Cyclic nucleotide-gated channels have been proposed to mediate the electrical response to light in the ventral photoreceptor cells of the horseshoe crab, Limulus polyphemus. However, a cyclic nucleotide-gated channel has not been identified from Limulus. We have cloned a putative full-length cyclic nucleotide-gated channel cDNA by screening cDNA libraries constructed from Limulus brain using a probe developed from Limulus ventral eye nerves. The putative full-length cDNA was derived from two overlapping partial cDNA clones. The open reading frame encodes 905 amino acids; the sequence shows 44% identity to that of the ␣ subunit of the bovine rod cyclic GMP-gated channel over the region containing the transmembrane domains and the cyclic nucleotide binding domain. This Limulus channel has a novel C-terminal region of ϳ200 amino acids, containing three putative Src homology domain 3 binding motifs and a putative coiled-coil domain. The possibility that this cloned channel is the same as that detected previously in excised patches from the photoreceptive membrane of Limulus ventral photoreceptors is discussed in terms of its sequence and its expression in the ventral eye nerves. Key Words: Cyclic nucleotide-gated channels-Limulus polyphemus. J. Neurochem. 72, 461-471 (1999).Cyclic GMP (cGMP) is the intracellular second messenger for phototransduction in vertebrate photoreceptors, where it directly activates cyclic nucleotide-gated (CNG) channels in the plasma membrane of the outer segment (Fesenko et al., 1985;Kaupp et al., 1989). It has also been proposed as an intracellular second messenger that activates ion channels in horseshoe crab (Limulus polyphemus) photoreceptors. Injection of cGMP into Limulus ventral photoreceptor cells partially depolarizes the photoreceptors, and channels in excised patches from the photoreceptive membranes can be activated directly by cGMP (Johnson et al., 1986;Bacigalupo et al., 1991).As a first step toward resolving the role of CNG channels in Limulus phototransduction, we have cloned a putative CNG channel cDNA, Lcng1, from Limulus. This putative Limulus CNG channel subunit (LCNG1) shares significant homology with the ␣ subunit of the bovine rod cGMP-gated channel. The sequence also reveals a novel long C terminus. RT-PCR experiments indicate that the Lcng1 gene is expressed in both Limulus brain and ventral eye nerve. MATERIALS AND METHODS Materials and tissue preparationHorseshoe crabs were either purchased from the Martin Fish Co. (Ocean City, MD, U.S.A.) or the Marine Biological Laboratory (Woods Hole, MA, U.S.A.) or caught in the Indian River of Florida. Dissected, live tissues were either used immediately or frozen in liquid nitrogen and stored at Ϫ80°C.Ventral eye nerves were cut at least several millimeters away from the brain to avoid brain tissue contamination. In desheathing nerves, both the sheath around the nerve and the modified patch of cuticle at the eye terminal were removed. Brains were dissected free of surrounding nerves. The coax extensor muscle of th...

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