Cellular UDP-glucose deficiency caused by a single point mutation in the UDP-glucose pyrophosphorylase gene

We previously isolated a mutant cell that is the only mammalian cell reported to have a persistently low level of UDP-glucose. In this work we obtained a spontaneous revertant whose UDP-glucose level lies between those found in the wild type and the mutant cell. The activity of UDP-glucose pyrophosphorylase (UDPG:PP), the enzyme that catalyzes the formation of UDP-glucose, was in the mutant 4% and in the revertant 56% of the activity found in the wild type cell. Sequence analysis of UDPG: PP cDNAs from the mutant cell showed one missense mutation, which changes amino acid residue 115 from glycine to aspartic acid. The substituted glycine is located within the largest stretch of strictly conserved residues among eukaryotic UDPG:PPs. The analysis of the cDNAs from the revertant cell indicated the presence of an equimolar mixture of the wild type and the mutated mRNAs, suggesting that the mutation has reverted in only one of the alleles. In summary, we demonstrate that the G115D substitution in the Chinese hamster UDPG:PP dramatically impairs its enzymatic activity, thereby causing cellular UDP-glucose deficiency.     

Progressive juvenile-onset punctate cataracts caused by mutation of the gammaD-crystallin gene

Cataracts are a significant public health problem. Here, we describe the genetic alteration responsible for a progressive form of cataract, segregating as an autosomal dominant trait in a three-generation pedigree. Unlike most autosomal dominant cataracts, these are not clinically apparent at birth but are initially observed in the first year or two of life. The opacification evolves relatively slowly, generally necessitating removal of the lens in childhood or early adolescence. A genome-wide search in our kindred revealed linkage at 2q33-35 where the gamma-crystallin gene cluster resides. A single base alteration resulting in an Arg- 14 --> Cys (R14C) substitution in gammaD-crystallin was subsequently identified. Protein modeling suggests that the effect of this mutation is a subtle one, affecting the surface properties of the crystallin molecule rather than its tertiary structure, consistent with the fact that the patients' lenses are normal at birth. This is the first gene defect shown to be responsible for a noncongenital progressive cataract, and studying the defective protein should teach us more about the mechanisms underlying cataract formation.     

Extensive phenotypic analysis of a family with growth hormone (GH) deficiency caused by a mutation in the GH-releasing hormone receptor gene

We studied the effects of aerosolized as well as intravenous infusion of acetylcholine on bronchial blood flow in six anesthetized sheep. Intravenous infusion of acetylcholine, at a dose of 2 microg/kg, increased bronchial blood flow from 45 +/- 15 (SE) to 74 +/- 30 ml/min, and vascular conductance increased by 76 +/- 22%. In contrast, aerosolized acetylcholine at doses of 2 and 20 microg/kg decreased bronchial vascular conductance by approximately 10%. At an aerosolized dose of 200 microg/kg, the bronchial vascular conductance increased by approximately 15%, and there was no further increase in conductance when the aerosolized dose was increased to 2,000 microg/kg. Pretreatment of animals with a nitric oxide synthase inhibitor, Nomega-nitro-L-arginine methyl ester hydrochloride, partially blocked the vasodilatory effects of intravenous acetylcholine and completely blocked the vasodilatory effects of high-dose aerosolized acetylcholine. These data suggest that aerosolized acetylcholine does not readily penetrate the vascular wall of bronchial circulatory system and, therefore, has minimal vasodilatory effects on the bronchial vasculature.     

