Mitochondrial DNA mutation at np 3243 in a family with maternally inherited diabetes mellitus

Mitochondrial DNA (mtDNA) gene defects may play a role in the development of maternally inherited diabetes mellitus and deafness (MIDD). A family from Southern Italy who showed maternal transmission of type 2 diabetes mellitus with three individuals affected is described. A 10.4 kb deletion and mutations at nucleotide positions (np) 3243, 7445 and 11778 in the mtDNA of six relatives were sought. The mitochondrial np 3243 mutation of the tRNA Leu (UUR) gene was identified in a boy affected by optic atrophy and mental retardation, as well as in his diabetic mother. No other mutations or deletions were found. Our study points out the variable phenotypic expression of the np 3243 mtDNA mutation. This may suggest the presence of other mitochondrial or nuclear mutations required to modulate the phenotype. A clinical and metabolic follow-up of all family members was necessary to understand the role of the np 3243 mutation, especially in one child affected by optic atrophy and mental retardation. Further studies will be aimed at investigating the prevalence of mutations and deletions of mtDNA in type 2 diabetes mellitus. Received: 20 November 1998 / Accepted in revised form: 20 July 1999     

Maternally Inherited Diabetes Mellitus: the Role of Mitochondrial DNA Defects

Several studies have shown a consistent maternal effect in the transmission of Type 2 diabetes (NIDDM). The mitochondrial encephalomyopathies are a group of diseases characterized by maternal inheritance and a variety of mitochondrial DNA defects. Diabetes is a feature of some of these disorders and therefore the hypothesis arose that mitochondrial DNA mutations might play a role in patients with diabetes but no other features of neurological disease. Recent studies have confirmed that a specific point mutation in the gene encoding the mitochondrial tRNA for leucine segregates with diabetes and nerve deafness in families from the UK, Holland, France and Japan. Mitochondrial gene deletions have also been reported. Affected subjects present with progressive insulin deficiency and may fall into the broad classifications of either Type 1 (IDDM) or Type 2 diabetes (NIDDM). Future studies are aimed at searching for other mitochondrial gene defects in diabetes and attempting to explain the mechanism of hyperglycaemia by the development of phenotypic expression systems. Although an exciting development in the genetics of diabetes, currently described mitochondrial mutations do not fully explain the maternal effect in the transmission of Type 2 diabetes.     

Mitochondrial medicine – recent advances

Mitochondria contain the respiratory chain enzyme complexes that carry out oxidative phosphorylation and produce the main part of cellular energy in the form of ATP. Mitochondrial DNA (mtDNA) encodes essential subunits of the respiratory chain and is thus critical for maintaining cellular energy production. The first pathogenic mtDNA mutations were reported in 1988, and today more than 50 disease-causing mtDNA mutations have been identified. In addition, mtDNA mutations have been implicated in ageing and in common disorders such as diabetes mellitus, heart failure and Parkinson's disease. This review will summarize recent advances in the rapidly expanding field of mitochondrial medicine.     

Mitochondrial gene defects in patients with NIDDM

Summary Non-insulin-dependent diabetes mellitus (NIDDM) has a strong genetic component and maternal factors have recently been implicated in disease inheritance. The mitochondrial myopathies are a group of diseases which often show maternal inheritance as a result of mtDNA defects; some patients have impaired glucose tolerance. Occasional families with maternally inherited diabetes and deafness associated with a deletion or point mutation of mtDNA have been reported. To assess the importance of mitochondrial gene defects in NIDDM, 150 unrelated diabetic subjects from Wales, UK and 68 unrelated patients with diabetes and at least one affected sibling from England, UK were studied. Southern blot analysis did not show any large mtDNA deletions or duplications. One patient had a mutation in the mitochondrial tRNA^leu(UUR) gene at bp 3243. This mutation is commonly associated with the syndrome of mitochondrial encephalomyopathy, lactic acidosis and stroke like episodes (MELAS). Study of this patient and his siblings showed a distinct form of late-onset diabetes associated with nerve deafness but no clinical features of the MELAS syndrome. No diabetic subject was shown to have the mtDNA mutation at position 8344 (tRNA^lys) which has previously been described in the syndrome of mitochondrial encephalomyopathy and red-ragged fibres (MERRF). The role of other mitochondrial gene defects in diabetes and the pathophysiological basis of glucose intolerance in patients with the MELAS mutation requires further elucidation.Abbreviations mtDNA mitochondrial DNA - tRNA transfer RNA - NIDDM non-insulin-dependent mellitus - bp base pair - PCR polymerase chain reaction     

