Kakihana Kazuhiko, Yamamoto Masahide, Iiyama Mitsuko, Miura Osamu
Department of Hematology, Graduate School of Medicine and Dentistry, Tokyo Medical and Dental University, Tokyo 113-8519, Japan.
Biochem Biophys Res Commun. 2005 Sep 23;335(2):424-31. doi: 10.1016/j.bbrc.2005.07.095.
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.
促红细胞生成素受体（EpoR）的刺激会导致细胞内Ca2+水平短暂升高，以及Jak2酪氨酸激酶的激活，从而刺激各种下游信号通路。在此，我们证明通用的Ca2+受体钙调蛋白（CaM）在体外以Ca2+依赖的方式与EpoR结合。使用造血细胞中各种EpoR突变体的结合研究表明，CaM与EpoR的膜近端65个氨基酸的胞质区域（氨基酸258 - 312）结合，该区域对于Jak2介导的EpoR信号激活至关重要。结构不相关的CaM拮抗剂W - 13和CMZ抑制了Jak2介导的EpoR信号通路的激活，而W - 13类似物W - 12则未显示出任何显著的抑制作用。此外，CaM的过表达增强了Epo诱导的EpoR酪氨酸磷酸化。W - 13而非W - 12也抑制了Epo诱导的增殖和存活。总之，这些结果表明CaM与膜近端EpoR胞质区域结合，并在Jak2介导的EpoR信号激活中起重要作用。
