Abstract
We have used peripheral blood T-lymphocyte cultures to analyze the hprt mutation in two Lesch-Nyhan syndrome males who are cousins and to confirm the carrier status of female members of the family. Both cDNA and genomic DNA sequencing studies show that this patient carries a hitherto undescribed single base deletion in the exon 5 donor splice site sequence (15:+1, δG, base number 31635). The largest cDNA product contained all nine hprt exons plus an insertion of 66 bases of intron 5, consistent with the use of a cryptic splice site in intron 5 (aag67/gtaagc). This splicing error would result in a chain terminating codon immediately after exon 5 (15:2–4, taa) and predicts a polypeptide of 133 amino acids. This loss of the normal splice donor site also results in multiple hprt mRNA species, combining the use of the cryptic splice site in intron 5 and splicing errors involving exons 2–6. In addition to defining a new Lesch-Nyhan mutation (hprtHenryville), these results provide insight into aberrant splicing of hprt mRNA in T-lymphocytes.
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Literature Cited
Lesch, M., and Nyhan, W.L. (1964). A familial disorder of uric acid metabolism and central nervous system function.Am. J Med Gener 36:561–570.
Sculley, D.G., Dawson, P.A., Emmerson, B.T., and Gordon, R.B. (1992). A review of the molecular basis of hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency.Hum. Genet.,90: 195–207.
Cole, J., and Skopek, T.R. (1994). ICPEMC Commitee on spontaneous mutation: Working paper 3: —Somatic mutant frequency, mutation rates and mutational spectra in the human population in vivo.Mutation Res. 304: 33–106.
Skopek, T.R., Recio, L., Simpson, D., Dallaire, L., Melançon, S.B., Ogier, H., O'Neill, J.P., Falta, M.T., Nicklas, J.A., and Albertini, R.J. (1990). Molecular analyses of a novel Lesch-Nyhan syndrome mutation (hprt Montreal) by use of T-lymphocyte cultures.Hum. Genet. 85: 111–116.
Sinnett, D., Lavergne, L., Melançon, S.B., Dallaire, L., Potier, M., and Labuda, D. (1988). Lesch-Nyhan syndrome: Molecular investigation of three French Canadian families using a hypoxanthine-guanme phosphoribosyltransferase cDNA probe.Hum. Genet. 81: 4–8.
Itiaba, K., Melançon, S.B., Dallaire, L., and Crawhall, J.C. (1978). Adenine phosphoribosyltransferase deficiency in association with sub-normal hypoxanthine phosphoribosyltransferase in families of Lesch-Nyhan patients.Biochem. Med. 19: 252–259.
O'Neill, J.P., McGinniss, M.J., Berman, J.K., Sullivan, L.M., Nicklas, J.A., and Albertini, R.J. (1987). Refinement of a T-lymphocyte cloning assay to quantify the in vivo thioguanine-resistant mutant frequency in humans.Mutagenesis 2: 87–94.
Yang, J-L., Maher, V.M., and McCormick, J.J. (1989). Amplification of cDNA from the lysate of a small clone of diploid human cells and direct DNA sequencing.Gene 83: 347–354.
Bronstein, S.M., Cochrane, J.E., Craft, T.R., Swenberg, J.A., and Skopek, T.R. (1991) Toxicity, Mutagenicity, and Mutational Spectra ofN-Ethyl-N-nitrosourea in Human Cell Lines with Different DNA Repair Phenotypes.Cancer Res. 51: 5188–5197.
Gibbs, R.A., Nguyen, P.N., Edwards A., Civitello, A.B., and Caskey, C.T. (1990). Multiplex DNA deletion detection and exon sequencing of the hypoxanthine phosphoribosyltransferase gene in Lesch-Nyhan families.Genomics 7: 235–244.
Edwards, A., Voss, H., Rice P., Civitello, A., Stegemann, J., Schwager, C., Zimmerman, J., Erfle, H., Caskey, C.T., and Ansorge, W. (1990). Automated DNA sequencing of the human HPRT locus.Genomics 6: 593–608.
Jolly, D.J., Okayama, H., Berg, P., Esty, A.C., Filpula, D., Bohlen, P., Johnson, G.G., Shively, J.E., Hunkapillar, T., and Freidmann, T. (1983) Isolation and characterization of a full-length expressible cDNA for human hypoxanthine phosphoribosyltransferase.Proc. Nat. Acad. Sci. U.S.A. 80: 477–481.
Andersson, B., Hou, S-M., and Lambert B. (1992). Mutations causing defective splicing in the humanhprt gene.Environ. Molec. Mutagenesis 20: 89–95.
Recio, L., Cochrane, J., Simpson, D., Skopek, T.R., O'Neill, J.P., Nicklas, J.A. and Albertini, R.J. (1990). DNA sequence analysis of in vivohprt mutation in human T-lymphocytes.Mutagenesis 5: 505–510.
Rossi, A.M., Tates, A.D., van Zeeland, A.A., and Vrieling, H. (1992). Molecular Analysis of Mutations Affectinghprt mRNA Splicing in Human Tlymphocytes in Vivo.Environ. Molec. Mutagenesis 19: 7–13.
Steingrimsdottir, H., Rowley, G., Dorado, G., Cole, J., and Lehmann, A.R. (1992). Mutations which alter splicing in the human hypoxanthine-guanine phosphoribosyltransferase gene.Nuc. Acids Res. 20: 1201–1208.
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Hunter, T.C., Melancon, S.B., Dallaire, L. et al. Germinal HPRT splice donor site mutation results in multiple RNA splicing products in T-lymphocyte cultures. Somat Cell Mol Genet 22, 145–150 (1996). https://doi.org/10.1007/BF02369904
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DOI: https://doi.org/10.1007/BF02369904