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By using homozygosity mapping and positional cloning, we have shown that adult-onset type II citrullinemia (CTLN2) is caused by mutations of the SLC25A13 gene, which is localized on chromosome 7q21.3 and encodes a mitochondrial solute carrier protein named citrin. So far, we have reported nine mutations, most of which cause loss of citrin, and we have established several methods for DNA diagnosis. These methods have shown that more than 90% of the patients diagnosed as suffering from CTLN2 by enzymatic analysis carry SLC25A13 mutations in both alleles, indicating that CTLN2 is caused by citrin deficiency. Furthermore, by using the same DNA diagnosis methods, we discovered that 70 neonates or infants suffering from a particular type of neonatal hepatitis carry the same SLC25A13 mutations. Since the symptoms of the neonates are different from those of the more severe CTLN2 and usually ameliorate without special treatment, we designated the neonatal disease neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD). We conclude that citrin deficiency causes NICCD in neonates and CTLN2 in adults through the additional effects of genetic or environmental modifiers. Since the function of citrin, together with that of an isoform, aralar, was found to be as a mitochondrial aspartate glutamate carrier, the various symptoms of NICCD and CTLN2 may be understood as caused by defective aspartate export from the mitochondria to the cytosol and defects in the malate aspartate shuttle. It is, however, still difficult to understand the cause of the hepatic deficiency of argininosuccinate synthetase protein in CTLN2. [ABSTRACT FROM AUTHOR]

The mitochondrial aspartate/glutamate carrier catalyzes an important step in both the urea cycle and the aspartate/malate NADH shuttle. Citrin and aralar1 are homologous proteins belonging to the mitochondrial carrier family with EF-hand Ca2+-binding motifs in their N-terminal domains. Both proteins and their C-terminal domains were overexpressed in Escherichia coli, reconstituted into liposomes and shown to catalyze the electrogenic exchange of aspartate for glutamate and a H+. Overexpression of the carriers in transfected human cells increased the activity of the malate/aspartate NADH shuttle. These results demonstrate that citrin and aralar1 are isoforms of the hitherto unidentified aspartate/glutamate carrier and explain why mutations in citrin cause type II citrullinemia in humans. The activity of citrin and aralar1 as aspartate/glutamate exchangers was stimulated by Ca2+ on the external side of the inner mitochondrial membrane, where the Ca2+-binding domains of these proteins are localized. These results show that the aspartate/glutamate carrier is regulated by Ca2+ through a mechanism independent of Ca2+ entry into mitochondria, and suggest a novel mechanism of Ca2+ regulation of the aspartate/malate shuttle. [ABSTRACT FROM AUTHOR]

The proband was born at 36 weeks, appropriate for gestational age, to nonconsanguineous white parents. There was no evidence of hyperbilirubinemia or intrahepatic cholestasis in the neonatal period, and she had normal newborn screen results. She presented with 3 episodes of life-threatening bleeding and anemia. The diagnostic evaluation for her bleeding diathesis revealed an abnormal clotting profile with no biochemical evidence for hepatocellular damage. She was incidentally noted to have severe growth deceleration that failed to respond to 502 kJ/kg (120 kcal/kg) per day of protein-hydrolyzed formula. An extensive diagnostic workup for failure to thrive, which was otherwise normal, included plasma amino acid analysis that revealed hyperglutaminemia and citrulline levels within the reference range. Testing of a repeat sample revealed isolated hypercitrullinemia. No argininosuccinic acid was detected. Her ammonia level and urine orotic acid were within the reference ranges. Subsequent plasma amino acid analysis exhibited a profile suggestive of neonatal intrahepatic cholestasis caused by citrin deficiency with elevations in citrulline, methionine, and threonine. Western blotting of fibroblasts demonstrated citrin deficiency, and a deletion for exon 3 was found in the patient's coding DNA of the SLC25A13 gene. On the basis of the experience with adults carrying this condition, the patient was given a high-protein, low-carbohydrate diet. The failure to thrive and bleeding diathesis resolved. When compliance with the dietary prescription was relaxed, growth deceleration was again noted, although significant bleeding did not recur. This is the first report of an infant of Northern European descent with citrin deficiency. The later age at presentation with failure to thrive and bleeding diathesis and without obvious evidence of neonatal intrahepatic cholestasis expands the clinical spectrum of citrin deficiency. This case emphasizes the importance of continued dietary control and growth monitoring in children with neonatal intrahepatic cholestasis caused by citrin deficiency and identifies a new metabolic entity responsible for failure to thrive. [ABSTRACT FROM AUTHOR]