Clinical,genetic, and biochemical characterization of a Leber hereditary optic neuropathy family containing both the 11778 and 14484 primary mutations

Four mitochondrial DNA (mtDNA) mutations at nps 3460, 11778, 14484, and 14459 account for roughly 90% of cases of Leber hereditary optic neuropathy (LHON) and are designated as “primary” LHON mutations since they act as major predisposition factors for LHON. Although each primary mutation can arise independently on different mtDNA backgrounds during human evolution, they characteristically do not co‐occur in LHON patients. We report here a family with the simultaneous occurrence of the 11778A and 14484C mutations. Neuro‐ophthalmological examination of the proband, a nine‐year‐old Caucasian female, revealed the bilateral optic atrophy, central scotomas, and reduced visual acuity typical of LHON. Her mother had normal appearing optic discs and is today visually asymptomatic. Analysis of the proband blood mtDNA revealed that she harbored both the 11778A (heteroplasmic, 94% mutant) and the 14484C (homoplasmic mutant) mutation. This genotype was maintained in proband lymphoblasts and transmitochondrial cybrids. The mother also had both mutations, with the 14484C mutation homoplasmic in all cell types examined. However, only 31% of her blood mtDNAs carried the 11778 mutation, which segregated to essentially 100% wild‐type in lymphoblast and cybrid mtDNA. Complex I‐linked respiration and specific enzyme activity were consistently lowest in proband lymphoblast and cybrid mitochondria compared to those from the mother, 11778A patients, 14484C patients, or controls, thus demonstrating both a deleterious synergistic interaction between the 11778A and 14484C mutations and the magnitude of 11778A‐associated complex I dysfunction. Remarkably, spontaneous vision recovery occurred in the proband, highlighting the complexities encountered when associating mtDNA genotype and complex I function with LHON expression. © 2001 Wiley‐Liss, Inc.     

Clinical, genetic, and biochemical characterization of a Leber hereditary optic neuropathy family containing both the 11778 and 14484 primary mutations.

Four mitochondrial DNA (mtDNA) mutations at nps 3460, 11778, 14484, and 14459 account for roughly 90% of cases of Leber hereditary optic neuropathy (LHON) and are designated as "primary" LHON mutations since they act as major predisposition factors for LHON. Although each primary mutation can arise independently on different mtDNA backgrounds during human evolution, they characteristically do not co-occur in LHON patients. We report here a family with the simultaneous occurrence of the 11778A and 14484C mutations. Neuro-ophthalmological examination of the proband, a nine-year-old Caucasian female, revealed the bilateral optic atrophy, central scotomas, and reduced visual acuity typical of LHON. Her mother had normal appearing optic discs and is today visually asymptomatic. Analysis of the proband blood mtDNA revealed that she harbored both the 11778A (heteroplasmic, 94% mutant) and the 14484C (homoplasmic mutant) mutation. This genotype was maintained in proband lymphoblasts and transmitochondrial cybrids. The mother also had both mutations, with the 14484C mutation homoplasmic in all cell types examined. However, only 31% of her blood mtDNAs carried the 11778 mutation, which segregated to essentially 100% wild-type in lymphoblast and cybrid mtDNA. Complex I-linked respiration and specific enzyme activity were consistently lowest in proband lymphoblast and cybrid mitochondria compared to those from the mother, 11778A patients, 14484C patients, or controls, thus demonstrating both a deleterious synergistic interaction between the 11778A and 14484C mutations and the magnitude of 11778A-associated complex I dysfunction. Remarkably, spontaneous vision recovery occurred in the proband, highlighting the complexities encountered when associating mtDNA genotype and complex I function with LHON expression.     

Asian-specific mtDNA backgrounds associated with the primary G11778A mutation of Leber's hereditary optic neuropathy

