Molecular analysis of the AGXT gene and linkage studies in primary hyperoxaluria type 1

Primary hyperoxaluria type 1 (PH1) is a rare, autosomal recessive error of metabolism, which leads to renal stones, loss of renal function and systemic accumulation of oxalate in affected individuals. PH1 is caused by deficiency of the hepatic peroxisomal enzyme alanineiglyoxylate aminotransferase (AGT) and diagnosis of PHI currently requires measurement of AGT enzyme activity in a liver biopsy. The gene AGXT coding for the AGT protein has been sequenced and mapped to chromosome 2q37.3. This project used two different molecular genetic approaches to further characterise the disease and improve its diagnosis. The frequency of known mutations and polymorphisms in the AGXT gene was determined in DNA from 79 unrelated PHI patients. The same samples were also screened for the presence of new mutations using single strand conformation polymorphism (SSCP) analysis. Fifteen new mutations and polymorphisms were identified and characterised by DNA sequencing. Expression of these variants in liver mRNA was studied whenever possible. The results and the potential consequences of the mutations for AGT are discussed in context with the clinical and enzymological data of the patients. The other aspect of the project was to identify new linkage markers suitable for family studies of PH 1. Three microsatellite markers were evaluated and their proximity to the AGXT locus was determined to establish the recombination risk. These markers were successfully introduced for the prenatal diagnosis of PH1