Autosomal dominant transmission of diabetes and congenital hearing impairment secondary to a missense mutation in the WFS1 gene

Aims Mutations of the WFS1 gene have been implicated in autosomal dominant diseases, such as low‐frequency sensorineural hearing impairment (LFSNHI) and/or diabetes mellitus and/or optic atrophy. The aim was to investigate WFS1 gene sequences in a family with diabetes mellitus and hearing impairment. Methods Three members of a family with a maternally inherited combination of diabetes mellitus and hearing impairment, but no specific mutations in its mitochondrial genome, were investigated for mutations in the WFS1 gene. Results This pedigree, in which the proband had non‐insulin‐dependent diabetes mellitus and congenital hearing impairment and his mother a triple combination of diabetes mellitus, hearing impairment and optic atrophy, was found to be associated with autosomal dominant transmission of the E864K mutation of the WFS1 gene. Conclusions In the light of this confirmatory study, we recommend the systematic analysis of WFS1 gene sequences in patients with parentally inherited diabetes mellitus and deafness (± optic atrophy), in particular when diabetogenic mtDNA mutations have been excluded.     

The mitochondrial tRNALeu(UUR) A to G 3243 mutation is associated with insulin-dependent and non-insulin-dependent diabetes in a Chinese population

The mitochondrial DNA tRNA^Leu(UUR) A to G 3243 mutation is associated with maternally inherited diabetes in Caucasians and Japanese. In a Hong Kong Chinese population we have detected the 3243 mutation in 2 of 74 unrelated subjects with well characterized insulin-dependent (Type 1) diabetes mellitus (IDDM) and 2 of 75 unrelated subjects with young onset (<35 years) non-insulin-dependent diabetes (NIDDM). The 3243 mutation has only previously been associated with IDDM in Japanese. Racial differences in association of the mitochondrial 3243 mutation with IDDM suggest the influence of other genes that may increase its diabetogenic pathogenicity in Oriental races. We also found a significant excess of maternal inheritance of diabetes in the young onset NIDDM cohort, with a ratio of diabetic mothers to fathers of 2.4:1, p < 0.005. The 3243 mutation, however, only accounts for a small proportion of the observed excess maternal inheritance, and further study is needed to search for other diabetes associated mitochondrial DNA mutations. © 1997 John Wiley & Sons, Ltd.     

Mitochondrial dysfunction as a cause of ageing

Mitochondrial dysfunction is heavily implicated in the ageing process. Increasing age in mammals correlates with accumulation of somatic mitochondrial DNA (mtDNA) mutations and decline in respiratory chain function. The age-associated respiratory chain deficiency is typically unevenly distributed and affects only a subset of cells in various human tissues, such as heart, skeletal muscle, colonic crypts and neurons. Studies of mtDNA mutator mice has shown that increased levels of somatic mtDNA mutations directly can cause a variety of ageing phenotypes, such as osteoporosis, hair loss, greying of the hair, weight reduction and decreased fertility. Respiratory-chain-deficient cells are apoptosis prone and increased cell loss is therefore likely an important consequence of age-associated mitochondrial dysfunction. There is a tendency to automatically link mitochondrial dysfunction to increased generation of reactive oxygen species (ROS), however, the experimental support for this concept is rather weak. In fact, respiratory-chain-deficient mice with tissue-specific mtDNA depletion or massive increase of point mutations in mtDNA typically have minor or no increase of oxidative stress. Mitochondrial dysfunction is clearly involved in the human ageing process, but its relative importance for mammalian ageing remains to be established.     

