The mammalian multisubunit photoreceptor cGMP phosphodiesterase PDE alpha beta gamma 2 (PDE6 family) is a peripherally membrane-associated enzyme. A novel subunit, termed PDE delta (HGMW-approved symbol, PDE6D; MW 17 kDa), is able to detach PDE partially from bovine rod outer segment membranes under physiological conditions. Cloning of human and mouse PDE delta cDNAs revealed that PDE delta is a nearly perfectly conserved polypeptide of 150 amino acids that shows partial sequence homology to photoreceptor RG4 of unknown function. Multiple-species Southern blot analysis demonstrates that the PDE delta gene has been well conserved during evolution and is detectable at high stringency in invertebrates. The human and mouse genes are contained in less than 8 kb of genomic DNA and consist of four exons and three introns (0.7-4 kb in human, 0.7-2.2 kb in mouse). The PDE delta gene structure is identical to that of the C27H5.1 gene identified in the eyeless nematode Caenorhabditis elegans. The human PDE delta gene (locus designation PDE6D) was localized to the long arm of chromosome 2 (2q35-q36) by fluorescence in situ hybridization. By synteny, the mouse PDE delta gene is predicted to reside on chromosome 1.