Genetic counseling and prenatal diagnosis for the mitochondrial DNA mutations at nucleotide 8993.

Mitochondrial genetics is complicated by heteroplasmy, or mutant load, which may be from 1%-99%, and thus may produce a gene dosage-type effect. Limited data are available for genotype/phenotype correlations in disorders caused by mtDNA mutations; therefore, prenatal diagnosis for mtDNA mutations has been hindered by an inability to predict accurately the clinical severity expected from a mutant load measured in fetal tissue. After reviewing 44 published and 12 unpublished pedigrees, we considered the possibility of prenatal diagnosis for two common mtDNA mutations at nucleotide 8993.

A series of mutations in the dihydropteridine reductase gene resulting in either abnormal RNA splicing or DHPR protein defects. Mutations in brief no. 244. Online.

Five novel mutations are described which result in the rare hyperphenylalaninemia DHPR-deficiency. Three of these are located at different intron/exon boundaries within the DHPR gene, and disrupt the maturation of the DHPR transcript such that little full-length mRNA can be detected by RT-PCR. Each mutation alters a conserved nucleotide within the splice site consensus sequence, and results in the skipping of an exon and, in one case, the activation of an inappropriate splicing signal.

Defective copper-induced trafficking and localization of the Menkes protein in patients with mild and copper-treated classical Menkes disease.

Menkes disease is an X-linked disorder of copper metabolism. An overall copper deficiency reduces the activity of copper-dependent enzymes accounting for the clinical presentation of affected individuals. The Menkes gene product (MNK) is a P-type ATPase and is considered to be the main copper efflux protein in most cells.

Trisomy 20p resulting from inverted duplication and neocentromere formation.

Normal human centromeres contain large tandem arrays of alpha-satellite DNA of varying composition and complexity. However, a new class of mitotically stable marker chromosomes which contain neocentromeres formed from genomic regions previously devoid of centromere activity was described recently. These neocentromeres are fully functional yet lack the repeat sequences traditionally associated with normal centromere function.

Pitfalls in the use of 2-octynoic acid as an in vivo model of medium-chain acyl-coenzyme A dehydrogenase deficiency: ketone turnover and metabolite studies in the rat.

2-Octynoic acid was administered by intraperitoneal injection to fasted Sprague-Dawley rats in an attempt to simulate medium-chain acyl-coenzyme A dehydrogenase (MCAD) deficiency. The resultant urine organic acid profile showed a mild dicarboxylic aciduria but lacked the glycine conjugates characteristic of MCAD deficiency. Further studies with infused 13C(4)-3-hydroxybutyrate and 13C(2)-acetoacetate demonstrated reduced ketone production in treated rats compared with control animals.

Defective chromosome segregation, microtubule bundling and nuclear bridging in inner centromere protein gene (Incenp)-disrupted mice.

INCENP is a chromosomal passenger protein which relocates from the centromere to thel spindle midzone during the metaphase-anaphase transition, ultimately being discarded in the cell midbody at the completion of cytokinesis. Using homologous recombination, we have generated Incenp gene-targeted heterozygous mice that are phenotypically indistinguishable from their wild-type littermates. Intercrossing the hetero-zygotes results in no live-born homozygous Incenp -disrupted progeny, indicating an early lethality.

Connexin26 deafness in several interconnected families.

Mutations in the connexin26 gene are the basis of much autosomal recessive sensorineural deafness. There is a high frequency of mutant alleles, largely accounted for by one common mutation, 35delG. We have studied a group of families, who had been brought together through marriages between Deaf persons, in which there are more than 30 Deaf people in four generations.

Intracellular localization and loss of copper responsiveness of Mnk, the murine homologue of the Menkes protein, in cells from blotchy (Mo blo) and brindled (Mo br) mouse mutants.

Menkes disease is an X-linked copper deficiency disorder that results from mutations in the ATP7A ( MNK ) gene. A wide range of disease-causing mutations within ATP7A have been described, which lead to a diversity of phenotypes exhibited by Menkes patients. The mottled locus ( Mo, Atp7a, Mnk ) represents the murine homologue of the ATP7A gene, and the mottled mutants exhibit a diversity of phenotypes similar to that observed among Menkes patients.

Direct evidence that mitochondrial iron accumulation occurs in Friedreich ataxia.

Friedreich ataxia (FRDA) is due to mutations in the FRDA gene (FRDA). When the gene homologous to FRDA is knocked out in yeast, there is accumulation of iron in mitochondria and reduced respiratory function. So far, there is only indirect evidence to support the hypothesis that FRDA is due to accumulation of mitochondrial iron leading to increased production of free radicals.

Directly inherited partial trisomy of chromosome 6p identified in a father and daughter by chromosome microdissection.