A transgenic model to study the pathogenesis of somatic mtDNA mutation accumulation in beta-cells

Low levels of somatic mutations accumulate in mitochondrial DNA (mtDNA) as we age; however, the pathogenic nature of these mutations is unknown. In contrast, mutational loads of >30% of mtDNA are associated with electron transport chain defects that result in mitochondrial diseases such as mitochondrial encephalopathy lactic acidosis and stroke-like episodes. Pancreatic beta-cells may be extremely sensitive to the accumulation of mtDNA mutations, as insulin secretion requires the mitochondrial oxidation of glucose to CO(2). Type 2 diabetes arises when beta-cells fail to compensate for the increased demand for insulin, and many type 2 diabetics progress to insulin dependence because of a loss of beta-cell function or beta-cell death. This loss of beta-cell function/beta-cell death has been attributed to the toxic effects of elevated levels of lipids and glucose resulting in the enhanced production of free radicals in beta-cells. mtDNA, localized in close proximity to one of the major cellular sites of free radical production, comprises more than 95% coding sequences such that mutations result in changes in the coding sequence. It has long been known that mtDNA mutations accumulate with age; however, only recently have studies examined the influence of somatic mtDNA mutation accumulation on disease pathogenesis. This article will focus on the effects of low-level somatic mtDNA mutation accumulation on ageing, cardiovascular disease and diabetes.     

A common mitochondrial DNA variant is associated with insulin resistance in adult life

Summary Mitochondrial DNA is maternally inherited. Mitochondrial DNA mutations could contribute to the excess of maternal over paternal inheritance of non-insulin-dependent diabetes mellitus (NIDDM). We therefore investigated the relationship between this variant, insulin resistance and other risk factors in a cohort which had been well characterised with respect to diabetes. Blood DNA was screened from 251 men born in Hertfordshire 1920–1930 in whom an earlier cohort study had shown that glucose tolerance was inversely related to birthweight. The 16 189 variant (T- > C transition) in the first hypervariable region of mitochondrial DNA was detected using the polymerase chain reaction and restriction digestion. DNA analysis showed that 28 of the 251 men (11 %) had the 16 189 variant. The prevalence of the 16 189 variant increased progressively with fasting insulin concentration (p < 0.01). The association was independent of age and body mass index and was present after exclusion of the patients with NIDDM or impaired glucose tolerance. We found that insulin resistance in adult life was associated with the 16 189 variant. This study provides the first evidence that a frequent mitochondrial variant may contribute to the phenotype in patients with a common multifactorial disorder. [Diabetologia (1998) 41: 54–58] Received: 20 May 1997 and in revised form: 7 August 1997     

Molecular and clinical aspects of mitochondrial diabetes mellitus.

This review provides a compact overview on the contribution of mutations in mtDNA to the pathogenesis of diabetes mellitus, with emphasis on the A3243G mutation in the tRNA(Leu, UUR) gene. This mutation associates in most individuals with maternally inherited diabetes and deafness (MIDD) whereas in some other carriers the MELAS syndrome or a progressive kidney failure is seen. Possible pathogenic mechanisms are discussed especially the question why particular mutations in mtDNA associate with distinct clinical entities. Mutations in mtDNA can affect the ATP production, thereby leading to particular clinical phenotypes such as muscle weakness. On the other hand mtDNA mutations may also alter the intracellular concentration of mitochondrial metabolites which can act as signalling molecules, such as Ca or glutamate. This situation may contribute to the development of particular phenotypes that are associated with distinct mtDNA mutations.     

Mitochondrial gene mutations in gestational diabetes mellitus

Mitochondrial DNA mutations have been implicated in many diseases including diabetes mellitus. Although gestational diabetes mellitus (GDM) has been suggested to have genetic determinant and to be etiologically indistinct with non-insulin-dependent diabetes mellitus (NIDDM), its association with mitochondrial gene mutations is still unknown. In this study, 137 patients with GDM and 292 non-diabetic pregnant controls were examined for mitochondrial DNA mutations from the nucleotide 3130-4260 encompassing tRNA-Leu gene and adjacent NADH dehydrogenase 1 gene by polymerase chain reaction, single-stranded conformation polymorphism, restriction fragment length polymorphism and DNA sequencing. One heteroplasmic mutation at the position of 3398 (T-C), which changed a highly conserved methionine to threonine in NADH dehydrogenase subunit 1, was identified in 2.9% GDM patients but not in the controls, indicating its association with GDM (P = 0.01). Two novel mutations, a heteroplasmic C3254A and a homoplasmic A3399T, were also found in GDM subjects, the functional meaning of which merits further investigation. G3316A and T3394C mutations implicated in NIDDM, were seen at higher frequencies in patients with GDM than the controls. Our results suggest that mitochondrial DNA mutations may contribute to the development of GDM in some patients.     

