title: Calmodulin physically interacts with the erythropoietin receptor and enhances Jak2-mediated signaling.
abstract: Stimulation of the erythropoietin receptor (EpoR) induces a transient increase in intracellular Ca2+ level as well as activation of the Jak2 tyrosine kinase to stimulate various downstream signaling pathways. Here, we demonstrate that the universal Ca2+ receptor calmodulin (CaM) binds EpoR in a Ca2+-dependent manner in vitro. Binding studies using various EpoR mutants in hematopoietic cells showed that CaM binds the membrane-proximal 65-amino-acid cytoplasmic region (amino acids 258-312) of EpoR that is critical for activation of Jak2-mediated EpoR signaling. Structurally unrelated CaM antagonists, W-13 and CMZ, inhibited activation of Jak2-mediated EpoR signaling pathways, whereas W-12, a W-13 analog, did not show any significant inhibitory effect. Moreover, overexpression of CaM augmented Epo-induced tyrosine phosphorylation of the EpoR. W-13, but not W-12, also inhibited Epo-induced proliferation and survival. Together, these results indicate that CaM binds to the membrane-proximal EpoR cytoplasmic region and plays an essential role in activation of Jak2-mediated EpoR signaling.
authors.affiliation: Department of Hematology, Graduate School of Medicine and Dentistry, Tokyo Medical and Dental University, Tokyo 113-8519, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Kakihana
authors.forename: Masahide
authors.initials: M
authors.lastname: Yamamoto
authors.forename: Mitsuko
authors.initials: M
authors.lastname: Iiyama
authors.forename: Osamu
authors.initials: O
authors.lastname: Miura
mesh_terms: D000818:Animals; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000737:chemistry*; D002460:Cell Line; D049109:Cell Proliferation; D002470:Cell Survival; D002845:Chromatography; D017077:Culture Media, Conditioned / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D047468:Immunoprecipitation; D053614:Janus Kinase 2; D051379:Mice; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry*; D011518:Proto-Oncogene Proteins / Q000737:chemistry*; D017467:Receptors, Erythropoietin / Q000737:chemistry*; D012685:Sepharose / Q000737:chemistry; D015398:Signal Transduction; D013449:Sulfonamides / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry; D015854:Up-Regulation
chemical_list: D002147:Calmodulin; D017077:Culture Media, Conditioned; D011518:Proto-Oncogene Proteins; D017467:Receptors, Erythropoietin; D013449:Sulfonamides; C040184:W 12; D014443:Tyrosine; C033772:N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide; D012685:Sepharose; D011505:Protein-Tyrosine Kinases; C507924:JAK2 protein, human; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2; D002118:Calcium