CHARACTERIZATION OF H4 - A MOUSE T-LYMPHOCYTE ACTIVATION MOLECULE FUNCTIONALLY ASSOCIATED WITH THE CD3 T CELL-RECEPTOR/

The monoclonal antibody C398.4A was produced by immunizing Armenian ha msters with the mouse T cell clone D10.G4.1. It recognizes a molecule selectively expressed by activated mouse T cells and was named H4. H4 is expressed on the T cell surface about 24 h after activation and pea ks at day 7. By contrast, it is not expressed by resting or activated B cells, macrophages, or fibroblasts. It is also expressed by CD4 or C D8 single-positive mature thymocytes. Immunoprecipitation showed that H4 is a disulfide-linked dimer, precipitating as a broad band at about 50-65 kDa under nonreducing conditions and at 25 and 29 kDa under red ucing conditions. Deglycosylation of the reduced H4 by N-glycanase gav e rise to a single band of about 21 kDa, suggesting that the two chain s may be differentially glycosylated forms of the same protein. The H4 expression pattern and biochemical features, together with cross-bloc king, co-capping, co-modulation, and immunoprecipitation preclearing e xperiments showed that H4 is different from other known co-stimulatory molecules such as CD69, CD2, Ly-6, CD25, OX-40, Mac-1 and LFA-1. By i n vitro kinase assay, H4 was found to co-precipitate a tyrosine kinase activity that phosphorylated substrates of about 29 and 25 kDa. Co-mo dulation and co-capping experiments showed that H4 is physically assoc iated with the CD3/T cell receptor. These data suggest that H4 may fun ction as a T cell-specific co-stimulatory molecule and play a role in the T cell response when the activation stimulus is limited either bec ause the antigen is only available in low concentration or has a low a gonistic activity.