A transgenic model to study the pathogenesis of somatic mtDNA mutation accumulation in β-cells

Low levels of somatic mutations accumulate in mitochondrial DNA (mtDNA) as we age; however, the pathogenic nature of these mutations is unknown. In contrast, mutational loads of >30% of mtDNA are associated with electron transport chain defects that result in mitochondrial diseases such as mitochondrial encephalopathy lactic acidosis and stroke-like episodes. Pancreatic β-cells may be extremely sensitive to the accumulation of mtDNA mutations, as insulin secretion requires the mitochondrial oxidation of glucose to CO₂. Type 2 diabetes arises when β-cells fail to compensate for the increased demand for insulin, and many type 2 diabetics progress to insulin dependence because of a loss of β-cell function or β-cell death. This loss of β-cell function/β-cell death has been attributed to the toxic effects of elevated levels of lipids and glucose resulting in the enhanced production of free radicals in β-cells. mtDNA, localized in close proximity to one of the major cellular sites of free radical production, comprises more than 95% coding sequences such that mutations result in changes in the coding sequence. It has long been known that mtDNA mutations accumulate with age; however, only recently have studies examined the influence of somatic mtDNA mutation accumulation on disease pathogenesis. This article will focus on the effects of low-level somatic mtDNA mutation accumulation on ageing, cardiovascular disease and diabetes.     

The heteroplasmic m.14709T>C mutation in the tRNAGlu gene in two Tunisian families with mitochondrial diabetes

Diabetes mellitus (DM) is a heterogeneous disorder characterized by the presence of chronic hyperglycemia. Genetic factors play an important role in the development of this disorder, and several studies reported mutations in nuclear genes implicated in the insulin function. Besides, DM can be maternally transmitted in some families, possibly due to the maternal mitochondrial inheritance. In fact, mitochondrial genes may be plausible causative agents for diabetes, since mitochondrial oxidative phosphorylation plays an important role in glucose-stimulated insulin secretion from beta cells.Materials and MethodsIn this report, we screened two Tunisian families with mitochondrial diabetes for the m.3243A>G and the m.14709T>C mutations, respectively, in the tRNA^Leu(UUR) and the tRNA^Glu genes.ResultsThe polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) and the sequence-specific primers by polymerase chain reaction (SSP-PCR) analysis in the leucocytes and the buccal mucosa in the members of the two families showed the absence of the m.3243A>G mutation and the presence of the heteroplasmic m.14709T>C mutation in the tRNA^Glu gene in the two tested tissues.ConclusionsWe conclude that the m.14709T>C mutation in the tRNA^Glu gene could be a cause of mitochondrial diabetes in Tunisian affected families. In addition, the heteroplasmic loads correlated with the severity and the onset of mitochondrial diabetes in one family but not in the other, suggesting the presence of environmental factors or nuclear modifier genes.     

Mitochondrial complex I activity is significantly decreased in a patient with maternally inherited type 2 diabetes mellitus and hypertrophic cardiomyopathy associated with mitochondrial DNA C3310T mutation: a cybrid study

Mitochondrial respiratory function in a patient with maternally inherited type 2 diabetes mellitus and hypertrophic cardiomyopathy associated with heteroplasmic mitochondrial DNA (mtDNA) C3310T mutation, which replaces the second amino acid of NADH dehydrogenase 1 (ND1) from a hydrophobic Proline to a hydrophilic Serine, was investigated. Mitochondrial respiratory function solely due to mtDNA C3310T mutation was investigated in cybrid system by the fusion of mtDNA-deleted (rho(0)) HeLa cells and exogenous mtDNA either from the proband or from controls. Total oxygen consumption of the proband cybrid cells was significantly decreased compared with those of controls (2.468+/-0.475 versus 2.871+/-0.484 micromol/h/10(7) cells, p=0.0392). Mitochondrial respiratory chain complex I activity of the proband cybrid cells was also significantly decreased compared with those of controls (0.191+/-0.080 versus 0.288+/-0.113 micromol/h/mg protein, p=0.0223). Furthermore, ATP content in the proband cybrid cells was also significantly decreased compared with those in controls (1.119+/-0.344 versus 1.419+/-0.378 pmol/10(5) cells, p=0.044). The present study indicates that mtDNA C3310T mutation may be a pathogenic mutation of maternally inherited type 2 diabetes mellitus and hypertrophic cardiomyopathy in the proband and the family.     

