K. Ishiguro et Ac. Sartorelli, CHARACTERIZATION OF THE DOMAINS OF THE ERYTHROPOIETIN RECEPTOR NECESSARY FOR INDUCTION OF CELL-GROWTH AND DIFFERENTIATION, Cytokine (Philadelphia, Pa. Print), 10(8), 1998, pp. 564-577
To define the cytoplasmic region(s) of the erythropoietin receptor (Ep
oR) necessary for promotion of growth and induction of differentiation
, mutated EpoR cDNAs containing truncations and conversions of tyrosin
e residues to phenylalanines were generated. Constructs mere introduce
d into IL-3 dependent Ba/F3 cells by electroporation, and individual t
ransfectants were propagated in methylcellulose-containing medium in t
he presence of erythropoietin (Epo), Truncated receptor at amino acid
350 was partially deficient in promoting cellular growth and the recep
tor lacking the box 2 region was not capable of inducing cellular grow
th. When a large number of cells,were screened in Epo-containing liqui
d medium, clones arose in which Epo-dependent growth was due to activa
tion of the endogenous EpoR gene. Analysis of the degree of Epo-depend
ent differentiation of the transfectants, based on the steady-state le
vels of beta(major)-globin mRNA, showed that the carboxyl terminal 133
amino acids and tyrosyl residues located at positions 429/431 and 460
/464 were not necessary for the induction of differentiation. Examinat
ion of clones from diverse origins demonstrated that an inverse relati
onship existed between the rate of Epo-induced cellular replication an
d the degree of Epo-induced differentiation. (C) 1998 Academic Press.