Cloning and characterization of rod opsin cDNA from the Old World monkey, Macaca fascicularis.

PURPOSE

Opsin sequences from different phyla have been compared to examine the structural and functional regions of this molecule and to assess the molecular evolution of genes in the animal kingdom. Of all the sequences reported, there is only one primate sequence (human) for rod opsin. To extend our knowledge of the primate family of opsins, the authors cloned and characterized the opsin cDNA for the Old World monkey Macaca fascicularis and determined its position in the phylogenetic tree of the opsin gene family.

METHODS

A fragment of the monkey opsin cDNA was generated using specific primers and the polymerase chain reaction. This fragment was then used as a probe to isolate several cDNA clones from a M. fascicularis retinal cDNA library. The nucleotide sequence of monkey opsin was determined and translated into an amino acid sequence. The amino acid sequences of opsins from a variety of organisms ranging from mollusks, arthropods, and vertebrates, including monkey, were then compared to identify highly conserved amino acid residues and to construct a phylogenetic tree of the rod opsins.

RESULTS

The monkey has a single major 2.2-kb transcript for opsin and minor transcripts of 1.5, 4.8, and 8.9 kb. The major transcript is highly abundant in the monkey retina and is localized to the inner segments of rod photoreceptor cells, typical of opsin mRNAs in other organisms. The sequence of monkey opsin closely resembles the human sequence at the nucleotide and the amino acid levels, with the latter having only 7 differences out of 348 residues. Comparison of the monkey sequence with the amino acid sequences from diverse organisms shows that all opsins share only 34 conserved amino acids, with the highest concentration of these residues occurring in the first cytoplasmic loop of rhodopsin. A hypothetical phylogenetic tree of the opsins suggests that both human and monkey opsin shared an ancestral gene with the mammalian Order Carnivora.

CONCLUSION

The opsins of man and Old World monkeys have diverged little from the time they shared a common ancestor. Comparison of opsins from diverse phyla, however, shows that only 34 residues have been conserved. These conserved residues are likely to be important in the structure and function of rhodopsin.