We studied 19 patients of Southeast Asian (SEA) ethnic ancestry with Leber's hereditary optic neuropathy (LHON) to investigate the mtDNA haplotypes associated with the primary mutation(s). Eighteen patients carried a mitochondrial DNA (mtDNA) G11778A mutation (Arg340His in the respiratory complex I ND4 subunit), while one had a T14484C mutation (Met64Val in the ND6 subunit). One patient had a class II LHON mtDNA mutation, G3316A. Sequencing data of the ND genes showed many single-nucleotide polymorphisms (62 SNPs in 17 individuals; 10 LHON patients and 7 normal controls) not previously reported in Europeans or Japanese. The SEA G11778A LHON mutation was associated mostly with two mtDNA haplogroups, M (47%) and a novel lineage, characterized by the gain of a 10394 DdeI site but absence of the 10397 AluI site, designated BM (37%). A significant association was observed between one SNP, A10398G, resulting in a Thr114Ala substitution in the ND3 subunit, and the primary LHON mutation. This SNP also characterizes haplogroup J, with which the European LHON 11778 and 14484 mutations show preferential association. The combination of A10398G and other SNPs, specific for the haplogroups J, M, or BM, might act synergistically to increase the penetrance of the LHON mutations, thus allowing their detection. Received: June 6, 2002 / Accepted: August 23, 2002 Acknowledgments We thank Dr. Mulia Sitepu, School of Medicine, University of Atmajaya, Jakarta, Drs. Norma Handoyo and Inakawati Rivai of the School of Medicine, University of Diponegoro, Semarang, and Dr. Tjahyono Ghondowiardjo of the School of Medicine, University of Indonesia, for referring their patients for DNA analysis. This work was supported by grants from PT Krakatau Steel and PT Inti through the Agency for Strategic Industries (Indonesia) and by a generous development fund from the National Development Planning Agency (BAPPENAS) of the Republic of Indonesia. Correspondence to:S. Marzuki     

Leber's hereditary optic neuropathy with 14484 mutation in Central Java,Indonesia

Leber's hereditary optic neuropathy (LHON) is a maternally inherited late-onset form of blindness characterized by acute or subacute bilateral retinal degradation resulting in a permanent loss of central vision. G11778A, C3460A, and T14484C mutations on mitochondrial DNA (mtDNA) are specific for LHON and account for most, but not all, worldwide LHON cases. A six-generation Indonesian LHON family with the T14484C mutation was analyzed. Polymerase chain reaction/restriction fragment length polymorphism analysis showed that all of the maternal lineages had the T14484C mutation in a homoplasmic form. Penetrance of the disease (33.3%) and male predominance (3:1) was similar to other worldwide LHON with the T14484C mutation. The incidence of offspring born to affected mothers was no different from that of unaffected mothers, and the age distribution of cases was no higher than that of asymptomatic carriers. Eight secondary mutations were sought but not detected. The patients of this family belonged to haplogroup M. These findings support the idea that the mtDNA backgrounds involved in the expression of LHON mutations in southeast Asians are different from those of Europeans.T. Nishioka and M. Tasaki contributed equally to this work     

Deciphering exome sequencing data: Bringing mitochondrial DNA variants to light

The expanding use of exome sequencing (ES) in diagnosis generates a huge amount of data, including untargeted mitochondrial DNA (mtDNA) sequences. We developed a strategy to deeply study ES data, focusing on the mtDNA genome on a large unspecific cohort to increase diagnostic yield. A targeted bioinformatics pipeline assembled mitochondrial genome from ES data to detect pathogenic mtDNA variants in parallel with the “in‐house” nuclear exome pipeline. mtDNA data coming from off‐target sequences (indirect sequencing) were extracted from the BAM files in 928 individuals with developmental and/or neurological anomalies. The mtDNA variants were filtered out based on database information, cohort frequencies, haplogroups and protein consequences. Two homoplasmic pathogenic variants (m.9035T>C and m.11778G>A) were identified in 2 out of 928 unrelated individuals (0.2%): the m.9035T>C (MT‐ATP6) variant in a female with ataxia and the m.11778G>A (MT‐ND4) variant in a male with a complex mosaic disorder and a severe ophthalmological phenotype, uncovering undiagnosed Leber's hereditary optic neuropathy (LHON). Seven secondary findings were also found, predisposing to deafness or LHON, in 7 out of 928 individuals (0.75%). This study demonstrates the usefulness of including a targeted strategy in ES pipeline to detect mtDNA variants, improving results in diagnosis and research, without resampling patients and performing targeted mtDNA strategies.     

