The human cone transducin alpha-subunit (GNAT2) gene has been completely characterized. The human GNAT2 transcription unit is 9967-bp in length and consists of eight exons with seven introns. The eight exons are identical to the reported cDNA sequence (Lerea et al., 1989). Northern blot analysis of RNA from human retinas and a retinoblastoma cell line, WERI-RB1, reveals a 1.7-kb transcript for GNAT2. Multiple transcription initiation sites were mapped for human retina and WERI-RB1 RNA by primer extension and S1 nuclease protection assays. This gene has seven initiation sites spanning 31 bp. The sequence upstream of the GNAT2 gene shows a TATA box consensus sequence at -29, a CCAAT box consensus sequence at -58 (reverse orientation), and a sequence (CCATAT) similar to the CCAAT box consensus at -76. The GNAT2 upstream sequence shows no significant identity with the upstream region of the human rod transducin alpha-subunit gene (GNAT1) or with the upstream regions of the color visual pigment genes, indicating that the expression of GNAT2 may be regulated differently than these other rod- and cone-specific proteins.