Amino acid residues 268-276 of the erythropoietin receptor contain an endocytosis motif and are required for erythropoietin-mediated proliferation.
Erythropoietin (EPO) promotes viability, proliferation and differentiation of mammalian erythroid progenitor cells via its specific cell surface receptor (EPO-R). We have previously shown that truncated EPO-Rs containing 267 amino acids or less were defective in internalization of (125)I-EPO, whereas internalization via a receptor derivative containing 276 amino acids was unaffected, thus directing focus to the nine amino acid residues FEGLFTTHK at positions 268-276 [Levin, Cohen, Supino, Yoshimura, Watowich, Neumann, FEBS Lett. 427 (1998) 164-170]. Here, a panel of EPO-R mutants was generated to determine the role of these residues in EPO endocytosis, down regulation of cell surface receptors and EPO-mediated signaling. While linking amino acid residues 268-276 to a truncated EPO-R (Delta+9 EPO-R) conferred both ligand uptake and ligand-independent down regulation of the respective receptor from the cell surface, Phe 272 was crucial for EPO endocytosis but not for ligand-independent down regulation. Additional receptor motifs probably play a role in EPO endocytosis and receptor down-regulation, as these processes were not adversely impaired in Delta268-276 EPO-R. A central role of residues 268-276, in particular Phe, was demonstrated by the inability of Delta268-276 and F268,272A EPO-Rs to support EPO-mediated signal transduction.
Flint-Ashtamker
Galit
G
Department of Cell Biology and Histology, Sackler Faculty of Medicine Tel-Aviv University, 69978, Ramat Aviv, Israel.
Eisen-Lev
Ronit
R
Cohen
Jacob
J
Jun-shen Huang
Lily
L
Neumann
Drorit
D
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002455:Cell Division; D002460:Cell Line; D015536:Down-Regulation; D004705:Endocytosis; D004921:Erythropoietin / Q000378:metabolism / Q000494:pharmacology*; D009154:Mutation; D017467:Receptors, Erythropoietin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
D017467:Receptors, Erythropoietin; D004921:Erythropoietin