Abstract
The V617F activating point mutation in Jak2 is associated with a proportion of myeloproliferative disorders. In normal hematopoietic cells, Jak2 signals only when associated with a growth factor receptor, such as the erythropoietin receptor (EpoR). We sought to identify the molecular requirements for activation of Jak2V617F by introducing a point mutation in the FERM domain (Y114A), required for receptor binding. Whereas BaF3.EpoR cells are readily transformed by Jak2V617F to Epo independence, we found that the addition of the FERM domain mutation blocked transformation and the induction of reactive oxygen species. Further, while cells expressing Jak2V617F had constitutive activation of STAT5, cells expressing Jak2V617F/Y114A did not, suggesting that signaling is defective at a very proximal level. In addition, expression of the Myc and Pim proto-oncogenes by Jak2V617F was found to be FERM domain dependent. An inducible constitutively active STAT5 mutant expressed in BaF3 cells was sufficient to induce Myc and Pim. Finally, the FERM domain in Jak2V617F was also required for abnormal hematopoiesis in transduced primary murine fetal liver cells. Overall, our results suggest that constitutive activation of Jak2 requires an intact FERM domain for a transforming phenotype, and is necessary for activation of the major target of Jak2, STAT5.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Cell Line, Tumor
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Cell Transformation, Neoplastic / genetics
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Cell Transformation, Neoplastic / metabolism*
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Enzyme Activation / genetics
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Erythropoietin / genetics
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Erythropoietin / metabolism
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Gene Expression Regulation, Neoplastic* / genetics
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Hematopoiesis, Extramedullary / genetics
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Humans
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Janus Kinase 2 / biosynthesis*
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Janus Kinase 2 / genetics
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Liver / embryology
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Mice
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Mutation, Missense*
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Myeloproliferative Disorders / genetics
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Myeloproliferative Disorders / metabolism*
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Myeloproliferative Disorders / pathology
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Protein Serine-Threonine Kinases / biosynthesis*
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Protein Serine-Threonine Kinases / genetics
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Protein Structure, Tertiary / genetics
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Proto-Oncogene Proteins / biosynthesis*
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins c-myc / biosynthesis*
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Proto-Oncogene Proteins c-myc / genetics
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Proto-Oncogene Proteins c-pim-1 / biosynthesis*
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Proto-Oncogene Proteins c-pim-1 / genetics
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Reactive Oxygen Species / metabolism
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Receptors, Erythropoietin / genetics
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Receptors, Erythropoietin / metabolism
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STAT5 Transcription Factor / genetics
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STAT5 Transcription Factor / metabolism
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Signal Transduction / genetics
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Transduction, Genetic
Substances
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MYC protein, human
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Myc protein, mouse
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PIM2 protein, human
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Pim2 protein, mouse
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Proto-Oncogene Proteins
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Proto-Oncogene Proteins c-myc
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Reactive Oxygen Species
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Receptors, Erythropoietin
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STAT5 Transcription Factor
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Erythropoietin
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JAK2 protein, human
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Jak2 protein, mouse
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Janus Kinase 2
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PIM1 protein, human
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Pim1 protein, mouse
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-pim-1