Biochemical comparisons of a new yS T cell lineage-specific marker (GD3.5Ag) with the WC1 family

In this report, we describe GD3.5, a new lineage-specific γδ T cell marker that is distinct from TCR, and WC1. FACS analysis indicated that GD3.5Ag is expressed on approximately 90% of the peripheral blood γδ T cell population and GD3.5 specifically stained γδ T cells and not αβ T cells or B-cells. Nonreducing Western blot analysis and immunoprecipitation experiments revealed a single 220-240kD glycoprotein recognized by GD3.5 compared to two IL-A29 reactive bands at 200kD and 300kD. Cross-immunoprecipitation experiments demonstrated that GD3.5 antigen and WC1 could be immunoprecipitated from lysates cleared of IL-A29/WC1 and GD3.5Ab/GD3.5Ag complexes, respectively. Digestion of WC1 and GD3.5 antigen with V-8 protease resulted in digestion profiles that clearly distinguished the glycoproteins. Additionally, GD3.5 antigen and WC1 possess disparate sensitivity to PNGase F, O-glycoprotease, and neuraminidase, indicating differences in N- and O-linked sugars and the presence of sialic acid residues. Lastly, surface expression of GD3.5 antigen, but not WC1, was reduced by very low-dose chymotrypsin and ConA treatment. Therefore, our data suggest that GD3.5 antigen is a previously uncharacterized, lineage-specific 78 T cell antigen. Furthermore, we show that GD3.5 and WC1 are sialomucins, which provides important clues to their function. Additionally, preliminary data suggest that GD3.5 antigen may play a role in γδ T cell adhesion to P-selectin.