Phylogenetic analysis of Leber's hereditary optic neuropathy mitochondrial DNA's indicates multiple independent occurrences of the common mutations

The mitochondrial DNAs (mtDNA) from 17 Caucasian 11778-positive and 30 Caucasian 11778-negative Leber's hereditary optic neuropathy (LHON) patients were PCR-amplified and subjected to high resolution restriction endonuclease analysis. Concurrently, all patient mtDNAs were screened for the common primary LHON mtDNA mutations at nucleotide pairs (nps) 3460, 11778, and 14484, the ambiguous intermediate-risk LHON mtDNA mutations at nps 5244 and 15257, and the secondary LHON mtDNA mutations at nps 3394, 4216, 4917, 7444, 13708, and 15812. Phylogenetic analysis was performed using mtDNA haplotype data from the 47 LHON patients and 175 non-LHON Caucasian controls. The superimposition of the LHON mutation screening results upon the Caucasian mtDNA phylogeny revealed (1) 35 different LHON haplotypes, (2) that all three common primary mutations have occurred multiple times in Caucasians, (3) that while recurrent mutation is common for the primary mutations, secondary mutations tend to be lineage-specific, (4) that the np 15257 mutation was confined to a single mtDNA lineage but may be etiologically important in some LHON cases since it was found in a LHON pedigree which lacked a common primary mutation; complete sequence analysis of the proband mtDNA revealed only a single other candidate missense mutation (at np 10663 of the ND4L gene) of uncertain pathological significance; and (5) that the np 14484 mutation may be less pathogenic than either the np 3460 or np 11778 mutations, as this mutation most commonly occurred on a single mtDNA lineage and almost always in association with secondary LHON mutations. A phylogenetic ageneous disease has thus provided key genetic data bearing on the relative pathogenicity of the LHON-associated mtDNA mutations. © 1995 Wiley-Liss, Inc.     

Mitochondrial DNA background modulates the assembly kinetics of OXPHOS complexes in a cellular model of mitochondrial disease

Leber's hereditary optic neuropathy (LHON), the most frequent mitochondrial disorder, is mostly due to three mitochondrial DNA (mtDNA) mutations in respiratory chain complex I subunit genes: 3460/ND1, 11778/ND4 and 14484/ND6. Despite considerable clinical evidences, a genetic modifying role of the mtDNA haplogroup background in the clinical expression of LHON remains experimentally unproven. We investigated the effect of mtDNA haplogroups on the assembly of oxidative phosphorylation (OXPHOS) complexes in transmitochondrial hybrids (cybrids) harboring the three common LHON mutations. The steady-state levels of respiratory chain complexes appeared normal in mutant cybrids. However, an accumulation of low molecular weight subcomplexes suggested a complex I assembly/stability defect, which was further demonstrated by reversibly inhibiting mitochondrial protein translation with doxycycline. Our results showed differentially delayed assembly rates of respiratory chain complexes I, III and IV amongst mutants belonging to different mtDNA haplogroups, revealing that specific mtDNA polymorphisms may modify the pathogenic potential of LHON mutations by affecting the overall assembly kinetics of OXPHOS complexes.     

中国人Leber 遗传性视神经病变的原发突变及临床特征

目的 分析中国人Leber遗传性视神经病变(Leber’s hereditary optic neuropathy,LHON)线粒体DNA3个原发致病基因突变遗传及其临床特征。方法 分别用突变特异性引物聚合酶链反应,异源双链-单链构象多态性,限制性片段长度多态性和DNA测序方法,对110个家系的156例LHON患者进行11778A、3460A、14484C 3个原发位点检测,并收集患者病史及其临床资料,进行统计学分析。结果 110例LHON先证者中,11778位点突变者100例,占90．9％;3460位点突变者2例,占1．8％;14484位点突变者8例,占7,3％。不同突变位点的LHON患者发病时视力分布：125人(250眼)11778位点突变患眼中发病时视力≤0．01(占17．6％),视力介于0.01至0．1之间(占52．1％),视力≥0．1(占30．3％);28人(56眼)14484位点突变患眼中无患眼视力低于0．01视力介于001至0．1之间(占12．7％),视力≥0．1(占87．3％);3人(6眼)3460位点突变患眼视力均介于0．03至0．08之间。视力恢复情况：250只11778位点突变的患眼中,6．97％的眼视力有所恢复,平均最终视力0．03(指数～0．07);56只14484位点突变的患眼中,50％的眼视力有所恢复,占50％,平均最终视力0．8(0．3～1．2)。结论 中国人LHON患者mtDNA 3个原发致病突变中,以11778A位点突变为主、14484C位点突变较少、3460A罕见。LHON的临床表现与致病突变位点有关,14484C突变患者的发病视力及视力恢复情况明显好于11778A患者。     

