Identification of nocturnin, a novel photoreceptor specific leucine zipper protein that is regulated by a retinal circadian clock

Purpose. The photoreceptors of Xenopus laevis retina contain a circadian clock that regulates many aspects of physiology. In a recent screen for rhythmic mRNAs from Xenopus retina, we identified a pair of mRNAs that are expressed specifically in photoreceptors for a few hours at the beginning of subjective night, both in cyclic light and in constant conditions. cDNA clones corresponding to these mRNAs encode the amino acid sequence of a protein which we have named nocturnin or "night factor." Methods. The cDNA clones for nocturnin were obtained by screening a Xenopus night-time retinal cDNA library using the original differential display PCR product as a probe, and were characterized by DNA sequencing. The authentic 5′ ends of the mRNAs were identified by primer extension and by 5′ RACE. Database searches for sequence similarities were done using BLAST and FASTA search algorithms. Results. Characterization of nocturnin cDNA clones revealed a single open reading frame of 388 amino acids. The two different mRNAs contain identical open reading frames, and differ only in the usage of alternate polyadenylation sites, resulting in an extended 3′ untranslated region in the larger message. The deduced amino acid sequence contains a consensus leucine zipper protein interaction domain near the N-terminus, and sequence similarity to a portion of a known transcription factor from yeast, CCR4 (Malvar, et al., Genetics 132: 951-962, 1992) over a region from amino acid 98 to the C-terminus. Similarities to 3 human expressed sequence tags were also found. Conclusions. Understanding the mechanism of circadian regulation requires elucidation of both the molecular components of the clock itself as well as those that are regulated by the clock. Sequence information from the nocturnin clones reveals that it may be a rhythmic photoreceptor-specific transcription factor. Further functional characterization will be required to discern whether nocturnin is a part of the central clock mechanism or is part of the clock output. In either event, it should provide insight into how the clock regulates rhythmic physiological events in the retina.