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 hamsters 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 peaks 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 CD8 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 reducing conditions. Deglycosylation of the reduced H4 by N‐glycanase gave rise to a single band of about 21 kDa, suggesting that the two chains may be differentially glycosylated forms of the same protein. The H4 expression pattern and biochemical features, together with cross‐blocking, co‐capping, co‐modulation, and immunoprecipitation preclearing experiments 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 in vitro kinase assay, H4 was found to co‐precipitate a tyrosine kinase activity that phosphorylated substrates of about 29 and 25 kDa. Co‐modulation and co‐capping experiments showed that H4 is physically associated with the CD3/T cell receptor. These data suggest that H4 may function as a T cell‐specific co‐stimulatory molecule and play a role in the T cell response when the activation stimulus is limited either because the antigen is only available in low concentration or has a low agonistic activity.

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