FUNCTIONAL INTERACTION OF ERYTHROPOIETIN AND STEM-CELL FACTOR RECEPTORS IS ESSENTIAL FOR ERYTHROID COLONY FORMATION

Production of mature erythrocytes requires multiple growth factors, bu t we do not know how their actions are coordinated, Here we show that erythroid progenitors from erythropoietin receptor (Epo-R)(-/-) fetal livers, infected in vitro with a retrovirus expressing the wild-type E po-R, require addition of both Epo and stem cell factor (SCF) to form colony-forming unit erythroid (CFU-E) colonies. Thus, a functional int eraction between KIT and the Epo-R, similar to what we reported in cul tured cells, is essential for the function of CFU-E progenitors, In co ntrast, CFU-E colony formation in vitro by normal fetal liver progenit ors requires only Epo; the essential interaction between activated KIT and the Epo-R must have occurred in vivo before or at the CFU-E proge nitor stage, Using truncated dominant-negative mutant Epo-Rs, me show that KIT does not activate the Epo-R by inducing its dimerization, but presumably does so by phosphorylating tyrosine residue(s) in its cyto solic domain, By expressing mutant Epo-Rs containing only one of eight cytosolic tyrosines, we show that either tyrosine residue Y464 or Y47 9 suffices for Epo-dependent cell proliferation, However, only Epo-R F 7Y479 is capable of supporting erythroid colony formation when express ed in Epo-R(-/-) fetal liver cells, indicating that Y464 either cannot send a differentiation signal or fails to respond to SCF/KIT activati on, This work employs a novel experimental system to study the functio n of growth factors and their receptors in normal hematopoiesis.