A mouse mutant with glutathionuria was discovered by screening for amino acidurias in the progeny of ethylnitrosourea-mutagenized mice. Total glutathione concentration was increased in both blood and urine but decreased in liver homogenates from affected mice. Glutathionuric mice exhibited lethargy, severe growth failure, shortened life spans and infertility. ␥-Glutamyl transpeptidase activity was deficient in kidney homogenates of glutathionuric mice. The glutathionuric phenotype in these mice is inherited as an autosomal recessive trait. This mouse mutant will be a useful animal model for the study of ␥-glutamyl transpeptidase physiology and glutathione metabolism.GSH is the most abundant cellular thiol and functions as the principal reducing reagent in all cell types (1). A partial listing of the antioxidative functions of GSH include: protection against mitochondrial damage, protection against oxygen toxicity in the lung, protection against lipid peroxidation, detoxification of electrophilic compounds through conjugation, preservation of proper sulfide bonds in proteins, a postulated function in anticarcinogenesis, and a role in the immune system (2). GSH metabolism also provides a source of cysteine for cells (3). ␥-Glutamyl transpeptidase (␥-GT; EC 2.3.2.2) 1 catalyzes the initial step in the degradation of GSH (4). ␥-GT is a key step in the ␥-glutamyl cycle (5), a series of degradative and synthetic reactions that mediate cellular GSH metabolism. Several reviews of ␥-GT physiology and function have been published (4 -6), but despite intensive investigation, the exact role ␥-GT plays in GSH metabolism or its putative contribution to renal amino acid transport have not been definitively determined. Bound to secretory endothelial cell membranes in several organs but predominantly in proximal renal tubule cells, ␥-GT participates in the transmembrane transport of GSH and in interorgan GSH exchange ( Fig. 1) (7). Meister (8) proposed that ␥-GT also contributes to amino acid transport in the proximal renal tubule through transpeptidation of GSH and subsequent tubule cell uptake of ␥-glutamyl amino acids. In vivo model systems that have lost ␥-GT activity are an exquisitely powerful tool for the study of ␥-GT function and its relationship to GSH metabolism. Administration of chemical inhibitors of ␥-GT to animals results in both glutathionuria and glutathionemia (9), but chemically treated animal models are limited by several drawbacks including the temporary nature of inhibition and the difficulty of long-term continuous inhibitor administration. Also, the degree and specificity of enzyme inhibition in various tissues (particularly the brain) of these chemically treated animals is unknown. Genetic ␥-GT deficiency has been described in only five humans (6), and the effects of various different disease states or environmental influences upon ␥-GT deficient individuals cannot be adequately evaluated given the rarity of the disorder. A genetic animal model of total ␥-GT deficiency overcomes the limitations of previ...
Abstract— Seven‐day‐old rats were injected intraperitoneally with l‐phenylalanine (1 g/kg) and the time course of brain polyribosome disaggregation and changes in brain levels of phenylalanine, tryptophan and tyrosine were determined. Disaggregation of brain polyribosomes preceded the increase in levels of phenylalanine in brain, and followed the same time course as depletion of tryptophan from brain. The effects of several metabolites of phenylalanine (which are formed in phenylketonuria) on protein synthesis in vitro was determined for brain and liver systems. None of the compounds tested was inhibitory at concentrations below 10 mM and in all cases hepatic protein synthesis was more sensitive to inhibition than was the corresponding system from brain. Ribosomal dimers, formed in brain after injection of phenylalanine, were incapable of supporting high levels of protein synthesis in vitro, a finding that suggested that the inhibition of protein synthesis in vitro in cell‐free systems of brain tissue after injection of phenylalanine into young rats was mediated by disaggregation of brain polyribosomes associated with tryptophan deficiency in brain.
A mouse mutant with sarcosinemia was found by screening the progeny of ethylnitrosourea-mutagenlzed mice for aminoacidurias. Paper chromatography, column chromatography, and gas chromatography-mass spectrometry identified high levels of sarcosine in the urine of the mutant mice. While sarcosine cannot be detected in the urine or plasma of normal mice, the urinary sarcosine level of 102 ± 58 mmol per g of creatinine in the mutant mice was at the upper range of the urinary levels (1.545 mmol of sarcosine per g of creatinine) observed in humans with sarcosinemia. Similarly, the plasma sarcosine level of 785 ± 153 pzmol/liter in the sarcosinemic mice was at the upper range of the plasma sarcosine levels (53-760 pzmol/liter) observed in affected humans. Sarcosine dehydrogenase [sarcosine:(acceptor) oxidoreductase (demethylating), EC 1.5.99.1] activity was deficient in sarcosinemic mice. The sarcosinuria phenotype in these mice was inherited as an autosomal recessive trait. This mouse mutant provides a useful genetic model for human sarcosinemia and for development of therapeutic approaches for genetic disease.Genetically modified mice are increasingly being used as models for diseases and for development of approaches for treatments such as gene therapy. Although some useful mouse strains arose spontaneously, two basic approaches have been developed to produce mouse mutants. When the gene is known beforehand, site-directed mutation induced by homologous recombination in embryo stem cells is a powerful approach for creating mouse mutants (1). When only the phenotype is known, germline mutagenesis followed by backcrossing and phenotypic screening has been successful. The latter approach utilizing ethylnitrosourea (ENU) mutagenesis has been used to develop hyperphenylalaninemic mice strains, deficient in phenylalanine hydroxylase or GTP cyclohydrolase activities (2, 3), and muscular dystrophic mice with defective dystrophin (4).Mouse models exist for a few of the inborn errors of metabolism (IEM) (5). Many genes related to IEM in humans have been defined and could be targeted for disruption in mice by homologous recombination. Others, however, have not been identified to date. In many IEM, such as cystinosis or Hartnup disease, the defective protein has yet to be identified. Furthermore, several disorders with poorly defined metabolic defects such as lactic acidosis or 3-methylglutaconic aciduria also have poorly defined enzymatic deficiencies (6). Genetic mouse models for these IEM should be useful for assigning the chromosomal location and for eventual isolation of the relevant genes (7).This report describes a mouse mutant with sarcosinemia found through metabolic screening of progeny of ENUmutagenized mice. The breeding scheme and ENU mutagenesis protocol were identical to that used for creating the hyperphenylalaninemic mutants (8). The phenotype testing was based on the metabolic screens that have been developed for newborn screening (9, 10). Of 135 pedigrees evaluated, one mutant strain was found to have ...
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