A study of 133 Chinese children with mitochondrial respiratory chain complex I deficiency

To investigate the clinical, enzymological and mitochondrial gene profiles of complex I deficiency in Chinese, clinical and laboratory data of the patients (79 boys, 54 girls) were retrospectively assessed. Activities of mitochondrial respiratory chain complexes in peripheral leucocytes were spectrophotometrically measured. The entire mitochondrial DNA (mtDNA) sequence was analyzed in 62 patients. Restriction fragment length polymorphism and gene sequencing analyses were performed in 15 families. Ninety‐one patients had isolated complex I deficiency; 42 had combined deficiencies of complex I and other complexes. The main clinical presentations were neuromuscular disorders (107 patients) and non‐neurological dysfunction (hepatopathy, renal damage and cardiomyopathy; 26 patients). In 32 of 62 patients who underwent mtDNA sequencing, 24 mutations were identified in 15 mitochondrial genes. The 12338T>C, 4833A>G and 14502T>C mutations were found in 12.9%, 11.3% and 4.8% patients, respectively. Seven patients had multiple mutations. Three novel mutations were identified. Chinese patients with complex I deficiency presented heterogeneous phenotypes and genotypes. Twenty‐four mutations were identified in 15 mitochondrial genes in 51.6% patients. mtDNA mutations were more common in isolated complex I deficiency than in combined complex deficiencies. The 12338T>C, 4833A>G and 14502T>C mutations were common.     

A simple oligonucleotide biochip capable of rapidly detecting known mitochondrial DNA mutations in Chinese patients with Leber's hereditary optic neuropathy (LHON)

Leber's hereditary optic neuropathy (LHON) is a maternally transmitted disease. Clinically, no efficient assay protocols have been available. In this study, we aimed to develop an oligonucleotide biochip specialized for detection of known base substitution mutations in mitochondrial DNA causing LHON and to investigate frequencies of LHON relevant variants in Anhui region of China. Thirty-two pairs of oligonucleotide probes matched with the mutations potentially linked to LHON were covalently immobilized. Cy5-lablled targets were amplified from blood DNA samples by a multiplex PCR method. Two kinds of primary mutations 11778 G > A and 14484 T > C from six confirmed LHON patients were interrogated to validate this biochip format. Further, fourteen Chinese LHON pedigrees and twenty-five unrelated healthy individuals were investigated by the LHON biochip, direct sequencing and pyrosequencing, respectively. The biochip was found to be able efficiently to discriminate homoplasmic and heteroplasmic mtDNA mutations in LHON. Biochip analysis revealed that twelve of eighteen LHON symptomatic cases from the 14 Chinese pedigree harbored the mutations either 11778G > A, 14484T > C or 3460G A, respectively, accounting for 66.7%. The mutation 11778G > A in these patients was homoplasmic and prevalent (55.5%, 10 of 18 cases). The mutations 3460G > A and 3394T > C were found to co-exist in one LHON case. The mutation 13708G > A appeared in one LHON pedigree. Smaller amount of sampling and reaction volume, easier target preparation, fast and high-throughput were the main advantages of the biochip over direct DNA sequencing and pyrosequencing. Our findings suggested that primary mutations of 11778G > A, 14484T > C or 3460G > A are main variants of mtDNA gene leading to LHON in China. The biochip would easily be implemented in clinical diagnosis.     

Poor outcome in Down syndrome fetuses with cardiac anomalies or growth retardation

The penetrance in Leber's hereditary optic neuropathy (LHON) pedigrees is determined primarily by a mutation in the mitochondrial genome (mtDNA), but secondary factors are also necessary for manifestation of the disorder. It has been proposed that mtDNA polymorphisms affect penetrance in LHON pedigrees. In particular, it has been postulated that one or more polymorphisms associated with European haplogroup J mtDNAs substantially increase the penetrance of the primary LHON mutation at nucleotide 14484. We report here a haplogroup H matrilineal pedigree (VIC14) in which the single affected member carries the 14484 LHON mutation, but who manifested a milder and atypical optic nerve disorder. In addition, during a population screen, we identified an individual who carried the 14484 mutation but who had normal vision. Finally, the 14484 mutation is under-represented among haplogroup H mtDNAs that carry a LHON mutation. These results, in conjunction with other studies that are reviewed, indicate that 14484 LHON mutations have a low penetrance when they arise in a haplogroup H mtDNA background.     