Mechanisms of expression of pyruvate dehydrogenase deficiency caused by an E1alpha subunit mutation

OBJECTIVE: To characterize the biochemical mechanisms of expression of the pyruvate dehydrogenase (PDH) E1alpha subunit exon 10 R302C missense mutation. BACKGROUND: Mutations in the X-linked E1alpha subunit gene are responsible for most cases of PDH deficiency, an important cause of neurodevelopmental defects and neurodegeneration with primary lactic acidemia. Although the disease shows extreme allelic heterogeneity, the R302C mutation has been defined in several unrelated cases. METHODS: Cell lines expressing selectively either the mutant or wild-type E1alpha alleles against identical genetic backgrounds were generated from the fibroblasts of a female heterozygous for the R302C mutation. Enzyme activity, mRNA, polypeptide expression, and turnover were studied in each. RESULTS: The residual PDH activity was below measurable levels in the cell line (B5) expressing only the mutant allele and normal in the wild-type polypeptide expressing (A10) cell line, confirming that the R302C mutation alone is sufficient to cause a severe PDH deficiency. The mutant polypeptide was less stable than the wild-type polypeptide, but the steady-state level of the mutant E1alpha protein was reduced only two- to threefold. CONCLUSIONS: The primary mechanism of expression of the R302C mutation must be limitation of catalytic efficiency. We speculate that catalysis may be inhibited in the mutant polypeptide because conformational changes are induced near serine 300, a residue that is particularly important as a regulatory phosphorylation site in the wild-type polypeptide.     

A novel nonsense mutation associated with an exon skipping in a patient with hereditary protein S deficiency type I

The genetic defect in a patient with hereditary type I protein S (PS) deficiency was investigated. All the exons and intron-exon junctions of the patient's PS gene were amplified by PCR and subjected to heteroduplex screening. Only the PCR product of exon 4 revealed heteroduplex bands. A novel nonsense mutation, Ser62 (TCA) to Stop (TGA) was found in exon 4. RT-PCR detected the aberrant mRNA in the patient's platelets, which was markedly reduced in amount and lacked the region of exon 4, suggesting that the nonsense mutation affected the mutated mRNA metabolism and induced exon skipping. The skipping of exon 4 causes an in-frame deletion of 29 amino acids which just construct the thrombin-sensitive region of the PS molecule. The loss of such an important domain as well as the quantitative decrease in the mutated mRNA appear to be responsible for the type I PS deficiency in this patient.     

The inlA gene of Listeria monocytogenes LO28 harbors a nonsense mutation resulting in release of internalin

Internalin is a surface protein that mediates entry of Listeria monocytogenes EGD into epithelial cells expressing the cell adhesion molecule human E-cadherin or its chicken homolog, L-CAM, which act as receptors for internalin. After observing that entry of L. monocytogenes LO28 into S180 fibroblasts, in contrast to that of EGD, did not increase after transfection with L-CAM, we examined both the expression and the structure of internalin in strain LO28. We discovered a nonsense mutation in inlA which results in a truncated protein released in the culture medium. Mutations leading to release of internalin were also detected in clinical and food isolates. These results question the role of internalin as a virulence factor in murine listeriosis.     

A homozygous nonsense mutation of the human growth hormone receptor gene in a Sardinian boy with Laron-type dwarfism

Patients with idiopathic intracranial hypertension may occasionally present with coexisting lower motor neuron facial weakness. This study reviews a 6-year experience at Mayo Clinic. The aim of this study was to determine the possible association of idiopathic intracranial hypertension and facial paresis. Two cases were identified. Both fulfilled the modified Dandy's diagnostic criteria for idiopathic intracranial hypertension. Treatment consisted of steroids in one, and emergent optic nerve sheath fenestration in the other. The cranial nerve palsies resolved in both cases.     

Mutation analysis of the 6-pyruvoyl-tetrahydropterin synthase gene in Chinese hyperphenylalaninemia caused by tetrahydrobiopterin synthesis deficiency

BACKGROUND: The systemic manifestations of acute pancreatitis are responsible for the majority of pancreatitis-associated morbidity and mortality and are now believed to be due to the actions of specific inflammatory cytokines. This report summarizes what is known about the role of cytokines in the pathogenesis of acute pancreatitis. METHODS: Comprehensive literature review of experimental pancreatitis as well as all reports of cytokine involvement during clinical pancreatitis. RESULTS: Several cytokines and other noncytokine inflammatory mediators are produced rapidly during pancreatitis. These mediators arise in many tissues in a predictable fashion independent of the animal model used or the underlying etiology in human disease. Preventing the activities of these mediators has a profound beneficial effect in experimental animals. CONCLUSIONS: A few recently described inflammatory mediators are believed to be primarily responsible for the systemic manifestations of acute pancreatitis and its associated distant organ dysfunction. The predictable nature in which they are produced may allow for novel approaches to treating this disease. Am J Surg.     

