title: The zebrafish period2 protein positively regulates the circadian clock through mediation of retinoic acid receptor (RAR)-related orphan receptor α (Rorα).
abstract: We report the characterization of a null mutant for zebrafish circadian clock gene period2 (per2) generated by transcription activator-like effector nuclease and a positive role of PER2 in vertebrate circadian regulation. Locomotor experiments showed that per2 mutant zebrafish display reduced activities under light-dark and 2-h phase delay under 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 the Ror/Rev-erb response element, implicating that Per2 plays dual roles in the zebrafish circadian clock. Cell transfection and co-immunoprecipitation 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 essential functions of Per2 in the zebrafish circadian clock and in particular provide strong evidence for a positive role of PER2 in the vertebrate circadian system.
authors.affiliation: From the Center for Circadian Clocks and School of Biology and Basic Medical Sciences, Medical College, Soochow University, Suzhou 215123, Jiangsu, China.
authors.forename: Mingyong
authors.initials: M
authors.lastname: Wang
authors.affiliation: From the Center for Circadian Clocks and School of Biology and Basic Medical Sciences, Medical College, Soochow University, Suzhou 215123, Jiangsu, China.
authors.forename: Zhaomin
authors.initials: Z
authors.lastname: Zhong
authors.affiliation: From the Center for Circadian Clocks and School of Biology and Basic Medical Sciences, Medical College, Soochow University, Suzhou 215123, Jiangsu, China.
authors.forename: Yingbin
authors.initials: Y
authors.lastname: Zhong
authors.affiliation: From the Center for Circadian Clocks and School of Biology and Basic Medical Sciences, Medical College, Soochow University, Suzhou 215123, Jiangsu, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: From the Center for Circadian Clocks and School of Biology and Basic Medical Sciences, Medical College, Soochow University, Suzhou 215123, Jiangsu, China han.wang88@gmail.com wanghan@suda.edu.cn.
authors.forename: Han
authors.initials: H
authors.lastname: Wang
mesh_terms: D056930:ARNTL Transcription Factors / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D047369:Chromatin Immunoprecipitation; D057906:Circadian Clocks / Q000502:physiology*; D004625:Embryo, Nonmammalian; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D007124:Immunoenzyme Techniques; D051379:Mice; D009043:Motor Activity; D056950:Period Circadian Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D018168:Receptors, Retinoic Acid / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015533:Transcriptional Activation; D015027:Zebrafish; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D056930:ARNTL Transcription Factors; D005136:Eye Proteins; D056950:Period Circadian Proteins; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D018168:Receptors, Retinoic Acid; D029961:Zebrafish Proteins; C498846:per2 protein, zebrafish; C521431:roraa protein, zebrafish
keywords: Circadian Clock; Gene Regulation; Period2; Rev-ErbAα (NR1D1); Rorα; Transgenic; Zebrafish