Fine Time Scaling of Purifying Selection on Human Nonsynonymous mtDNA Mutations Based on the Worldwide Population Tree and Mother–Child Pairs

A high‐resolution mtDNA phylogenetic tree allowed us to look backward in time to investigate purifying selection. Purifying selection was very strong in the last 2,500 years, continuously eliminating pathogenic mutations back until the end of the Younger Dryas (～11,000 years ago), when a large population expansion likely relaxed selection pressure. This was preceded by a phase of stable selection until another relaxation occurred in the out‐of‐Africa migration. Demography and selection are closely related: expansions led to relaxation of selection and higher pathogenicity mutations significantly decreased the growth of descendants. The only detectible positive selection was the recurrence of highly pathogenic nonsynonymous mutations (m.3394T>C‐m.3397A>G‐m.3398T>C) at interior branches of the tree, preventing the formation of a dinucleotide STR (TATATA) in the MT‐ND1 gene. At the most recent time scale in 124 mother–children transmissions, purifying selection was detectable through the loss of mtDNA variants with high predicted pathogenicity. A few haplogroup‐defining sites were also heteroplasmic, agreeing with a significant propensity in 349 positions in the phylogenetic tree to revert back to the ancestral variant. This nonrandom mutation property explains the observation of heteroplasmic mutations at some haplogroup‐defining sites in sequencing datasets, which may not indicate poor quality as has been claimed.     

The molecular basis of cystathionine ß‐synthase (CBS) deficiency in UK and US patients with homocystinuria

The molecular basis of cystathionine ß‐synthase (CBS) deficiency has been studied in 536 patient alleles with 130 different mutations described. To date, no study has reported on the incidence of any of the reported mutations in patients from the UK and the US. We developed a new antisense oligonucleotide (ASO) PCR/hybridization method to screen for 12 of the most frequent CBS mutations in 14 unrelated patients from the UK and 38 unrelated patients from the US, a total of 104 independent alleles. We determined 16/28 (57%) and 28/76 (37%) of the affected alleles in the UK and US patients, respectively. Four different mutations were identified in the UK patients (c.374G>A, R125Q; c.430G>A, E144K; c.833T>C, I278T; c.919G>A, G307S) and 8 mutations identified in the patients from the US (c.341C>T, A114V; c.374G>A, R125Q; c.785C>T, T262M; c.797G>A, R266K; c.833T>C, I278T; c.919G>A, G307S; g.13217A>C (del ex 12); c.1330G>A, D444N). The I278T was the predominant mutation in both populations, present in 8 (29%) of 28 independent alleles from the UK and in 14 (18%) of 76 independent alleles from the US. The incidence of the G307S mutation was 21% in the UK patients and 8% in the US patients. The spectrum of mutations observed in the patients from the UK and US is closer to that which is observed in Northern Europe and bears less resemblance to that observed in Ireland. © 2003 Wiley‐Liss, Inc.     

Population genetics and disease susceptibility: characterization of central European haplogroups by mtDNA gene mutations, correlation with D loop variants and association with disease

Mitochondrial (mt)DNA haplogroups in a German control group (n = 67) were characterized by screening mitochondrial coding regions encompassing most of the ND, tRNA and cyt b genes. We used a PCR-SSCP screening approach followed by direct sequencing of polymorphic mtDNA fragments. Five major mtDNA lineages, diverging in at least nine different haplogroups, could be defined by characteristic polymorphic sites in mitochondrial genes. Additional sequencing of two hypervariable segments (HVS-I and II) of the non-coding displacement (D) loop in all control subjects revealed that certain D loop variants were strongly correlated with lineages and haplogroups, while others represented hotspots occurring frequently in different haplogroups. The existence of identified lineages and haplogroups received support from data in the literature, obtained by use of different approaches. Subsequently, we investigated four disease groups for association with these haplogroups: (i) LHON patients (n = 55) carrying at least one of the primary/intermediate LHON mutations at nt 3460, 11778, 14484 and/or 15257; (ii) patients suffering from Wolfram or DIDMOAD syndrome (n = 8); (iii) MELAS patients (n = 9); (iv) a group of children, who died from 'sudden infant death syndrome' (SIDS) (n = 9). The distribution patterns among the haplogroups of the disease groups (LHON, DIDMOAD and SIDS) differed considerably from the control population. LHON and DIDMOAD were significantly under-represented in the most frequent German haplogroup DC, but were concentrated in a mtDNA lineage defined by polymorphisms at nt 4216 + 11251 + 16126. As this lineage diverged into two precisely defined haplogroups, LHON and DIDMOAD could be assigned to the two haplogroups separately. Strikingly, SIDS was often found in association with two rare German haplogroups. MELAS patients were equally distributed among German haplogroups and, moreover, did not reveal any accumulation of specific D loop variants. We conclude that certain European mtDNA haplogroups define a genetic susceptibility basis for various disorders.      

