Molecular genetics of a patient with Mohr-Tranebjaerg Syndrome due to a new mutation in the DDP1 gene.

The deafness-dystonia syndrome (DDS) or Mohr-Tranebjaerg syndrome (MTS, MIM 304700) is a rare X-linked recessive neurological disorder resulting from loss-of-function mutations in the nuclear DDP1/TIMM8A gene, involved in the transport and sorting of proteins to the mitochondrial inner membrane. A Mohr-Tranebjaerg patient and his mother were subjected to clinical and molecular studies. Screening of mutations were performed in TIMM8A, TIMM13, and other mitochondrial protein transport genes by conformation sensitive gel electrophoresis (CSGE), followed by direct DNA sequencing of tissue samples from the patient. Mitochondrial DNA of the patient was also sequenced at the genes for COX subunits and some mitochondrial tRNAs. Respiratory chain activities in a muscle biopsy and cultured fibroblasts from the patient were assessed using biochemical methods. mRNA expression of TIMM8A and TIMM13 was determined by RT-PCR in cultured fibroblasts. We identified a new case of Mohr-Tranebjaerg syndrome and report the characteristics of a new pathogenic de novo mutation (c.112C>T, pGln38X) in the TIMM8A gene. Biochemical measures of respiratory chain complex activities in muscle biopsy and fibroblasts did not show a major deficiency or alteration. mRNA expression studies demonstrated increased TIMM8A mRNA levels in cultured fibroblasts from the patient. Phenotypic differences among published cases seem not to be related with the mutation location or type. Our results support the idea that dysfunctions of mitochondrial protein transport, in addition to OXPHOS deficiency, can be the basis of important mitochondrial pathologies.

Altered expression of master regulatory genes of adipogenesis in lipomas from patients bearing tRNA(Lys) point mutations in mitochondrial DNA.

The mechanisms underlying the appearance of lipomas in patients bearing mutations in the tRNA(Lys) gene of mitochondrial DNA are unknown. We investigated changes in gene expression patterns in lipomas from three patients bearing A8344G or G8363A tRNA(Lys) gene mutations. Uncoupling protein-1 mRNA was detected in the lipomas, in contrast with undetectable expression in normal adipose tissue. However, expression of other markers of brown fat, such as PGC-1alpha, was unaltered. PPARgamma and retinoblastoma gene expression was down regulated in the lipomas, but C/EBPalpha mRNA was not affected. The expression of Pref-1 was dramatically down regulated. Thus, lipomatosis due to tRNA(Lys) mutations is associated with a pattern of altered expression of master regulators of adipogenesis consistent with enhanced proliferation but maintenance of adipocyte features, and with a distorted pattern of brown versus white adipocyte differentiation.

[Deletions of the mitochondrial DNA associated to chronic progressive external ophthalmoplegia with ragged-red fibers in 2 Brazilian patients]. / Deleciones en el ADN mitocondrial asociadas a oftalmoplejía crónica extrínseca progresiva con fibras rojas rasgadas en 2 pacientes brasileños.

BACKGROUND AND OBJECTIVE: Our purpose was to report the neurological manifestations and molecular-genetic analysis of mitochondrial DNA associated with chronic progressive external ophthalmoplegia (CPEO) and raged red fibers (RRFs). PATIENTS AND METHOD: Two patients, a male and a female (32 and 28 year-old, respectively), were studied due to progressive palpebral ptosis associated with RRFs in muscle biopsy. Both patients were subjected to neurological, histochemical and enzymatic analysis of muscular biopsy, analysis of cerebro-spinal fluid, and molecular analysis of mitochondrial DNA. RESULTS: Symptoms started at ages 24 and 17 years. Initial symptoms were palpebral ptosis, progressive limitation of vertical and horizontal gaze, fatigue and exercise intolerance, and weakness of proximal muscles. Brain MRIs were normal in both patients. Both patients had deletions of muscle mitochondrial DNA with similar size (5,425 and 5,112 base pairs) and location. CONCLUSIONS: CPEO with RRFs is usually associated with huge deletions in mitochondrial DNA. Fatigue and proximal muscle weakness can be found during the follow-up.

