IDENTIFICATION OF FUNCTIONAL DOMAINS IN MURINE GRANULOCYTE-MACROPHAGECOLONY-STIMULATING FACTOR USING MONOCLONAL-ANTIBODIES TO SYNTHETIC PEPTIDES

Granulocyte-macrophage (GM)-CSF is an important hematopoietic cytokine that regulates proliferation and differentiation of macrophages, neut rophils, and eosinophils. In this study, we generated mAb to five synt hetic peptides that correspond to regions along the murine GM-CSF mole cule. The ability of anti-peptide mAb to bind to and inhibit biologic activity of murine (m) GM-CSF was determined. mAb with the highest neu tralization titers were derived from mice immunized with peptide II, w hich correspond to amino acids 27 to 38 of mGM-CSF. Immunochemical stu dies showed that peptide II specifically blocked binding of anti-pepti de II mAb to GM-CSF. mAb to two other peptides in the N-terminal half corresponding to residues 7 to 17 and 47 to 58, respectively, of mGM-C SF also inhibited GM-CSF-dependent proliferation and differentiation o f murine bone marrow precursors for macrophages and granulocytes. Anti -peptide mAb also inhibited growth of a murine hematopoietic cell line FDCP1 and a murine T cell line HT-2, which was shown to be dependent on GM-CSF for growth in vitro. Biologic activity of both natural and r ecombinant mGM-CSF was neutralized by anti-peptide mAb. These findings indicate that epitopes in the N-terminal region of mGM-CSF are import ant for biologic activity, and the epitope defined by peptide II (resi dues 27 to 38) lies within a particularly important functional domain of the mGM-CSF molecule.