Cloning of opsin cDNAs from the brain of Anolis carolinensis
Von Schantz M., Soni BG., Foster RG.
Purpose. We have previously shown that a range of different anti-opsin antibodies labelled CSF-contacting neurons in the septal area of the brain of the lizard Anolis carolinensis. This makes these cells strong candidates for the deep brain photoreceptors that mediate circadian responses to light in this species. In an attempt to characterize these putative photoreceptors and their transduction machinery we have started to isolate cDNA clones encoding their opsin(s). Methods. Initially, we used degenerate opsin primers in reverse transcription PCR with RNA from Anolis brain. The amplified DNA was subcloned and sequenced. Subsequently, a λgt11 cDNA library was constructed with Anolis brain mRNA. This library was screened with the PCR-amplified cDNA, as well as a chicken pineal opsin cDNA. Results. An 0.4 kb PCR product was obtained. Sequence analysis showed it to be identical with the rhodopsin-like gene previously cloned from an Anolis genomic library by Kawamura and Yokoyama (Gene 1995;149:267-270). Screening of the library with both probes gave several positive clones, the analysis of which is in progress. Conclusions. The detection of a message for the putative Anolis pineal opsin in the brain indicates that it is expressed in the deep brain photoreceptor as well as (or possibly instead of) in the pineal organ. We intend to further investigate this by cDNA cloning from pineal and ocular photoreceptors, as well as screening for other transduction genes.