Src transduces erythropoietin-induced differentiation signals through phosphatidylinositol 3-kinase.
In this study, we examined the molecular mechanism of erythropoietin-initiated signal transduction of erythroid differentiation through Src and phosphatidylinositol 3-kinase (PI3-kinase). Antisense oligonucleotides against src but not lyn inhibited the formation of erythropoietin-dependent colonies derived from human bone marrow cells and erythropoietin-induced differentiation of K562 human erythroleukaemia cells. Antisense p85alpha oligonucleotide or LY294002, a selective inhibitor of PI3-kinase, independently inhibited the formation of erythropoietin-dependent colonies. In K562 cells, Src associated with PI3-kinase in response to erythropoietin. Antisense src RNA expression in K562 cells inhibited the erythropoietin-induced activation of PI3-kinase and its association with erythropoietin receptor. PP1, a selective inhibitor of the Src family, reduced erythropoietin-induced tyrosine phosphorylation of erythropoietin receptor and its association with PI3-kinase in F-36P human erythroleukaemia cells. The coexpression experiments and in vitro kinase assay further demonstrated that Src directly tyrosine-phosphorylated erythropoietin receptor, and associated with PI3-kinase. In vitro binding experiments proved that glutathione S-transferase-p85alpha N- or C-terminal SH2 domains independently bound to erythropoietin receptor, which was tyrosine-phosphorylated by Src. Taken together, Src transduces the erythropoietin-induced erythroid differentiation signals by regulating PI3-kinase activity.
Kubota
Y
Y
Department of Transfusion Medicine, School of Medicine and Environmental Health Sciences, Kagawa Medical University, Kagawa 761-0793, Japan. ketsuken@kms.ac.jp
Tanaka
T
T
Kitanaka
A
A
Ohnishi
H
H
Okutani
Y
Y
Waki
M
M
Ishida
T
T
Kamano
H
H
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects; D019556:COS Cells; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002522:Chlorocebus aethiops; D002867:Chromones / Q000494:pharmacology; D003114:Colony-Forming Units Assay; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015672:Erythroid Precursor Cells / Q000166:cytology / Q000187:drug effects*; D004921:Erythropoietin / Q000494:pharmacology / Q000502:physiology*; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects; D016391:Genes, src; D006801:Humans; D020014:K562 Cells / Q000187:drug effects; D046911:Macromolecular Substances; D009025:Morpholines / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000502:physiology; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000502:physiology*; D017467:Receptors, Erythropoietin / Q000187:drug effects* / Q000502:physiology; D011994:Recombinant Proteins; D015398:Signal Transduction / Q000502:physiology*; D014162:Transfection; D000077191:Wortmannin; D018909:src Homology Domains; D019061:src-Family Kinases / Q000502:physiology
D000730:Androstadienes; D002867:Chromones; D004791:Enzyme Inhibitors; D004921:Erythropoietin; D046911:Macromolecular Substances; D009025:Morpholines; D009363:Neoplasm Proteins; D016376:Oligonucleotides, Antisense; D019869:Phosphatidylinositol 3-Kinases; D016392:Proto-Oncogene Proteins pp60(c-src); D017467:Receptors, Erythropoietin; D011994:Recombinant Proteins; D000077191:Wortmannin; D019061:src-Family Kinases; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C060902:lyn protein-tyrosine kinase