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A 6-week-old child presented with hypotonia, myopathy, and a rapidly worsening dilated cardiomyopathy with severe atrial and ventricular arrhythmias and pulmonary hypertension, which proved fatal at age 3 months. Biochemical analysis showed a combined deficiency of the enzymatic activities of complexes I and IV and molecular studies identified a T14709C mutation in the mitochondrial tRNA glutamic acid gene. A review of symptomatology in patients with this mutation shows that it mainly presents in childhood or young adults with mild myopathy and diabetes mellitus. Infants with a high, nearly homoplasmic mutant load can present with more severe symptoms including cardiomyopathy. Families with this mitochondrial DNA mutation should be aware that increased mutant load in a subsequent generation may result in severe and often fatal cardiac symptoms.

A 6-year-old boy had progressive muscle weakness since age 4 and emotional problems diagnosed as Asperger syndrome. His mother and two older siblings are in good health and there is no family history of neuromuscular disorders. Muscle biopsy showed ragged-red and cytochrome coxidase (COX)-negative fibers. Respiratory chain activities were reduced for all enzymes containing mtDNA-encoded subunits, especially COX. Sequence analysis of the 22 tRNA genes revealed a novel G10406A base substitution, which was heteroplasmic in multiple tissues of the patient by RFLP analysis (muscle, 96%; urinary sediment, 94%; cheek mucosa, 36%; blood, 29%). The mutation was not detected in any accessible tissues from his mother or siblings. It appears that this mutation arose de novo in the proband, probably early in embryogenesis.

A 14-year-old boy had exercise intolerance, weakness, ataxia, and lactic acidosis. Because his muscle biopsy showed a mosaic pattern of fibers staining intensely with the succinate dehydrogenase reaction but not at all with the cytochrome c oxidase reaction, we sequenced his mitochondrial DNA and found a novel mutation (C14680A) in the gene for tRNAGlu. The mutation was present in accessible tissues from the asymptomatic mother but not from a brother with Asperger syndrome. These data expand the clinical heterogeneity of mutations in this mitochondrial gene.

Objective: To document novel homozygous mutations in the gene for deoxyguanosine kinase (DGK) in 3 children with mitochondrial DNA depletion., Design: Clinical features included liver failure, hypotonia, and nystagmus in 2 siblings, and liver cirrhosis, optic dysplasia, nystagmus, and microcephaly in the third patient. We sequenced the whole coding region of the DGK gene., Results: We identified 2 novel homozygous mutations, G352A and C269T, that lead to truncated proteins., Conclusion: These data confirm that DGK mutations typically affect the liver and brain.

The majority of patients with MELAS (mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes) have the A3243G point mutation. The much rarer T3271C mutation has been reported predominantly in Japanese subjects. Our objective was to better define the clinical phenotype and mutation load in patients with MELAS and the T3271C mutation in mitochondrial DNA. We present clinical and molecular genetic data in two pedigrees with the T3271C mutation. The age at onset was 8 years in one proband and 14 years in the other. Both patients had migrainelike headache, seizures, and strokelike episodes. Mutation loads were quantified in multiple tissues from the patients and from family members by polymerase chain reaction-restriction fragment length polymorphism analysis. The symptoms in both probands were typical of MELAS, and, contrary to previous reports, onset was early. Hearing loss was less common than in typical MELAS, and ragged red fibers were absent. The proportion of mutant genomes was consistently and markedly greater in DNA from urinary sediment than from blood. In the mother of one proband, mutant genomes were detected only in DNA from hair follicles and cheek mucosa The phenotype of patients with the T3271C mutation might not be as distinct as that of the A3243G mutation, as previously described. Our data also suggest that urine is a better source of DNA than blood for diagnosis and that multiple tissues should be studied in maternal relatives, especially when the mutation cannot be detected in blood.

Testing for common mutations in mitochondrial DNA (mtDNA), including the A3243G MELAS (mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes) mutation, is routinely done in DNA isolated from blood. Since the blood level of the A3243G mutation may be low in probands and even lower in asymptomatic or oligosymptomatic maternal relatives, we assessed the proportion of A3243G mutant genomes in other accessible tissues. We studied five tissues (leukocytes, skin fibroblasts, hair roots, urinary sediment, and cheek mucosa) in 61 individuals from 22 families harboring the A3243G mutation. Although mutational loads varied widely among these tissues, as a rule DNA from urinary sediment had the highest and blood the lowest proportion of mutant genomes; individual hair roots differed markedly from one another; fibroblasts and cheek mucosa tended to have higher mutation loads than blood but lower than urinary sediment. In all individuals in whom the mutation was detectable in blood, it was also detected in other tissues. Conversely, in some individuals the mutation was undetectable in blood but clearly present in other tissues. We conclude that urinary sediment and cheek mucosa are tissues of choice for the diagnosis of mtDNA mutations, as they are easy to obtain and the mutation load is almost always greater than in blood.

