Tyrosine phosphorylation of the erythropoietin receptor: role for differentiation and mitogenic signal transduction.
The erythropoietin (Epo) receptor belongs to the cytokine receptor superfamily. Although the cytokine receptors do not possess a tyrosine kinase consensus sequence in the intracellular domain, rapid stimulation of a tyrosine kinase activity occurs after activation by the ligand. We and others have shown that Epo induces the tyrosine phosphorylation of its cognate receptor as well as phosphorylation of other proteins. In this report, we examined the role of the receptor tyrosine residues in signal transduction. Eight tyrosine residues are located within the intracellular domain of the murine Epo receptor. A single tyrosine residue is present in the region previously shown to be sufficient for proliferative signal transduction. This tyrosine (Tyr 343) was mutated to phenylalanine. Moreover, mutant receptors were also generated with either a tyrosine residue or a phenylalanine residue at position 343 and with a COOH terminal truncation that removed the 7 other tyrosine residues. Expression vectors carrying these mutated receptors were transfected into the interleukin-3-dependent murine cell line Ba/F3. Epo-induced growth was sustained efficiently by all these receptors, although receptors without any tyrosine residues conferred a significantly reduced mitogenic activity. Moreover, all receptors were able to mediate Epo-dependant accumulation of beta-globin mRNA. The mutated receptors all induced the tyrosine phosphorylation of several cellular proteins after Epo stimulation. However, the truncated receptors induced the phosphorylation of a reduced number of proteins, suggesting that phosphorylated tyrosines of the receptor could have a role in the recruitment either of a tyrosine kinase or of tyrosine kinase substrate proteins. The receptors were all able to mediate Epo-induced activation of phosphatidylinositol 3-kinase, although truncated receptors no longer bound phosphatidylinositol 3-kinase.
Gobert
S
S
Institut Cochin de Génétique Moléculaire (ICGM), Université René Descartes, Paris, France.
Porteu
F
F
Pallu
S
S
Muller
O
O
Sabbah
M
M
Dusanter-Fourt
I
I
Courtois
G
G
Lacombe
C
C
Gisselbrecht
S
S
Mayeux
P
P
D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects; D004920:Erythropoiesis / Q000187:drug effects / Q000502:physiology*; D004921:Erythropoietin / Q000502:physiology*; D005786:Gene Expression Regulation / Q000187:drug effects; D005914:Globins / Q000096:biosynthesis / Q000235:genetics; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000378:metabolism; D007377:Interleukin-3 / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation / Q000187:drug effects; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D019000:Phosphotyrosine; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D017467:Receptors, Erythropoietin / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017384:Sequence Deletion; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D014162:Transfection; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism*
D007377:Interleukin-3; D017467:Receptors, Erythropoietin; D011993:Recombinant Fusion Proteins; D004921:Erythropoietin; D019000:Phosphotyrosine; D014443:Tyrosine; D005914:Globins; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D011505:Protein-Tyrosine Kinases