Cytogenetic analysis of a 4 year old girl with developmental delay and dysmorphic features showed extra chromosomal material of unknown origin on 20p (46,XX,add(20)(p13)). Familial chromosome studies showed direct inheritance of add(20)(p13) from the father, who had a similar, albeit milder, phenotype. Fibroblast chromosome studies of the father showed no karyotype mosaicism.

Should we genetically test everyone for haemochromatosis?

The increasing availability of DNA-based diagnostic tests has raised issues about whether these should be applied to the population at large in order to identify, treat or prevent a range of diseases. DNA tests raise concerns in the community for several reasons. There is the possibility of stigmatisation and discrimination between those who test positive and those who don't.

Prenatal diagnosis and discrimination against the disabled.

Two versions of the argument that prenatal diagnosis discriminates against the disabled are distinguished and analysed. Both are shown to be inadequate, but some valid concerns about the social effects of prenatal diagnosis are highlighted.

Should doctors intentionally do less than the best?

The papers of Burley and Harris, and Draper and Chadwick, in this issue, raise a problem: what should doctors do when patients request an option which is not the best available? This commentary argues that doctors have a duty to offer that option which will result in the individual affected by that choice enjoying the highest level of wellbeing. Doctors can deviate from this duty and submaximise--bring about an outcome that is less than the best--only if there are good reasons to do so. The desire to have a child which is genetically related provides little, if any, reason to submaximise.

Should we clone human beings? Cloning as a source of tissue for transplantation.

The most publicly justifiable application of human cloning, if there is one at all, is to provide self-compatible cells or tissues for medical use, especially transplantation. Some have argued that this raises no new ethical issues above those raised by any form of embryo experimentation. I argue that this research is less morally problematic than other embryo research.

Respiratory chain complex I deficiency: an underdiagnosed energy generation disorder.

Objective: To define the spectrum of clinical and biochemical features in 51 children with isolated complex I deficiency.

Background: Mitochondrial respiratory chain defects are one of the most commonly diagnosed inborn errors of metabolism. Until recently there have been technical problems with the diagnosis of respiratory chain complex I defects, and there is a lack of information about this underreported cause of respiratory chain dysfunction.

Genotype and intellectual phenotype in untreated phenylketonuria patients.

Previous studies have shown that genotype correlates with biochemical phenotype in treated phenylketonuria. If there is a strong correlation between genotype and intellectual phenotype of untreated patients, it would be possible to determine which individuals would have normal intelligence without treatment. In this study, 42 families with untreated phenylketonuria were analyzed to examine whether there was an association between genotype and untreated intellectual phenotype.

The role of GMXCXXC metal binding sites in the copper-induced redistribution of the Menkes protein.

The Menkes protein (MNK or ATP7A) is a transmembrane, copper-transporting CPX-type ATPase, a subgroup of the extensive family of P-type ATPases. A striking feature of the protein is the presence of six metal binding sites (MBSs) in the N-terminal region with the highly conserved consensus sequence GMXCXXC. MNK is normally located in the trans-Golgi network (TGN) but has been shown to relocalize to the plasma membrane when cells are cultured in media containing high concentrations of copper.

Catenary cultures of embryonic gastrointestinal tract support organ morphogenesis, motility, neural crest cell migration, and cell differentiation.

The embryonic gastrointestinal tract develops from a simple tube into a coiled, flexed, and regionalized structure. The changes in gut morphology coincide with the differentiation of multiple cell types in concentric layers, and include colonization by migratory neuron precursors, and the development of gastrointestinal motility. We describe a reliable method for growing embryonic mouse intestine in vitro by the attachment of segments of intestinal tract by their cut ends, with the intervening region suspended in the culture medium.

Duplex PCR for Autosomal Dominant Spinocerebellar Ataxia Testing: A Nonradioactive Rapid Screening Method.

Background: Current DNA diagnostic testing for spinocerebellar ataxias (SCAs) 1, 2, 3, and 6 involves four separate tests based on radioactively-labeled poly merase chain reaction (PCR). Although this approach allows accurate allele sizing, it also is time-consuming and requires manipulation with hazardous radioactive isotopes. Methods and Results: A rapid screening approach was developed based on a nonradioactive and duplexed PCR assay coupling SCA 1 and SCA 3 together and SCA 2 and SCA 6 together.

A False-Positive Diagnosis for the Common MELAS (A3243G) Mutation Caused by a Novel Variant (A3426G) in the ND1 Gene of Mitochondria DNA.

Background: Several mutations in mitochondrial DNA (mtDNA) are associated with the syndrome of mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS). The "common" MELAS mutation, A3243G in the tRNA leucine (UUR) gene, affects approximately 80% of cases and is associated with respiratory chain complex I deficiency. Methods and Results: The A3243G mutation creates an ApaI restriction endonuclease site and can be detected by polymerase chain reaction (PCR) amplification of a region of mtDNA containing nt 3243, followed by ApaI digestion and electrophoretic analysis of the resulting fragments.