Worm Breeder's Gazette 10(3): 52
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
Because the 520-600 nm wavelength range of photosensitivity of C. elegans (Burr 1985, Photochem. Photobiol. 41, 577-582) is within the range expected for rhodopsin-like visual pigments, we investigated the possibility that the C. elegans genome contains an opsin gene. We used a 1.6 kb Drosophila ninaE opsin cDNA [cRh1], provided by Charles Zuker, to probe Southern blots of C. elegans genomic DNA digested with EcoRI. At a moderate stringency level (62 C; 1xSSPE) the probe identified 4 strongly and several weakly hybridizing bands ranging in size from 1.5 to 7.7 kb. Some opsin genes cross-hybridize at moderate to high stringencies (e.g. bovine rod-cell opsin cDNA [cRhB-rod] with vertebrate rod-cell opsin genes and cRh1 with the Drosophila Rh2 gene). Many require low stringencies (e.g. cRhB-rod with vertebrate cone- cell opsin genes and cRhB-rod with Drosophila opsin genes), while others do not cross-hybridize (e.g. cRh1 with the Drosophila UV- sensitive opsin genes Rh3 and Rh4). Though other signal proteins of the opsin family (adrenergic receptors and muscarinic acetylcholine receptors) have several structural and functional features in common with opsins, their genes do not cross-hybridize with opsin genes even within the same species (human) at low stringencies. In the light of these observations, our results at moderate stringency suggest that C. elegans has opsin gene(s) with relatively close resemblance to the ninaE gene of Drosophila. We have screened a C. elegans genomic library in Lambda Charon 4 phage (constructed by T. Snutch) with cRh1 under conditions identical to those used for the genomic blots. Thirteen positive phage giving strong hybridization signals at moderate stringency were grouped into 5 classes (designated s401-s405) based on the restriction fragment patterns produced by combined digestion with EcoRI and HindIII. Representative phage clones have been sent to Alan Coulson and John Sulston to be localized on the chromosomal contig map. The 5 classes were further characterized using probes produced by clevage of cRh1 with PstI. The 0.8 kb segment contains the major 3' portion of the coding region of the ninaE opsin (Zuker, et al. 1985, Cell 40, 851-858), including the sequences encoding the second extracellular loop and the 11-cis retinal binding site, both of which are highly conserved in opsins but not the other signal proteins. This probe hybridizes strongly with 7 different restriction fragments derived from s401-s405. The 0.6 kb PstI segment of cRh1 contains 5' noncoding sequences and the 5' third of the coding region, including the highly conserved first cytoplasmic loop, the putative binding site for G-protein in all of the signal proteins of the opsin family. It hybridizes weakly with the same fragments detected by the 0.8 kb probe and weakly with 2 unique fragments in s403. Thus, restriction and hybridization data suggest that our cloned DNA fragments may represent as many as 5 different opsin genes of C. elegans which may have greater sequence identity to the Rh1 (Drosophila ninaE) gene than that between Rh1 and some other opsin genes.