Intercellular adhesion molecule-2 (ICAM-2), a cell surface glycoprotein, is a second counter-receptor for lymphocyte function-associated Ag-1 (LFA-1). We report here the isolation and characterization of the cDNA and the gene that encode murine ICAM-2 (Accession numbers X65493 and X65490, respectively). The deduced sequence of the cDNA has 60% amino acid identity with its human counterpart and has the same expression pattern in cells and tissues. Furthermore, COS cells transfected with mouse ICAM-2 complementary and genomic DNA bind to purified human LFA-1, demonstrating the conservation of the function of ICAM-2 as a ligand for LFA-1 and conservation across species of sequences that are critical for binding to human LFA-1. COS cells transfected with the ICAM-2 cDNA do not react with mAb PA3, previously suggested to define ICAM-2 in the mouse. The mouse ICAM-2 gene was isolated and its structural organization determined. The gene is present in a single copy in the mouse genome and contains four exons spanning about 5.0 kb of DNA. The exon/intron architecture correlates to the structural domains of the protein and resembles that of other Ig superfamily members. The gene for ICAM-2, which is constitutively expressed in endothelial cells, has several conserved sequence motifs in its promoter region, including a direct repeat, and lacks transcription factor-binding sites present in the ICAM-1 gene, which is inducible in endothelial cells.