Entrainment of circadian clocks to environmental cues such as photoperiod ensures that daily biological rhythms stay in synchronization with the Earth's rotation. The vertebrate pineal organ has a conserved role in circadian regulation as the primary source of the nocturnal hormone melatonin. In lower vertebrates, the pineal has an endogenous circadian clock as well as photoreceptive cells that regulate this clock. The zebrafish opsin protein Exo-rhodopsin (Exorh) is expressed in pineal photoreceptors and is a candidate to mediate the effects of environmental light on pineal rhythms and melatonin synthesis. We demonstrate that Exorh has an important role in regulating gene transcription within the pineal. In developing embryos that lack Exorh, expression of the exorh gene itself and of the melatonin synthesis gene serotonin N-acetyl transferase 2 (aanat2) are significantly reduced. This suggests that the Exorh protein at the cell membrane is part of a signaling pathway that positively regulates transcription of these genes, and ultimately melatonin production, in the pineal. Like many other opsin genes, exorh is expressed with a daily rhythm: mRNA levels are higher at night than during the day. We found that the transcription factor Orthodenticle homeobox 5 (Otx5) activates exorh transcription, while the putative circadian clock component Period 3 (Per3) represses expression during the day, thereby contributing to the rhythm of transcription. This work identifies novel roles for Exorh and Per3, and gives insight into potential interactions between the sensory and circadian systems within the pineal.
Bibliographical noteFunding Information:
The authors thank Drs. Kathleen Molyneaux, Greg Matera, Marge Sedensky, Phil Morgan, and Marnie Halpern for their helpful comments on the manuscript, Drs. David Klein (National Institutes of Health), Yoav Gothilf (Tel Aviv University), and Bernard and Christine Thisse (Institut de Génétique et Biologie Moléculaire et Celluaire) for generously sharing plasmids, Dr. Yoshitaka Fukada (The University of Tokyo) for generously providing the 1055):GFP transgenic line and the exorh (− exorh cDNA, Dr. Mario Caccamo (Sanger Center) for advice on exorh genomic sequence, and Ms. Allisan Aquilina-Beck, Ms. Kristine Ilagan, and Mr. Brian Chen for their expert technical assistance. This work was supported in part by Research Grant No. 5-FY02-259 from the March of Dimes Birth Defects Foundation and Research Grant (J.O.L), No.T32 HD07104-29 Normal and Abnormal Development from the NIH/CHHD Institutional Pre-Doctoral Research Training Grant (L.X.P.), and Phi Beta Kappa Students Research Awards (O.P. and R.R.N.).
- Circadian rhythm
- Orthodenticle homeobox 5
- Period 3
- Pineal organ
- Serotonin N-acetyl transferase
- Transcriptional regulation