The gene for one of the activating members of the paired Ig-like receptor family, Pira6, was isolated from a genomic library and sequenced. The first of 9 exons in the approximately 8.2 kb Pira6 gene encodes the 5' untranslated region, the translation initiation site, and approximately half of the signal sequence. The second exon encodes the rest of the signal sequence, exons 3-8 each encode a single Ig-like extracellular domain, and exon 9 encodes the transmembrane region, cytoplasmic tail and 3' UTR with four polyadenylation signals and six mRNA instability sequences. A soluble form of PIR-A6 may be generated by alternative splicing. The exonic sequences account for approximately 42% of the Pira6 gene and approximately 34% for the single inhibitory Pirb gene, thus defining Pira and Pirb as genes with relatively short intronic sequences. Extensive sequence homology was found between Pira6 and Pirb from approximately 2 kb upstream of the ATG initiation site to the beginning of intron 8. The Pir genes appear to be distributed in three regions of the proximal end of chromosome 7 based on the present data and an analysis of currently available mouse genomic sequence databases. One region contains a single Pir gene which is almost identical to Pira6, and the other two contain multiple Pir genes in opposite transcriptional orientations. Potential binding sites for hemopoiesis-specific and ubiquitous transcription factors were identified upstream of the Pira6 transcription start sites that reside within the initiator consensus sequence motif. These results provide important clues to the coordinate regulation observed for PIR-A and PIR-B expression during hematopoiesis.