Biogenesis of mitochondria and genetics of mitochondrial defects

Mitochondria are formed by the concerted action of two genetic systems: the nucleocytoplasmic system and the intrinsic mitochondrial system. The genetic contribution of the mitochondria is modest because the genetic potential of mtDNA of mammals is restricted to the equivalent of about 16 000 base pairs. For various animals and man the complete base sequence of mtDNA is known and all possible polypeptide genes have now been assigned to subunits of the respiratory enzymes. The mtDNA sequences are not present on the nuclear genome.From a genetic point of view it is important that the inheritance of mtDNA is strictly maternal. Mutations of mtDNA primarily lead to impairments of energy metabolism. In view of the indispensability of oxidative phosphorylation for obligatory aerobic organisms, such mutations should be lethal. However, there are various inborn errors of metabolism with tissue-specific manifestations, which are maternally inherited. The question discussed is whether these diseases can be explained on the basis of mutations of mitochondrial gene products. Tissue specificity poses a special problem, since it is not very attractive to assume that there is a heterogenous population of mtDNA molecules in the fertilized egg. Therefore, one should rather think in terms of a double mutational event, one tissue-specific cytoplasmic and the other general mitochondrial. These mutations only give rise to metabolic disturbances if they are expressed together in the same cell.     

Irritable Bowel Syndrome May Be Associated with Maternal Inheritance and Mitochondrial DNA Control Region Sequence Variants

Background and Aim

Mitochondrial dysfunction has been implicated in various functional disorders that are co-morbid to irritable bowel syndrome (IBS) such as migraine, depression and chronic fatigue syndrome. The aim of the current case–control pilot study was to determine if functional symptoms in IBS show a maternal inheritance bias, and if the degree of this maternal inheritance is related to mitochondrial DNA (mtDNA) polymorphisms.

Methods

Pedigrees were obtained from 308 adult IBS patients, 102 healthy controls, and 36 controls with inflammatory bowel disease (IBD), all from Caucasian heritage, to determine probable maternal inheritance. Two mtDNA polymorphisms (16519T and 3010A), which have previously been implicated in other functional disorders, were assayed in mtDNA haplogroup H IBS subjects and compared to genetic data from 344 published haplogroup H controls.

Results

Probable maternal inheritance was found in 17.5 % IBS, 2 % healthy controls and 0 % IBD controls (p < .0001). No difference was found between IBS and control for 3010A, and a trend was found for 16519T (p = 0.05). IBS with maternal inheritance were significantly more likely to have the 16519T than controls (OR 5.8; 95 % CI 1.5–23.1) or IBS without maternal inheritance (OR 5.2; 95 % CI 1.2–22.6).

Conclusions

This small pilot study shows that a significant minority (1/6) of IBS patients have pedigrees suggestive of maternal inheritance. The mtDNA polymorphism 16519T, which has been previously implicated in other functional disorders, is also associated with IBS patients who display maternal inheritance. These findings suggest that mtDNA-related mitochondrial dysfunction may constitute a sub-group within IBS. Future replication studies in larger samples are needed.     

A homoplasmic mitochondrial transfer ribonucleic acid mutation as a cause of maternally inherited hypertrophic cardiomyopathy

