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.     

Characterization of a lectin-like protein isolated from Lachesis muta snake venom

A lectin-like protein was isolated from L. muta venom by gel filtration on BIO Gel P-100 followed by column Chromatography on DEAE-sephades A-50. The protein eluted at 0.4 M Nacl in 0.01 Tris pH 7.3 and exhibited agglutinin activity toward 0+ human erythrocytes. The protein is a dimer with Mr 28 kDa. Amino acid analysis revealed high content of tryptophan and acid recidues and low content of cysteine and methionine residues. No neutral carbohydrates and sialic acid were detected. Circular dichroic spectrum shows 78% of B structure and 1% of alpha structure. In vitro experiments with erythrocytes from rat, rabbit and dog revealed strong agglutination while red blood cells from mice, sheep and goat were not agglutinated. In vivo experiments using anesthetized rats, a sharp and prolonged fall in the blood pressure was observed at protein dose of 1.5 mg/kg. Double dose of protein caused the death of the animal.     

Prevention of bromobenzene toxicity by N-acetylmethionine: correlation between toxicity and the impairment in O- and S-methylation of bromothiocatechols

Bromobenzene (800 mg/kg, ip) caused severe liver necrosis with massive hemorrhage in the golden Syrian hamster within the first 24 hr. Kidney injury was also observed. Treatment with N-acetylmethionine (NAM) at an ip dose of 1200 mg/kg at 5 hr after bromobenzene administration significantly protected the liver and kidney against injuries. Plasma glutamate pyruvate transaminase and blood urea nitrogen levels were substantially decreased in the NAM-treated animals. Histological evaluations confirmed these results. When the urinary neutral and phenolic metabolites of bromobenzene from NAM-treated and untreated hamsters were isolated and compared by GC and GC/MS, a striking result was observed in terms of O- and S-methylated thiol-containing metabolite formation. The NAM-treated animals showed approximately a 8- to 14-fold increase in the excretion of the four isomeric O- and S-methylated bromothiocatechols. These thiocatechols, which are now known to be the 3,4-series metabolites of bromobenzene, can undergo methylation at either the thiol or the hydroxyl functional group. The excretion of 3-S- and 4-S-methylated bromodihydrobenzene thiolols was also increased significantly in the NAM-treated hamster, but other neutral and phenolic metabolites were relatively unchanged. These results suggest that bromobenzene toxicity in the Syrian hamster may be associated with impaired methylation capabilities, an impairment that could be due to methionine and glutathione depletion.     

Effect of heme and vacuole deficiency on FRE1 gene expression and ferrireductase activity in Saccharomyces cerevisiae

Doppler sonography of the fetal descending aorta, renal artery, middle cerebral artery and umbilical artery in a population of 74 fetuses with a abdominal circumference below the fifth percentile of the reference limits were done. All fetuses were free from structural and chromosomal abnormalities. The pulsatility- and the resistance indices as well as the ratios between these indices from peripheral and cerebral vessels were calculated and correlated to the fetal distress. The measurement of the pulsatility index of the middle cerebral artery provided the best results in predicting the development of fetal distress. Better results were achieved by the use of ratios of pulsatility-indices of various vessels than by the examination of the vessels alone. Our results suggest the usefulness of the examination of the middle cerebral artery and their ratios compared to the renal artery.     

Sheep pulmonary adenomatosis: a unique model of retrovirus-associated lung cancer

Sheep pulmonary adenomatosis (SPA) is a contagious bronchiolo-alveolar carcinoma of sheep associated with an exogenous type D/B retrovirus known as jaagsiekte sheep retrovirus (JSRV). SPA represents a unique model for lung cancer, and studies on its aetiopathogenesis can provide further insight into the mechanisms of epithelial neoplasms.