Novel nonsense mutation in the hypoxanthine guanine phosphoribosyltransferase gene and nonrandom X-inactivation causing Lesch-Nyhan syndrome in a female patient

BACKGROUND: Patient views are important in the evaluation of the quality of health care. The use of surveys needs to be evaluated to determine their cost-effectiveness and benefits. OBJECTIVES: To determine the costs of conducting patient opinion surveys in general practice and to find out how effective patient surveys are in stimulating changes which are beneficial for patient care. METHOD: Postal questionnaire to all 102 medical audit advisory groups (MAAGs) and 98 family health services authorities (FHSAs) in England and Wales, followed by postal questionnaire to 302 general practices reported to have conducted surveys, sampled by the type or questionnaire used. Numbers of MAAGs and FHSAs reporting surveys in general practice; types of questionnaire used; estimated costs; changes made; and benefits identified were measured. RESULTS: Eighty-five (83%) MAAGs and 75 (77%) FHSAs responded. One hundred and fifty-four (96%) of MAAGs or FHSAs reported survey activity. Types of questionnaire used were 1) designed by the practice, 2) designed by the MAAG or FHSA, possibly in collaboration with a practice, or 3) standard 'off-the-shelf'. One hundred and thirty-three (44%) practices responded. Total costs to a practice of conducting a survey ranged from nothing to over 2200 Pounds. Questionnaires designed by the practice are likely to be more costly than other designs. Some practices had surveys provided free of charge by MAAG or FHSA. Sixty-one per cent of practices said changes had been implemented and a further 22% of practices said changes were planned. The most common change was to appointment systems. Benefits were identified for patients, staff, the practice, the MAAG or FHSA and the NHS. Surveys also brought benefits in relationships and understanding. Only 8.2% of practices felt the costs of surveys outweighed the benefits. CONCLUSIONS: Many practices are surveying patients' opinions. Surveys can be costly but MAAGs and FHSAs can provide expertise and resources. Surveys using any of the types of questionnaire are likely to lead to changes and identifiable benefits. Benefits of surveys are perceived by the majority of practices to outweigh the costs.     

Clinical features of primary ovarian failure caused by a point mutation in the follicle-stimulating hormone receptor gene

The recent finding that a mutation in the FSH receptor gene causes ovarian dysgenesis prompted the present study to determine the phenotype caused by this mutation. Twenty-two patients with ovarian dysgenesis and a 566C-->T mutation in the FSH receptor gene (designated FSH-resistant ovaries or FSHRO) were compared with 30 clinically similar patients with ovarian dysgenesis (designated ODG) who did not have this mutation. The genealogical studies suggested a founder effect of the FSH receptor gene mutation in Finland. Clinically, both groups of patients were characterized by primary or early secondary amenorrhea, variable development of secondary sex characteristics, and high serum levels of FSH and LH. Notable differences were observed in median adult height (FSHRO patients were shorter) and the occurrence of follicles judged by transvaginal sonography (observed in 6 of 8 FSHRO vs. 1 of 11 ODG) and ovarian histology (present in all 9 FSHRO vs. 1 of 4 ODG). These findings suggest that a subset of ovarian dysgenesis patients with the FSH receptor mutation 566C-->T is pathogenetically distinct, possibly due to residual receptor activity, and that these patients can be tentatively identified by demonstrating the presence of ovarian follicles and confirmed by mutation analysis.     

