ISOLATION, CLONING, AND CHARACTERIZATION OF A TRP HOMOLOG FROM SQUID (LOLIGO-FORBESI) PHOTORECEPTOR-MEMBRANES

The invertebrate phototransduction system is a valuable model of the u biquitous inositol lipid signalling system. Taking advantage of the ab ility to obtain relatively large amounts of retinal material from the cephalopod eye, partial protein sequence data were obtained for a 92-k Da component isolated from a detergent-insensitive cytoskeletal fracti on of a squid retinal microvillar membrane preparation. Degenerate oli gonucleotides, designed on the basis of these sequence data, were used to isolate a full-length cDNA, encoding the 92-kDa component, using b oth cDNA library screening and 5'-rapid amplification of cDNA ends (5' -RACE) techniques. Comparison of the amino acid sequence encoded by th is cDNA with entries in the OWL composite protein sequence database re veals greatest sequence similarity with the products of the Drosophila trp and trpl genes. Greatest variation from the Drosophila Trp protei n is seen in the carboxyl-terminal region, which is considerably trunc ated in the squid protein and which accounts for most of the substanti al difference in molecular weight seen between these proteins. This va riation may be significant as the carboxyl-terminal domain has been sh own to be in the regulation of several ligand-gated channels. The carb oxyl-terminal domain has been expressed and shown to interact with cal modulin in a calcium-dependent fashion, thereby supporting this hypoth esis. The likely occurrence of other homologues in a variety of system s suggests that this is a novel and important family of regulated ion channels involved in calcium signalling.