title: Hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency: identification of point mutations in Japanese patients with Lesch-Nyhan syndrome and hereditary gout and their permanent expression in an HPRT-deficient mouse cell line.
abstract: Two different single nucleotide transitions of hypoxanthine-guanine phosphoribosyltransferase (HPRT) were identified in a Japanese patient with Lesch-Nyhan syndrome (LNS) and a patient with hereditary gout. HPRT enzyme activities in the two patients were severely deficient, but the size and amount of mRNA were normal according to Northern analysis. Entire coding regions of HPRT cDNAs were amplified by PCR and sequenced. A G-to-A substitution at base 208 in exon 3, which predicted glycine 70 to arginine, was detected in the LNS patient (identical mutation with HPRT Utrecht). A C-to-A substitution at base 73 in exon 2, which predicted proline 25 to threonine, was detected in the gout patient (designated HPRT Yonago). We transfected normal HPRT cDNA, mutant cDNA with HRPT Utrecht or mutant cDNA with HPRT Yonago, respectively, to HPRT-deficient mouse cells and isolated permanent expression cell lines. The HPRT-deficient mouse cells had no detectable HPRT activity and a very low amount of HPRT mRNA. When the HPRT-deficient mouse cells were transfected with normal human cDNA, HPRT enzyme activity increased to 21.8% that of normal mouse cells. The mouse cells transfected with HPRT Utrecht showed no increase in HPRT activity; however, when the mouse cells were transfected with HPRT Yonago, the activity increased to 2.4% that of normal activity. The proliferative phenotypes of these cells in HAT medium and in medium containing 6-thioguanine were similar to those of skin fibroblasts from the patients. This series of studies confirmed that each of the two point mutations was responsible for the decreases in HPRT enzyme activity, and the proliferative phenotypes in HAT medium and medium containing 6-thioguanine.
authors.affiliation: Division of Child Neurology, Faculty of Medicine, Tottori University, Yonago, Japan.
authors.forename: J
authors.initials: J
authors.lastname: Tohyama
authors.forename: E
authors.initials: E
authors.lastname: Nanba
authors.forename: K
authors.initials: K
authors.lastname: Ohno
mesh_terms: D000293:Adolescent; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002478:Cells, Cultured; D002648:Child; D004247:DNA / Q000032:analysis; D017931:DNA Primers; D005347:Fibroblasts; D015971:Gene Expression Regulation, Enzymologic; D006073:Gout / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000172:deficiency* / Q000235:genetics* / Q000378:metabolism; D007564:Japan; D007926:Lesch-Nyhan Syndrome / Q000201:enzymology / Q000235:genetics*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection
chemical_list: D017931:DNA Primers; D012333:RNA, Messenger; D004247:DNA; D007041:Hypoxanthine Phosphoribosyltransferase