An activating splice donor mutation in the thrombopoietin gene causes hereditary thrombocythaemia

Essential thrombocythaemia (ET) is a chronic myeloproliferative syndrome due to sustained proliferation of megakaryocytes, which results in elevated numbers of circulating platelets, thrombotic or haemorrhagic episodes and occasional leukaemic transformation1. The cause of ET is unknown. Hereditary thrombocythaemia (HT) with autosomal−dominant transmission has been described with manifestations similar to those of sporadic ET2−8. As the thrombopoietin gene (THRO) encodes a lineage−restricted growth factor with profound stimulatory effects on megakaryopoiesis and platelet production9,10, we tested the hypothesis that HT results from a mutation in the human THRO gene. In a Dutch family with eleven affected individuals7, the thrombopoietin protein (TPO) concentrations in serum were consistently elevated in individuals with HT. We derived an intragenic CA marker for the human THRO gene and performed linkage analysis in fourteen informative meioses in this family. This resulted in a lod score of 3.5 at =0. A GC transversion was found in the splice donor site of intron 3 of the THRO gene in all affected family members. This mutation leads to THRO mRNAs with shortened 5'−untranslated regions (UTR) that are more efficiently translated than the normal THPO transcripts. We conclude that a splice donor mutation in THPO leads to systemic overproduction of TPO and causes thrombocythaemia.