Background: The mendelian forms of progressive external ophthalmoplegia (PEO) associated with multiple mitochondrial DNA deletions are clinically heterogeneous disorders transmitted as dominant or recessive traits. Autosomal dominant PEO is caused by mutations in at least 3 genes: adenine nucleotide translocator-1 (ANT1), encoding the muscle-specific adenine nucleotide translocator; chromosome 10 open reading frame 2 (C10orf2), encoding Twinkle helicase; and polymerase gamma (POLG), encoding the alpha subunit of polymerase gamma. Mutations in POLG can also cause autosomal recessive PEO, which is often associated with multisystemic disorders., Objective and Methods: To further investigate the frequency and genotype-phenotype correlations of mutations in the POLG gene, we used single-stranded conformational polymorphism analysis and direct sequencing to screen 30 patients with familial or sporadic PEO and multiple mitochondrial DNA deletions in muscle but without mutations in ANT1 and C10orf2., Results: Four unrelated patients had novel POLG mutations. A woman with PEO and mental retardation had a heterozygous Gly1076Val mutation. Two patients, one with PEO, exercise intolerance, and gastrointestinal dysmotility and the other with PEO, neuropathy, deafness, and hypogonadism, both had a Pro587Leu change. The fourth patient, who was compound heterozygous for Ala889Thr and Arg579Trp mutations, had PEO, gastrointestinal dysmotility, and neuropathy. These mutations were not detected in 120 healthy control alleles., Conclusions: Our results demonstrate that POLG mutations account for a substantial proportion of patients (13%) with PEO and multiple mitochondrial DNA deletions and cause both clinically and genetically heterogeneous disorders.

Cactaceae are a diverse group of plants with a wide variety of morphologies and reproductive strategies. Many species have segmented stems in which terminal cladodes may be separated from main stem cladodes with varying amounts of resistance. Previous results demonstrated that lignified xylem cells in tensile portions of stem joints provide the main resistance to separation of cladodes within cactus plants. The purpose of the present study was to determine if stem joints of Opuntia laevis would produce additional lignified xylem cells in response to additional externally applied stresses. Normal average stress levels, which accompany the addition of a new cladode, were applied to 12 plants. In contrast, double the average stress levels were applied to 13 other plants. After exposure to the two stress regimens for 6 months, the amount and location of lignified xylem cells in joint segments were similar for both stress treatments. So, although the results support the hypothesis that lignified xylem cells act as the main resistance to stress at joints of cladodes, doubling the normal amount of applied stress was insufficient to alter the amount or location of lignified xylem cells in stem joints. These results indicate that normal amounts of lignified xylem cells can resist up to two times the normal amount of stress for 6 months without producing additional lignified xylem cells.

Terminal meristems are responsible for all primary growth of roots. It has been asserted that all cells of root meristems are actively dividing and that the stem cell (proliferative) population expands exponentially. Lengths of cells in roots just proximal to the root cap/root initial boundary were used to determine the numbers of cortex and stele cells in the meristem. Meristem cells were defined as cells that did not have significantly different cell lengths from initial cells at the boundary. Data show that, for five of the six species (Allium cepa, Pisum sativum, Pyrus communis, Triticum aestivum, Vicia faba, and Zea mays) tested, only the first 15 stele and the first 10-35 cortex cells in median longitudinal sections would be in the meristem. For T. aestivum, no discrete meristem was found because all cells proximal to initial cells were longer than initial cells. In addition to this subject area, distributions of lengths of cells in the root meristem using this definition, for the six species were compared with a theoretical cell-age distribution for exponentially dividing cells, to determine if distributions of cell lengths were similar to a theoretical distribution of exponentially dividing cells. For all species tested, distributions of cell lengths were not similar to a theoretical cell-age distribution. From the data of this study with six plant species, we conclude that either contiguous proliferative cell populations of root meristems are very small or the proliferative cell population is not continuous. In addition, such populations do not resemble a theoretical exponential cell-age distribution. Moreover, it seems that the proliferative capacities of cells within terminal root segments differ markedly among species and are not easily characterized.