De Toni-Debré-Fanconi syndrome due to a palindrome-flanked deletion in mitochondrial DNA.

We report the molecular findings in a child presenting with sideroblastic anemia and proximal tubulopathy. Analysis of mitochondrial DNA (mtDNA) from fibroblasts showed the presence of a 3.3-kb single deletion in 50% of the genomes. This mutation is, unlike other previously reported deletions in tubulopathy patients, not flanked by direct repeat sequences but by palindrome sequences at the deletion breakpoints, suggesting an unusual mechanism for production of deletion. These findings further expand our knowledge of the syndrome of anemia and tubulopathy due to single deletions of mtDNA.

Peripheral neuropathy with ataxia in childhood as a result of the G8363A mutation in mitochondrial DNA.

Peripheral neuropathy has been identified in children with mitochondrial encephalomyopathies but not as a main clinical landmark. Here we report the clinical, electrophysiologic, biochemical, and genetic findings in a family who harbors the G8363A mutation in the tRNALys gene of mitochondrial DNA. Affected individuals presented with peripheral neuropathy and ataxia as the main clinical sign. Additional involvement included muscle weakness and multiple lipomatosis. Other common clinical characteristics associated with the G8363A mutation, such as cardiomyopathy and myoclonus epilepsy, were not observed. These findings suggest that a mitochondrial disease should be considered in the differential diagnosis of children with heredoataxic syndrome and peripheral neuropathy of unknown origin.

Bilateral striatal necrosis associated with a novel mutation in the mitochondrial ND6 gene.

We report the molecular findings in two independent patients presenting with progressive generalized dystonia and bilateral striatal necrosis in whom we have identified a mutation (T14487C) in the mitochondrial ND6 gene. The mutation is heteroplasmic in all samples analyzed, and it fulfills all accepted criteria of pathogenicity. Transmitochondrial cell lines harboring 100% mutant mitochondrial DNA showed a marked decrease in the activity of complex I of the respiratory chain supporting the pathogenic role of T14487C.

Liver failure caused by herpes simplex virus thymidine kinase plus ganciclovir therapy is associated with mitochondrial dysfunction and mitochondrial DNA depletion.

Herpes simplex virus thymidine kinase (HSV-tk) converts ganciclovir (GCV) into an active compound, which can be incorporated into DNA molecules and terminate DNA synthesis. Gene transfer of HSV-tk followed by GCV administration has been used with success to treat experimental cancer and this strategy has entered into clinical trials. Although it is thought that the cytotoxic effect occurs mainly in tumoral dividing cells, where mitotic activity favors integration of the genotoxic compound into nuclear DNA, there are concerns of potential damage to normal nondividing cells. In the present work we have explored the mechanisms of HSV-tk/GCV toxicity and in particular whether this therapy may cause lesions of mitochondrial DNA (mtDNA) and mitochondrial dysfunction. We found that the administration of GCV to rats injected with adenovirus encoding HSV-tk induced hepatocellular damage characterized by the presence of apoptotic bodies, ballooning of hepatocytes, and severe hepatic steatosis with mitochondria enlargement and cristae dissolution at the ultrastructural level. Remarkably, Southern blot analysis showed substantial reduction in the amount of mtDNA in the liver. Using radiolabeled GCV we could demonstrate incorporation of this compound into both nuclear and mtDNA in HSV-tk-transduced rat hepatocytic cell line MCA-RH7777 and subsequent alteration of mitochondrial function. Our observations confirm that GCV can damage both nuclear and mtDNA in cells transduced with HSV-tk and that this effect could be responsible for severe mitochondrial dysfunction and toxicity in normal nondividing cells. These data are relevant for the design of clinical trials using adenoviral vectors encoding HSV-tk.