Understanding the receptor activation mechanism is essential for the rational design of pharmacologically active ligand molecules. However, the activation mechanism of most cytokine receptors remains still unclear, and while agonism and antagonism have been described for ligand-mimetic peptides, there has been no report of inverse agonism that has been characterized for G protein-coupled receptors (GPCRs). To explore the activation mechanism of cytokine receptors, here we tried to investigate how agonism and antagonism could be altered by randomizing antibody variable region of an antibody/cytokine receptor chimera recognizing hen egg lysozyme (HEL) as an agonist. Based on our previous finding that the co-expression of V(H)-gp130 and V(L)-erythropoietin receptor (EpoR) chimeras transduced strict and efficient HEL-dependent cell growth signal, a V(H)-gp130 library encoding four randomized CDR2 residues was retrovirally infected to IL-3-dependent Ba/F3 cells already transfected with V(L)-EpoR. The selection without IL-3 resulted in a clonal expansion of the transduced cells, and interestingly some of which showed HEL dose-dependent growth suppression. Our results clearly indicate that agonism and antagonism of the antibody/cytokine receptor chimera can be readily switched by a subtle modification of the ligand binding domain as well as that of GPCRs, also implying the existence of inverse agonism in cytokine receptor superfamily.