4‐1BB T‐cell antigen binds to mature B cells and macrophages, and costimulates anti‐μ‐primed splenic B cells

Abstract 4‐1BB is expressed on activated murine T cells and may function as an accessory signaling molecule during T‐cell activation. To identify putative 4‐1BB ligands, a fusion protein consisting of the extracellular domain of 4‐1BB fused to human placental alkaline phosphatase (4‐1BB‐AP) was constructed. Alkaline phosphatase activity could then be used as an indicator of the relative amount of bound 4‐1BB. These studies indicated that 4‐1BB‐AP specifically bound to the surface of various mature B and macrophage cell lines. 4‐1BB‐AP bound at low levels to T cell lines (non‐activated and anti‐CD3‐activated), pre‐B‐cell lines, and an immature macrophage cell line. 4‐1BB‐AP did not bind to a glial tumor cell line, HeLa cells, or COS cells. In addition, 4‐1BB‐AP bound at higher levels to F(ab′) 2 anti‐μ‐activated primary B cells compared to anti‐CD3‐activated primary T cells. Scatchard analysis indicated that the A20 B cell lymphoma expressed 3680 binding sites per cell with a K d of 1.86 n M . Affinity cross‐linking studies demonstrated that a major cell surface species of 120 kDa bound to 4‐1BB‐AP 4‐1BB‐AP also bound to a minor species of approximately 60 kDa. The addition of paraformaldehyde‐fixed SF21 cells expressing recombinant 4‐1BB synergized with F(ab′) 2 anti‐μ in inducing splenic B cell proliferation suggesting that 4‐1BB may function as a regulator of B cell growth.