Leber遗传性视神经病变的临床实践指南

Leber遗传性视神经病变(Leber’s hereditary optic neuropathy,LHON)为母系遗传病,线粒体基因组m.11778G>A、m.14484T>C和m.3460G>A突变是其主要的分子基础,但其发病亦受到核基因、线粒体遗传背景和环境因素的共同影响。本指南的编写在参考国内外相关领域的基础和临床研究以及其他国家发布的指南和共识的基础上,结合了中国人群的实际资料,旨在总结关于LHON的遗传学知识和临床处置要点,以提高临床医师对该病的诊断水平,并为患者临床管理的规范化提供建议。     

Leber''s hereditary optic neuropathy: correlations between mitochondrial genotype and visual outcome.

Leber's hereditary optic neuropathy (LHON) is a maternally inherited disease associated with mitochondrial DNA (mtDNA) mutations. We describe the distribution of seven different mtDNA mutations and the clinical findings in 334 LHON patients belonging to 29 families. Mutations described only in LHON at nucleotide positions 11778, 3460, and 14484 were found in 15, two, and nine families respectively. In three families none of these mutations was found. Mutations described in LHON but also in controls at nucleotide positions 15257, 13708, 4917, and 4216 were found in one, 10, three and 12 families respectively. Combinations of mtDNA mutations were found in most families. The patient population mainly consisted of 79.2% to 89.5% males except for one family with only 10 of 17 patients being males (58.9%, p approximately 0.036). In 11 families only the 11778 mutation was found; in this group (WX) the affected males had a mean age of onset of 29.2 years and a mean visual outcome of 0.113. In seven families the 14484, 13708, and 4216 mutations were found; in this group (MA) the affected males had a mean age of onset of 22.0 years and a mean visual outcome of 0.442. In two families no mutation was found at all; in this group (YX) the affected males had a mean age of onset of 18.9 years and a mean visual outcome of 0.167. The mean age of onset in the WX group is significantly higher than in the MA group (p < or = 0.001) and in the YX group (p approximately 0.01). The mean visual outcome in the MA group is significantly better than in the WX group (p </= 0.001) and the YX group (p = 0.05). No significant clinical differences were found between families exhibiting only the 11778 mutation and those with additional mutations at np 13708, 4917, or 4216, suggesting that these mutations are of little phenotypic importance. Other mutations were present in relatively small numbers of patients. These results show that the clinical severity is dependent on the mitochondrial genotype.     

线粒体tRNAThr A15951G可能是与Leber遗传性视神经病变相关的基因突变

目的 进一步分析中国汉族Leber遗传性视神经病变(Leber's hereditary optic neuropathy,LHON)家系的临床和分子遗传学特征,阐明LHON的分子致病机制.方法 对2例具有典型LHON临床特征的先证者和家系其他成员进行眼科学及其临床检查.对这2个家系先证者使用24对有部分重叠的引物进行线粒体DNA(mitochondrial DNA,mtDNA)全序列扩增分析.结果 检查发现这些家系成员中视力损害的外显率分别为5.3％(1/19)、18.2％(4/22).经mtDNA测序分析,并没有发现mtDNA G11778A、G3460A和T14484C 3个常见的突变,在tRNAThr上发现了A15951G同质性突变位点.线粒体DNA全序列分析显示2个家系呈现mtDNA多态性,都属于东亚单倍型D4b1.A15951G突变位于线粒体tRNAThr高度保守区(通用位点为71位),可能导致tRNA空间结构和稳定性发生改变,线粒体蛋白合成功能受损,最终发生视力损害.结论 线粒体tRNAThr A15951G可能是与Leber遗传性视神经病变相关的致病性线粒体基因突变.     

