IDENTIFICATION AND CHARACTERIZATION OF AN ALTERNATIVE CYTOTOXIC T-LYMPHOCYTE-ASSOCIATED PROTEIN-4 BINDING MOLECULE ON B-CELLS

To determine whether alternative cytotoxic T lymphocyte-associated pro tein 4 (CTLA4) binding proteins exist on B cells, we constructed (i) m CTLA4hIgG consisting of the extracellular region of a mouse CTLA4 mole cule and the Fc portion of a human IgG1 molecule and (ii) PYAAhIgG, a mutant mCTLA4hIgG, having two amino acid substitutions on the conserve d MYPPPY motif in the complementarity-determining region 3-like region and lacking detectable binding to both B7-1 and B7-2 molecules. Using these fusion proteins (mCTLA4hIgG and PYAAhIgG), we demonstrated that a mouse immature B-cell line, WEH1231 cells, expressed alternative CT LA4 binding molecules (ACBMs) that were distinct from both B7-1 and B7 -2. ACBMs were 130-kDa disulfide-linked proteins. More importantly, AC BMs were able to provide costimulatory signal for T-cell proliferation in the presence of anti-CD3 monoclonal antibodies. In addition, we de monstrated that more than 20% of B220(+) cells obtained from normal mo use spleen expressed ACBMs.