The zebrafish Period2 protein positively regulates the circadian clock through mediation of RAR-related orphan receptor alpha (Ror{alpha}) [Cell Biology]

December 28th, 2014 by Wang, M., Zhong, Z., Zhong, Y., Zhang, W., Wang, H.

We report characterization of a null mutant for zebrafish circadian clock gene period2 (per2) generated by Transcription Activator-Like Effector Nuclease (TALEN) and a positive role of PER2 in vertebrate circadian regulation. Locomotor experiments showed that per2 mutant zebrafish display reduced activities under LD (light-dark) and 2-hour phase delay under DD (constant darkness), and quantitative real-time PCR analyses showed up-regulation of cry1aa, cry1ba, cry1bb and aanat2 but down-regulation of per1b, per3 and bmal1b in per2 mutant zebrafish; suggesting that Per2 is essential for the zebrafish circadian clock. Luciferase reporter assays demonstrated that Per2 represses aanat2 expression through E-box and enhances bmal1b expression through RORE, implicating that Per2 plays dual roles in the zebrafish circadian clock. Cell transfection and co-IP assays revealed that Per2 enhances bmal1b expression through binding to orphan nuclear receptor Rorα. The enhancing effect of mouse PER2 on Bmal1 transcription is also mediated by RORα even though it binds to REV-ERBα. Moreover, zebrafish Per2 also appears to have tissue-specific regulatory roles in numerous peripheral organs. These findings help define the Per2's essential functions in the zebrafish circadian clock, and in particular provide strong evidence for a positive role of PER2 in the vertebrate circadian system.
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