Mutational and phenotypical spectrum of phenylalanine hydroxylase deficiency in Denmark

We describe the genotypes of the complete cohort, from 1967 to 2014, of phenylketonuria (PKU) patients in Denmark, in total 376 patients. A total of 752 independent alleles were investigated. Mutations were identified on 744 PKU alleles (98.9%). In total, 82 different mutations were present in the cohort. The most frequent mutation c.1315+1G>A (IVS12+1G>A) was found on 25.80% of the 744 alleles. Other very frequent mutations were c.1222C>T (p.R408W) (16.93%) and c.1241A>G (p.Y414C) (11.15%). Among the identified mutations, five mutations; c.532G>A (p.E178K), c.730C>T (p.P244S), c.925G>A (p.A309T), c.1228T>A (p.F410I), and c.1199+4A>G (IVS11+4A>G) have not been reported previously. The metabolic phenotypes of PKU are classified into four categories; ‘classical PKU’, ‘moderate PKU’, ‘mild PKU’ and ‘mild hyperphenylalaninemia’. In this study, we assigned the phenotypic outcome of three of the five novel mutations and furthermore six not previously classified mutations to one of the four PKU categories.     

Two mtDNA mutations 14487T>C (M63V,ND6) and 12297T>C (tRNA Leu) in a Leigh syndrome family

Mitochondrial cytopathies are characterized by a large variability of clinical phenotypes and severity. The 14487T>C mutation in mtDNA has been recently described to be associated with Leigh syndrome. The 12297T>C mutation has been described in isolated dilated cardiomyopathy patients. Here, we report a family with multiple members who harbor both mutations, with only a few individuals who are affected with Leigh syndrome. Mitochondrial whole genome sequencing analysis in the proband’s muscle specimen detected two nearly homoplasmic mutations: 14487T>C (M63V in ND6) and 12297T>C in the tRNA ^Leu (CUN) gene. These two mutations were also detected in the blood, urine sediments, hair follicles, and buccal swab samples of all matrilineal relatives tested. All individuals tested were nearly homoplasmic for the 12297T>C mutation, but had variable degrees of heteroplasmy for 14487T>C. We also screened for the frequency of these two mutations. Of 268 patients with Leigh or Leigh-like disease, one case was found to harbor the 14487T>C mutation (0.3%), and one had the 12297T>C mutation (0.3%). Neither mutation was detected in the 88 patients meeting MELAS syndrome criteria nor in the 56 patients with respiratory chain complex I or I + III deficiency. In conclusion, the 14487T>C mutation appears as the primary etiology of Leigh syndrome in this family, demonstrating the high level of heteroplasmy needed for a clinically significant phenotype with this mutation. The 12297T>C mutation was not associated with dilated cardiomyopathy for the family members who were clinically evaluated and who were shown by testing to be nearly homoplasmic for that mutation.     

Mitochondrial DNA haplogroup diversity in Basques: a reassessment based on HVI and HVII polymorphisms.

This study provides a more complete characterization of the mitochondrial genome variability of the Basques, including data on the hypervariable segment HVII of the D-loop region, which remains relatively unknown. To that end, genomic DNA from 55 healthy men living in the Arratia Valley (Biscay province) and the Goiherri region (Guipúzcoa province) was examined by direct sequencing. Three-generation pedigree charts were compiled to ensure the collection from autochthonous individuals. The most notable findings emerging from the analysis of haplogroup composition are: (i) lack of U8a mitochondrial lineage, a rare subhaplogroup recently identified in Basques and proposed as a Paleolithic marker, (ii) low frequency of haplogroup V, which conflicts with results of earlier analyses describing high frequencies in southwestern Europe, and (iii) high frequency of haplogroup J, especially subhaplogroups J1c1 and J2a. The frequency of haplogroup J does not coincide with previous mtDNA studies in present-day Basques, but is congruent with frequencies found in prehistoric and historic Basque populations. In explaining divergence in haplogroup composition between modern Basque samples, we hypothesized spatial heterogeneity promoted by population fragmentation due to extreme limitation of dispersal opportunities during the Pleistocene glaciations. Similarities between extinct and extant Basque populations as for the high frequency of lineage J, as well as the abundance of this haplogroup in northern Spain endorse a shift in the focus of attention of mtDNA analysts. A refined dissection of haplogroup J might provide more solid evidence about the process of postglacial recolonization of Europe, and thus about the shaping of the European gene pool.