A novel nonsense mutation, S466Xa in exon 10 of the cystic fibrosis transmembrane conductance regulator gene

The first recorded occurrence of Aedes (Stegomyia) albopictus (Skuse) in Curitiba, Paraná, South of Brazil, was described. The collection was carried out by means of aspirator in human bait.     

Selective complement C1s deficiency caused by homozygous four-base deletion in the C1s gene

The authors report the case of a 50 year old man with pseudowanthoma elastica with a history of myocardial infarction and severe aortic regurgitation. Angiography showed multiple coronary artery aneurysms and aneurysmal dilatation of the aortic annulus. The outcome after triple coronary bypass surgery with aortic valve replacement in a valved Bentall conduit was favourable. Pseudoxanthoma elastica is a rare condition in which the prognosis depends on the degree of vascular involvement. In this context, coronary artery aneurysms and aneurysmal dilatation of the aorta are rare complications.     

Genetic cholesteryl ester transfer protein deficiency is extremely frequent in the Omagari area of Japan. Marked hyperalphalipoproteinemia caused by CETP gene mutation is not associated with longevity

Low levels of HDL cholesterol have been clearly demonstrated to be associated with an increased incidence of coronary heart disease, strongly suggesting that HDL particles have an antiatherogenic function. However, little information has been available concerning the atherogenicity of a marked hyperalphalipoproteinemia (HALP). There is no agreement about whether plasma cholesteryl ester transfer protein (CETP) deficiency is associated with an antiatherogenic state or not, although this disorder was reported to be one of the major causes of marked HALP. In the current study, we have found a unique area (Omagari City, Akita Prefecture, Japan) where CETP deficiency caused by a G-to-A mutation at the 5' splice donor site of intron 14 in the CETP gene is extremely frequent. In Omagari City, the mutation was detected more than 20 times more frequently and the prevalence of a marked HALP with plasma HDL cholesterol > or = 2.58 mmol/L (100 mg/dL) was 5 to 10 times higher than in other areas of Japan. This discovery has made it possible to perform a large population-based study concerning the atherogenicity of a marked elevation of HDL cholesterol in a genetically more homogeneous population. There was a statistically significant U-shaped relationship between HDL cholesterol levels and the incidence of ischemic changes in electrocardiograms. In cases of HDL cholesterol < 1.81 mmol/L (70 mg/dL), the incidence increased in proportion to the levels of HDL cholesterol. The frequency of the CETP gene mutation was higher in patients with coronary heart disease than in healthy control subjects. In subjects aged > 80 years, the prevalence of both marked HALP and the intron 14 splicing defect was significantly lower than in the younger generation. The current study indicated for the first time that a marked HALP caused by CETP gene mutation may not represent a longevity syndrome, suggesting the importance of reevaluation of the clinical significance and pathophysiology of a marked HALP.     

Fracture of the sacrum caused by bone deficiency

This case report discusses a very rare condition; Papillon-Lefèvre Syndrome. This is an autosomal recessive condition in which the main features are hyperkeratosis of the palms and the soles and severe periodontal destruction.     

CMO I deficiency caused by a point mutation in exon 8 of the human CYP11B2 gene encoding steroid 18-hydroxylase (P450C18)

Corticosterone methyloxidase I (CMO I) deficiency is an autosomal recessive disorder of aldosterone biosynthesis. To determine further the molecular genetic basis of CMO I deficiency, a patient of Turkish origin that suffered from CMO I deficiency was studied. Nucleotide sequencing of the PCR-amplified exons from the genomic DNA of this patient revealed a single point mutation CTG (leucine) CCG (proline) at codon 461 in exon 8 of CYP11B2, which is involved in the putative heme binding site of steroid 18-hydroxylase (P450(C18)). The expression study using a cDNA introducing the point mutation revealed that the amino acid substitution totally abolishes the P450(C18)p3 enzyme activities required for conversion of 11-deoxycorticosterone to aldosterone, even though the mutant product was detected in the mitochondrial fraction of the transfected cells. These results suggest that this point mutation causes CMO I deficiency.