GUANYLATE-CYCLASE-INHIBITORY PROTEIN IS A FROG RETINAL CA2-BINDING PROTEIN RELATED TO MAMMALIAN GUANYLATE-CYCLASE-ACTIVATING PROTEINS()

Two guanylate-cyclase-activating proteins (GCAP) encoded by a tail-to- tail gene array have been characterized in the mammalian retina. Using frog retina as a model, we obtained evidence for the presence of a ph otoreceptor Ca2+-binding protein closely related to GCAP. This protein (206 amino acids) does not stimulate guanylate cyclase (GC) in low [C a2+], but inhibits GC in high [Ca2+], and is therefore termed guanylat e-cyclase-inhibitory protein (GCIP). Sequence analysis indicates that GCIP and GCAP1 and GCAP2 have diverged substantially, but conserved do mains present in all vertebrate GCAP are present in GCIP. Moreover, pa rtial characterization of the GCIP gene showed that the positions of t wo introns in the GCIP gene are identical to positions of correspondin g introns of the mammalian GCAP gene array. As to the major difference s between GCIP and GCAP, the fourth EF hand Ca2+-binding motif of GCIP is disabled for Ca2+ binding, and GCIP does not stimulate GC. Monoclo nal and polyclonal antibodies raised against recombinant GCIP identifi ed high levels of GCIP in the inner segments, somata and synaptic term inals of frog cone photoreceptors. The results suggest that GCIP is a Ca2+-binding protein of the GCAP/recoverin subfamily. Its localization in frog cones closely resembles that of GC in mammalian cones. GCIP i nhibits GC at high free [Ca2+], competing with GCAP1 and GCAP2 for GC regulatory sites.