THE BETA-CHAIN OF THE INTERLEUKIN-3 RECEPTOR FUNCTIONALLY ASSOCIATES WITH THE ERYTHROPOIETIN RECEPTOR

Interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating fac tor (GM-CSF) receptors share a common beta chain (beta(c)) and both cy tokines enhance erythropoietin (Epo)-dependent in vitro erythropoiesis by primary hematopoietic progenitors and factor-dependent cells. Thes e data suggest that the Epo receptor and beta(c) may functionally inte ract. To determine whether such interactions can be documented, we stu died a murine factor-dependent cell line (Ba/F3), which endogenously e xpresses IL-3R. First, Ba/F3 cells were transfected with murine EpoR, making them responsive to both IL-3 and Epo. Next, the EpoR expressing cells were transfected with murine beta(c). This resulted in an enhan ced sensitivity of these cells to Epo, which was especially pronounced at low Epo concentrations, Ba/F3-EpoR were then treated with antisens e oligodeoxynucleotides to the murine beta. Control sense and nonsense had no effect on Epo-dependent growth, but the antisense markedly and specifically inhibited Epo-dependent growth. In contrast, the antisen se did not affect beta-globin message levels (another Epo-responsive e ffect in these cells) detectable by Northern blot. Finally, Western bl ot analysis of proteins immunoprecipitated from cells expressing both receptors with antibody against beta and blotted with antibody against EpoR, or immunoprecipitated with antibody against EpoR and blotted wi th antibody against beta, showed that EpoR and beta coimmunoprecipitat e. These data show that the beta chain functionally and physically ass ociates with the EpoR. This suggests that these cytokine receptors exi st as a large supercomplex and offers the first molecular explanation for the synergistic effects of IL-3 and GM-CSF with Epo during erythro poiesis. (C) 1997 by The American Society of Hematology.