Evidence for specific receptors for interleukin 3 on lymphokine-dependent cell lines established from long-term bone...

Utilizing 125I-labelled IL 3, an assay for specific IL 3 receptors on continuous cell lines was developed. The 32D-cl 23 and FDC-P1 cell lines examined were originally derived from long-term cultures of murine bone marrow supplemented with WEHI-3-conditioned media. Both of these cell lines have been shown to be absolutely dependent on IL 3, a component of WEHI-3-conditioned media, for growth in vitro. The binding of radiolabeled IL 3 to these cells was found to be specific, saturable, reversible, and time and temperature dependent. The specificity of IL 3 binding was demonstrated by the failure of a number of different lymphokines, hormones, and proteins to compete for the binding of radiolabeled IL 3. The IL 3-dependent cell lines uniquely express detectable receptors, whereas other cell lines of various types and origin known to be IL 3 independent had no detectable specific binding of IL 3. A Kd of 5.4 X 10(-11) M and a maximum binding capacity of 6.25 fmol per 1 X 10(6) cells was calculated for 32D-cl 23 from equilibrium binding studies. A similar analysis for FDC-P1 yielded a Kd of 1.7 X 10(-11) and a maximum binding capacity of 1.2 fmol per 1 X 10(6) cells. The data are consistent with the presence of 4000 to 5000 specific high-affinity binding sites/cell on 32D-cl 23, whereas FDC-P1 has 1500 to 2500 specific sites/cell. The Kd calculated for the FDC-P1 cell line from kinetic binding data was 2.5 X 10(-11) M, which agreed well with the Kd of 1.7 X 10(-11) M calculated from equilibrium data.