BT-IgSF is a newly identified cell surface glycoprotein belonging to the immunoglobulin superfamily (IgSF). We have previously shown that the expression of the BT-IgSF gene was highly restricted to brain and testis, and its transcript was detected in both neurons and glial cells. In this study, to explore its function, we generated cells overexpressing BT-IgSF proteins and analyzed their phenotypes. We found that the constitutive expression of BT-IgSF in the myeloid leukemia cell line TF-1-fms did not alter the growth rates, but caused the formation of large cell aggregates. The cell aggregates were also observed with mutant BT-IgSF lacking its cytoplasmic tail, the amino acid sequences of which were highly conserved among the BT-IgSF subgroup proteins. The neutralizing antibody to beta(1) integrin did not diminish the cell aggregate formation. These results indicate that BT-IgSF functions as a cell adhesion molecule, that its cytoplasmic tail is not essential for the function, and that beta(1) integrin is not involved in the function. We confirmed the cell adhesion function using NIH/3T3 fibroblastic cells expressing BT-IgSF in an inducible system. Flow cytometric analyses with the cells demonstrated that the cell aggregation mediated by BT-IgSF was through homophilic molecular interaction, and in a Ca(2+)/Mg(2+)-independent manner. Coupled with its restricted pattern of the expression, the cell adhesion-inducing function of BT-IgSF suggests a role of the cell surface molecule in the development/function of the central nervous system and spermatogenesis.