ -ATPase subunits 8 and 9, and in mtDNA-less rho O petite mutants, but a significant increase in the sensitivity to catabolite repression was observed in these mutants in particular in the mtDNA-less strains.

Microphotometric analysis of NADH-tetrazolium reductase deficiency in fibroblasts of patients with Leber hereditary optic neuropathy.

We employed a microphotometric approach to examine whether a defect in the mitochondrial respiratory complex I expected in Leber hereditary optic neuropathy (LHON) as the consequence of a mtDNA (11778G>A) mutation in the ND4 gene coding for a subunit of the respiratory complex I can be detected at the single-cell level. Genetically stable fibroblast cell lines were established from skin biopsies of two members of a Chinese Indonesian family with LHON. The fibroblasts were homoplasmic for the 11778G>A mutation. The activity of the respiratory complex I was examined histochemically by staining for NADH-tetrazolium reductase. The histochemical staining showed a typical pattern with an apparent concentration of the activity around the nucleus, suggested as the reflection of the gradient in the thickness of the unsectioned fibroblast cells. Microphotometric quantification of the staining intensity showed that the activity is linear for at least 60 min. The activity shows a discontinuity in its Arrhenius kinetics with a break point at 13.0-13.5 degrees C (activation energy at 50-58 J/mol and 209-238 J/mol above and below the break temperature, respectively), indicating the membrane association of the NADH-tetrazolium reductase activity. Both patients showed lower fibroblast NADH-tetrazolium reductase activity, with a reduction of degrees 30%. Our results demonstrate the utility of microphotometric analysis in the study of biochemical defects associated with mutations in the mtDNA.

The association of the autoantigens of primary biliary cirrhosis with the mitochondrial H+-ATPase--a reassessment.

The claimed association between the M2 autoantigens of primary biliary cirrhosis (PBC) and the mitochondrial H+-ATPase has been re-examined in view of the recent reports that PBC autoantibodies react specifically with the lipoate acetyl transferases of 2-oxo acid dehydrogenases. Study of F0F1-ATPase purified from human and yeast mitochondria, and the comparison between immunoprecipitates obtained with antibodies against the H+-ATPase beta subunit and anti-M2 antibodies of PBC, established that the M2 antigens are not associated with the H+-ATPase complex. The M2 antigens did copurify with a crude bovine heart F1-ATPase preparation, but not with F1-ATPase from yeast, human heart or human liver.

Meiotic breakpoint mapping of a proposed X linked visual loss susceptibility locus in Leber's hereditary optic neuropathy.

Leber's hereditary optic neuropathy (LHON) is a maternally inherited degenerative disorder characterised by an acute or subacute optic nerve degeneration resulting in visual failure. Mitochondrial DNA mutations have been reported and a nuclear modifier gene(s) on the X chromosome is thought to play an important role in the onset of this disorder. We analysed a LHON family with a novel and more accurate approach using 27 X chromosomal microsatellite markers. Meiotic breakpoint mapping and two point lod score did not point to any particular area on the X chromosome which might contain the X susceptibility locus.

A tRNA(Lys) mutation in the mtDNA is the causal genetic lesion underlying myoclonic epilepsy and ragged-red fiber (MERRF) syndrome.

Skeletal muscle mtDNA of three patients with mitochondrial encephalomyopathy, characterized clinically by myoclonic epilepsy and ragged-red fiber (MERRF) syndrome, has been sequenced to determine the underlying molecular defect(s). An A-to-G substitution of nt 8344 in the tRNA(Lys) gene, a substitution suggested to be associated with MERRF encephalomyopathy, was detected in these patients. Abnormal patterns of mitochondrial translation products were observed in the skeletal muscle of patients, consistent with the expected consequential defect in protein synthesis. The genealogical studies of the three patients, as well as mtDNA from one published MERRF patient and from nine other normal and disease controls, revealed that the tRNA(Lys) mutations in the MERRF patients have arisen independently. These observations provided evidence that the base substitution is a causal mutation for MERRF.