1. Murphy, S., Peterson, P., Hand, H. & Laszlo, J. Experience of the Polycythemia Vera Study Group with essential thrombocythemia: a final report on diagnostic criteria, survival, and leukemic transition by treatment. Semin. Hematol. 34, 29−39 (1997) | PubMed  | ISI | ChemPort |
2. Pickers, M. & Speck, B. Thrombocythaemia: familial occurrence and transition into blastic crisis. Acta Haematol. 51, 257−265 (1974) | PubMed  |
3. Slee, P.H.T.J., van Everdingen, J.J.E., Geraedts, J.P.M., te Velde, J. & den Ottolander, G.J. Familial myeloproliferative disease. Acta Med. Scand. 210, 321−327 (1981) | PubMed  | ISI | ChemPort |
4. Eyster, M.E. et al. Familial essential thrombocythemia. Am. J. Med. 80, 497−502 (1986) | PubMed  | ISI | ChemPort |
5. Fernandez-Robles, E., Vermylen, C., Martiat, P., Ninane, J. & Cornu, G. Familial essential thrombocythemia. Pediatr. Hematol. Oncol. 7, 373−376 (1990) | PubMed  | ChemPort |
6. Williams, E.G. & Shahidi, NT. Benign familial thrombocytosis. Am. J. Hematol. 37, 124−125 (1991) | PubMed  | ISI | ChemPort |
7. Schlemper, R.J., van der Maas, A.P.C.& Eikenboom, J.C.J. Familial essential thrombocythemia: clinical characteristics of 11 cases in one family. Ann. Hematol. 68, 153−158 (1994) | PubMed  | ISI | ChemPort |
8. Kikuchi, M. et al. Familial thrombocytosis. Br. J. Haematol. 89, 900−902 (1995) | PubMed  | ISI | ChemPort |
9. Kaushansky, K. Thrombopoietin: understanding and manipulating platelet production. Annu. Rev. Med. 48, 1−11 (1997) | Article | PubMed  | ISI | ChemPort |
10. Eaton, D.L. & de Sauvage, F.J. Thrombopoietin: the primary regulator of megakaryocytopoiesis and thrombopoiesis. Exp. Hematol. 25, 1−7 (1997) | PubMed  | ISI | ChemPort |
11. van Dijken, P.J., Woldendorp, K.H. & van Wouwe, J.P. Familial thrombocytosis in infancy presenting with a leukaemoid reaction. Acta Paediatr. 85, 1132−1134 (1996) | PubMed  | ChemPort |
12. Kaushansky, K. et al. Promotion of megakaryocyte progenitor expansion and differentiation by thec-MpI ligandthrombopoietin. Nature 369, 568−571 (1994) | Article | PubMed  | ISI | ChemPort |
13. Zhou, W., Toombs, C.F., Zou, T., Quo, J. & Robinson, M.O. Transgenic mice overexpressing human c-mpl ligand exhibit chronic thrombocytosis and display enhanced recovery from 5-fluorouracil or antiplatelet serum treatment. Blood 89, 1551−1559 (1997) | PubMed  | ISI | ChemPort |
14. Van, X.Q. et al. Chronic exposure to retroviral vector encoded MGDF (mpl-ligand) induces lineage-specific growth and differentiation of megakaryocytes in mice. Blood, 86, 4025−4033 (1995) | PubMed  |
15. Tahara, T. et al. A sensitive sandwich ELISA for measuring thrombopoietin inhuman serum: serum thrombopoietin levels in healthy volunteers and in patients with haemopoietic disorders. Br. J. Haematol. 93, 783−788 (1996) | Article | PubMed  | ISI | ChemPort |
16. Krawczak, M., Reiss, J. & Cooper, D.N. The mutational spectrum of single base-pair substitutions in mRNA splice junctions of human genes: causes and consequences. Hum. Genet. 90, 41−54 (1992) | PubMed  | ISI | ChemPort |
17. Chang, M.S. et al. Cloning and characterization of the human megakaryocyte growth and development factor (mgdf) gene. J. Biol. Chem. 270, 511−514 (1995) | Article | PubMed  | ISI | ChemPort |
18. Sohma, Y. et al. Molecular cloning and chromosomal localization of the human thrombopoietin gene. FEBS Lett 353, 57−61 (1994) | Article | PubMed  | ISI | ChemPort |
19. Kamura, T., Handa, H., Hamasaki, N. & Kitaijma, S. Characterization of the human thrombopoietin gene promoter-a possible role of an Ets transcription factor, E4TF1/GABP. 7. Biol. Chem. 272, 11361−11368(1997)
20. Nielsen, H., Engelbrecht, E., Brunak, S. & von Heijne, G. Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng. 10, 1−6 (1997) | Article | PubMed  | ISI | ChemPort |
21. Stoffel, R., Wiestner, A. & Skoda, R.C. Thrombopoietin in thrombocytopenic mice: evidence against regulation at the mRNA level and for a direct regulatory role of platelets. Blood 87, 567−573 (1996) | PubMed  | ISI | ChemPort |
22. Sachs, A.B., Sarnow, P.& Hentze, M.W. Starting at the beginning, middle, and end: translation initiation in eukaryotes. Cell 89, 831−838 (1997) | Article | PubMed  | ISI | ChemPort |
23. Jackson, R.J. & Wickens, M. Translational controls impinging on the 5'-untranslated region and initiatio factor proteins. Curr. Opin. Genet. DeveL 7, 233−241 (1997) | Article | ISI | ChemPort |
24. D'Andrea, A.D. Cytokine receptors in congenital hematopoietic disease. N. Engl. J. Med. 330, 839−846 (1994) | Article | PubMed  | ChemPort |
25. Ott, J. A computer program for linkage analysis of general human pedigrees. Am. J. Hum. Genet. 28, 528−529 (1967)
26. Chomczynski, P. & Sachhi, N. Single-step method of RNA isolation by acid guanidium thiocyanate-phenol-chloroform extraction. Analyt. Biochem. 162, 156−159 (1987) | Article | PubMed  | ISI | ChemPort |
27. Seed, B. & Aruffo, A. Molecular cloning of the CD2 antigen, the T-cell erythrocyte receptor, by a rapid immunoselection procedure. Proc. Natl. Acad. Sci. USA 84, 3365−3369 (1987) | PubMed  | ChemPort |
28. Berger, J., Hauber, J., Hauber, R., Geiger, R. & Cullen, B.R. Secreted placenta! alkaline phosphatase: a powerful new quantitative indicator of gene expression in eukaryotic cells. Gene 66, 1−10 (1988) | Article | PubMed  | ISI | ChemPort |

	Top

	Top