Constitutive activation of the JAK2/STAT5 signal transduction pathway correlates with growth factor independence of megakaryocytic leukemic cell lines.
The factor-independent Dami/HEL and Meg-01 and factor-dependent Mo7e leukemic cell lines were used as models to investigate JAK/STAT signal transduction pathways in leukemic cell proliferation. Although Dami/HEL and Meg-01 cell proliferation in vitro was independent of and unresponsive to exogenous cytokines including granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), IL-6, thrombopoietin (TPO), and tumor necrosis factor-alpha (TNF-alpha), the growth of Mo7e cells was dependent on hematopoietic growth factors. When these cell lines were cultured in medium without cytokines, a constitutively activated STAT-like DNA-binding factor was detected in nuclear extracts from both Dami/HEL and Meg-01 cells. However, the STAT-like factor was not detectable in untreated Mo7e cells, but was activated transiently in Mo7e cells in response to cytokine treatments. The constitutively activated and cytokine-induced STAT-like DNA-binding factor in these three cell lines was identified as STAT5 by oligonucleotide competition gel mobility assays and by specific anti-STAT antibody gel supershift assays. Constitutive activation of JAK2 also was detected in the factor-independent cell lines, but not in Mo7e cells without cytokine exposure. Meg-01 cells express a p185 BCR/ABL oncogene, which may be responsible for the constitutive activation of STAT5. Dami/HEL cells do not express the BCR/ABL oncogene, but increased constitutive phosphorylation of Raf-1 oncoprotein was detected. In cytokine bioassays using growth factor-dependent Mo7e and TF-1 cells as targets, conditioned media from Dami/HEL and Meg-01 cells did not show stimulatory effects on cell proliferation. Our results indicate that the constitutive activation of JAK2/STAT5 correlates with the factor-independent growth of Dami/HEL and Meg-01 cells. The constitutive activation of JAK2/STAT5 in Dami/HEL cells is triggered by a mechanism other than autocrine cytokines or the BCR/ABL oncoprotein.
Liu
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Division of Medical Oncology and Hematology, the Department of Internal Medicine, University of South Florida College of Medicine, H. Lee Moffitt Cancer Institute, Tampa 33612, USA. liur@moffitt.usf.edu
Fan
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Garcia
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Jove
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Zuckerman
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D002455:Cell Division / Q000187:drug effects; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004789:Enzyme Activation; D016044:Fusion Proteins, bcr-abl / Q000502:physiology; D015973:Gene Expression Regulation, Leukemic; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006133:Growth Substances / Q000494:pharmacology*; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology; D015850:Interleukin-6 / Q000494:pharmacology; D053614:Janus Kinase 2; D007947:Leukemia, Megakaryoblastic, Acute / Q000378:metabolism / Q000473:pathology*; D008533:Megakaryocytes / Q000187:drug effects / Q000378:metabolism*; D008894:Milk Proteins; D009363:Neoplasm Proteins / Q000502:physiology; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D050799:STAT5 Transcription Factor; D015398:Signal Transduction / Q000187:drug effects; D013926:Thrombopoietin / Q000494:pharmacology; D015534:Trans-Activators / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D017077:Culture Media, Conditioned; D004268:DNA-Binding Proteins; D006133:Growth Substances; D007377:Interleukin-3; D015850:Interleukin-6; D008894:Milk Proteins; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D014409:Tumor Necrosis Factor-alpha; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D013926:Thrombopoietin; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl; C507924:JAK2 protein, human; D053614:Janus Kinase 2; D019908:Proto-Oncogene Proteins c-raf