Mice deficient in the urea-cycle enzyme, carbamoyl phosphate synthetase i, die during the early neonatal period from hyperammonemia
✍ Scribed by J. Paul Schofield; Dr. J. Paul Schofield; Timothy M. Cox; C. Thomas Caskey; Maki Wakamiya
- Publisher
- John Wiley and Sons
- Year
- 1999
- Tongue
- English
- Weight
- 122 KB
- Volume
- 29
- Category
- Article
- ISSN
- 0270-9139
No coin nor oath required. For personal study only.
✦ Synopsis
Ammonia liberated during amino acid catabolism in mammals is highly neurotoxic and is detoxified by the five enzymes of the urea cycle that are expressed within the liver. Inborn errors of each of the urea cycle enzymes occur in humans. Carbamoyl phosphate synthetase I (CPSase I; EC 6.3.4.16) is located within the inner mitochondrial matrix and catalyzes the initial rate-limiting step of the urea cycle. Unless treated, complete deficiency of CPSase I, a rare autosomal recessive disease, causes death in newborn infants. Survivors are often mentally retarded and suffer frequent hyperammonemic crises during intercurrent illness or other catabolic stresses. Biochemically, CPSase I deficiency is characterized by high levels of blood ammonia, glutamine, and alanine, with low or absent citrulline and arginine levels. As a first step toward the development of gene therapy directed to the hepatocyte, we have generated a CPSase I-deficient mouse by gene targeting. Mice with homozygous disruption of CPSase I (CPSase [؊/؊] mice) die within 36 hours of birth with overwhelming hyperammonemia, and without significant liver pathology. This animal is a good model of human CPSase I deficiency. (HEPATOLOGY 1999;29:181-185.)
The hepatic urea cycle detoxifies ammonia liberated by the degradation of amino acids, producing urea. 1 The cycle comprises two mitochondrial enzymes, carbamoyl phosphate synthetase (CPSase I) and ornithine transcarbamylase (OTC), and three cytoplasmic enzymes: argininosuccinic acid synthetase, argininosuccinate lyase, and arginase. 2 The predominant expression of CPSase I and OTC within periportal hepatocytes effectively restricts the urea cycle to the liver. 3 In
Methods
All animals used in these experiments received humane care under the UK Animals (Scientific Procedures) Act, 1986.
Construction of Targeting Vector (Fig. 1). The production of carbamoyl phosphate from ammonia, carbon dioxide, and water involves two partial reactions, each requiring the binding of adenosine triphosphate. To disrupt the CPSase I gene, a construct was designed to interrupt the sequences encoding the most 5Ј nucleotidebinding domain. The mouse CPSase I cDNA and the genomic organization was unknown, although this gene is highly conserved throughout evolution. 8,9 We were able to predict the mouse genomic structure around the most 5Ј nucleotide-binding domain by the Abbreviations: CPSase I, carbamoyl phosphate synthetase I; OTC, ornithine transcarbamylase; PCR, polymerase chain reaction; ES, embryonic stem cell.