OBJECTIVES: The purpose of this study was to understand the clinical and molecular features of familial hypertrophic cardiomyopathy (HCM) in which a mitochondrial abnormality was strongly suspected. BACKGROUND: Defects of the mitochondrial genome are responsible for a heterogeneous group of clinical disorders, including cardiomyopathy. The majority of pathogenic mutations are heteroplasmic, with mutated and wild-type mitochondrial deoxyribonucleic acid (mtDNA) coexisting within the same cell. Homoplasmic mutations (present in every copy of the genome within the cell) present a difficult challenge in terms of diagnosis and assigning pathogenicity, as human mtDNA is highly polymorphic. METHODS: A detailed clinical, histochemical, biochemical, and molecular genetic analysis was performed on two families with HCM to investigate the underlying mitochondrial defect. RESULTS: Cardiac tissue from an affected child in the presenting family exhibited severe deficiencies of mitochondrial respiratory chain enzymes, whereas histochemical and biochemical studies of the skeletal muscle were normal. Mitochondrial DNA sequencing revealed an A4300G transition in the mitochondrial transfer ribonucleic acid (tRNA)(Ile) gene, which was shown to be homoplasmic by polymerase chain reaction/restriction fragment length polymorphism analysis in all samples from affected individuals and other maternal relatives. In a second family, previously reported as heteroplasmic for this base substitution, the mutation has subsequently been shown to be homoplasmic. The pathogenic role for this mutation was confirmed by high-resolution Northern blot analysis of heart tissue from both families, revealing very low steady-state levels of the mature mitochondrial tRNA(Ile). CONCLUSIONS: This report documents, for the first time, that a homoplasmic mitochondrial tRNA mutation may cause maternally inherited HCM. It highlights the significant contribution that homoplasmic mitochondrial tRNA substitutions may play in the development of cardiac disease. A restriction of the biochemical defect to the affected tissue has important implications for the screening of patients with cardiomyopathy for mitochondrial disease.     

Somatic alterations in mitochondrial DNA and mitochondrial dysfunction in gastric cancer progression

Energy metabolism reprogramming was recently identified as one of the cancer hallmarks.One of the underlying mechanisms of energy metabolism reprogramming is mitochondrial dysfunction caused by mutations in nuclear genes or mitochondrial DNA(mtDNA).In the past decades,several types of somatic mtDNA alterations have been identified in gastric cancer.However,the role of these mtDNA alterations in gastric cancer progression remains unclear.In this review,we summarize recently identified somatic mtDNA alterations in gastric cancers as well as the relationship between these alterations and the clinicopathological features of gastric cancer.The causative factors and potential roles of the somatic mtDNA alterations in cancer progression are also discussed.We suggest that point mutations and mtDNA copy number decreases are the two most common mtDNA alterations that result in mitochondrial dysfunction in gastric cancers.The two primary mutation types(transition mutations and mononucleotide or dinucleotide repeat instability)imply potential causative factors.Mitochondrial dysfunction-generated reactive oxygen species may be involved in the malignant changes of gastric cancer.The search for strategies to prevent mtDNA alterations and inhibit the mitochondrial retrograde signaling will benefit the development of novel treatments for gastric cancer and other malignancies.     

The G1888A variant in the mitochondrial 16S rRNA gene may be associated with Type 2 diabetes in Caucasian-Brazilian patients from southern Brazil.

AIM: To compare the frequencies of the G1888A variant in the mitochondrial 16S rRNA gene between patients with Type 2 diabetes and non-diabetic control subjects from southern Brazil. METHODS: We analysed 520 Type 2 diabetic patients (389 Caucasian- and 131 African-Brazilians) and 530 control subjects (400 Caucasian- and 130 African-Brazilians). DNA samples were amplified by polymerase chain reaction and digested with the RsaI enzyme. Variant frequency in patients and control subjects was compared using chi2 test, Fisher's exact test or odds ratio test. We also investigated the frequency of the G1888A variant in a subgroup of the patients with a maternal history of Type 2 diabetes plus two or more features of maternally inherited diabetes and deafness. RESULTS: The 1888A allele does not seem to be associated with Type 2 diabetes in African-Brazilians (frequency of 3.8% in patients and 0.8% in control subjects; PFisher=0.213). However, in Caucasian-Brazilians, the 1888A allele was significantly associated with diabetes (12.3% in patients vs. 3.5% in control subjects; OR=3.881; 95% CI 2.106-7.164; P<0.001) and also with higher levels of insulin resistance. The majority of the patients carrying the 1888A allele did not have clinical features of maternally inherited diabetes and deafness. CONCLUSION: The present study indicates the association of the mitochondrial G1888A variant with Type 2 diabetes and insulin resistance in Caucasian-Brazilian patients from southern Brazil. However, further studies are required to confirm its effects on mitochondrial function and the role of these effects on the pathogenesis of Type 2 diabetes.