Lnk inhibits Tpo-mpl signaling and Tpo-mediated megakaryocytopoiesis.
Thrombopoietin (Tpo) is the primary cytokine regulating megakaryocyte development and platelet production. Tpo signaling through its receptor, c-mpl, activates multiple pathways including signal transducer and activator of transcription (STAT)3, STAT5, phosphoinositide 3-kinase-Akt, and p42/44 mitogen-activated protein kinase (MAPK). The adaptor protein Lnk is implicated in cytokine receptor and immunoreceptor signaling. Here, we show that Lnk overexpression negatively regulates Tpo-mediated cell proliferation and endomitosis in hematopoietic cell lines and primary hematopoietic cells. Lnk attenuates Tpo-induced S-phase progression in 32D cells expressing mpl, and Lnk decreases Tpo-dependent megakaryocyte growth in bone marrow (BM)-derived megakaryocyte culture. Consistent with this result, we found that in both BM and spleen, Lnk-deficient mice exhibited increased numbers of megakaryocytes with increased ploidy compared with wild-type mice. In addition, Lnk-deficient megakaryocytes derived from BM and spleen showed enhanced sensitivity to Tpo during culture. The absence of Lnk caused enhanced and prolonged Tpo induction of STAT3, STAT5, Akt, and MAPK signaling pathways in CD41+ megakaryocytes. Furthermore, the Src homology 2 domain of Lnk is essential for Lnk's inhibitory function. In contrast, the conserved tyrosine near the COOH terminus is dispensable and the pleckstrin homology domain of Lnk contributes to, but is not essential for, inhibiting Tpo-dependent 32D cell growth or megakaryocyte development. Thus, Lnk negatively modulates mpl signaling pathways and is important for Tpo-mediated megakaryocytopoiesis in vivo.
Tong
Wei
W
Whitehead Institute for Biomedical Research, Nine Cambridge Center, Cambridge, MA 02142, USA.
Lodish
Harvey F
HF
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D015153:Blotting, Western; D001854:Bone Marrow Cells / Q000378:metabolism; D002455:Cell Division; D016207:Cytokines / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005822:Genetic Vectors; D005838:Genotype; D049452:Green Fluorescent Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D008533:Megakaryocytes / Q000378:metabolism / Q000473:pathology*; D008565:Membrane Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008894:Milk Proteins / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008938:Mitosis; D015233:Models, Statistical; D009363:Neoplasm Proteins / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D040201:Platelet Membrane Glycoprotein IIb / Q000096:biosynthesis; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D018121:Receptors, Cytokine / Q000378:metabolism*; D053628:Receptors, Thrombopoietin; D012190:Retroviridae / Q000235:genetics; D050796:STAT3 Transcription Factor; D050799:STAT5 Transcription Factor; D015398:Signal Transduction; D013154:Spleen / Q000166:cytology; D013926:Thrombopoietin / Q000378:metabolism*; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D016207:Cytokines; D004268:DNA-Binding Proteins; D049452:Green Fluorescent Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D008565:Membrane Proteins; D008894:Milk Proteins; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D009363:Neoplasm Proteins; D019869:Phosphatidylinositol 3-Kinases; D040201:Platelet Membrane Glycoprotein IIb; D017346:Protein Serine-Threonine Kinases; D011506:Proteins; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D018121:Receptors, Cytokine; D053628:Receptors, Thrombopoietin; D050796:STAT3 Transcription Factor; D050799:STAT5 Transcription Factor; D013926:Thrombopoietin; D015534:Trans-Activators; C490777:Lnk protein, mouse; C490778:SH2B3 protein, human; C494087:Stat3 protein, mouse; C082218:MPL protein, human