CHARACTERIZATION OF MOUSE AND HUMAN GTP CYCLOHYDROLASE-I GENES - MUTATIONS IN PATIENTS WITH GTP CYCLOHYDROLASE-I DEFICIENCY

GTP cyclohydrolase I is the first and rate-limiting enzyme for the bio synthesis of tetrahydrobiopterin in mammals. Previously, we reported t hree species of human GTP cyclohydrolase I cDNA in a human liver cDNA library (Togari, A., Ichinose, H., Matsumoto, S., Fujita, IC., and Nag atsu, T. (1992) Biochem. Biophys. Res. Commun. 187, 359-365). Furtherm ore, very recently, we found that the GTP cyclohydrolase I gene is cau sative for hereditary progressive dystonia with marked diurnal fluctua tion, also known as DOPA-responsive dystonia (Ichinose, H., Ohye, T., Takahashi, E., Seki, N., Hori, T., Segawa, M., Nomura, Y., Endo, K., T anaka, H., Tsuji, S., Fujita, K., and Nagatsu, T. (1994) Nature Geneti cs 8, 236-242). To clarify the mechanisms that regulate transcription of the GTP cyclohydrolase I gene and to generate multiple species of m RNA, we isolated genomic DNA clones for the human and mouse GTP cycloh ydrolase I genes. Structural analysis of the isolated clones revealed that the GTP cyclohydrolase I gene is encoded by a single copy gene an d is composed of six exons spanning similar to 30 kilobases. We sequen ced all exon/intron boundaries of the human and mouse genes. Structura l analysis also demonstrated that the heterogeneity of GTP cyclohydrol ase I mRNA is caused by an alternative usage of the splicing acceptor site at the sixth exon. The transcription start site of the mouse GTP cyclohydrolase I gene and the 5'-flanking sequences of the mouse and h uman genes were deter mined. We performed regional mapping of the mous e gene by fluorescence in situ hybridization, and the mouse GTP cycloh ydrolase I gene was assigned to region C2-3 of mouse chromosome 14. We identified missense mutations in patients with GTP cyclohydrolase I d eficiency and expressed mutated enzymes in Escherichia coli to confirm alterations in the enzyme activity.