title: Transgenic zebrafish model to study translational control mediated by upstream open reading frame of human chop gene.
abstract: Upstream open reading frame (uORF)-mediated translational inhibition is important in controlling key regulatory genes expression. However, understanding the underlying molecular mechanism of such uORF-mediated control system in vivo is challenging in the absence of an animal model. Therefore, we generated a zebrafish transgenic line, termed huORFZ, harboring a construct in which the uORF sequence from human CCAAT/enhancer-binding protein homologous protein gene (huORF(chop)) is added to the leader of GFP and is driven by a cytomegalovirus promoter. The translation of transgenic huORF(chop)-gfp mRNA was absolutely inhibited by the huORF(chop) cassette in huORFZ embryos during normal conditions, but the downstream GFP was only apparent when the huORFZ embryos were treated with endoplasmic reticulum (ER) stresses. Interestingly, the number and location of GFP-responsive embryonic cells were dependent on the developmental stage and type of ER stresses encountered. These results indicate that the translation of the huORF(chop)-tag downstream reporter gene is controlled in the huORFZ line. Moreover, using cell sorting and microarray analysis of huORFZ embryos, we identified such putative factors as Nrg/ErbB, PI3K and hsp90, which are involved in huORF(chop)-mediated translational control under heat-shock stress. Therefore, using the huORFZ embryos allows us to study the regulatory network involved in human uORF(chop)-mediated translational inhibition.
authors.affiliation: Institute of Molecular and Cellular Biology, College of Life Science, National Taiwan University, Taipei, Taiwan.
authors.forename: Hung-Chieh
authors.initials: HC
authors.lastname: Lee
authors.forename: Yi-Jiun
authors.initials: YJ
authors.lastname: Chen
authors.forename: Yu-Wei
authors.initials: YW
authors.lastname: Liu
authors.forename: Kai-Yen
authors.initials: KY
authors.lastname: Lin
authors.forename: Shaio-Wen
authors.initials: SW
authors.lastname: Chen
authors.forename: Cheng-Yung
authors.initials: CY
authors.lastname: Lin
authors.forename: Yi-Chin
authors.initials: YC
authors.lastname: Lu
authors.forename: Pei-Chun
authors.initials: PC
authors.lastname: Hsu
authors.forename: Sheng-Chung
authors.initials: SC
authors.lastname: Lee
authors.forename: Huai-Jen
authors.initials: HJ
authors.lastname: Tsai
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D002460:Cell Line; D059865:Endoplasmic Reticulum Stress; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D008957:Models, Genetic; D016366:Open Reading Frames; D014176:Protein Biosynthesis; D038621:Regulatory Sequences, Ribonucleic Acid; D015398:Signal Transduction; D051742:Transcription Factor CHOP / Q000096:biosynthesis / Q000235:genetics*; D014158:Transcription, Genetic; D015027:Zebrafish / Q000196:embryology / Q000235:genetics*; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: C497249:DDIT3 protein, human; D018841:HSP90 Heat-Shock Proteins; D038621:Regulatory Sequences, Ribonucleic Acid; D029961:Zebrafish Proteins; D051742:Transcription Factor CHOP