Pineal expression-promoting element (PIPE), a cis-acting element, directs pineal-specific gene expression in zebrafish.
The pineal gland, sharing morphological and biochemical similarities with the retina, plays a unique and central role in the photoneuroendocrine system. The unique development of the pineal gland is directed by a specific combination of the expressed genes, but little is known about the regulatory mechanism underlying the pineal-specific gene expression. We isolated a 1.1-kbp fragment upstream of the zebrafish exo-rhodopsin (exorh) gene, which is expressed specifically in the pineal gland. Transgenic analysis using an enhanced green fluorescent protein reporter gene demonstrated that the proximal 147-bp region of the exorh promoter is sufficient to direct pineal-specific expression. This region contains three copies of a putative cone rod homeobox (Crx)Otx-binding site, which is known to be required for expression of both retina- and pineal-specific genes. Deletion and mutational analyses of the exorh promoter revealed that a previously uncharacterized sequence TGACCCCAATCT termed pineal expression-promoting element (PIPE) is required for pineal-specific promoter activity in addition to the CrxOtx-binding sites. By using the zebrafish rhodopsin (rh) promoter that drives retina-specific expression, we created a reporter construct having ectopic PIPE in the rh promoter at a position equivalent to that in the exorh promoter by introducing five nucleotide changes. Such a slight modification in the rh promoter induced ectopic enhanced green fluorescent protein expression in the pineal gland without affecting its retinal expression. These results identify PIPE as a critical cis-element contributing to the pineal-specific gene expression, in combination with the CrxOtx-binding site(s).
Asaoka
Yoichi
Y
Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Japan.
Mano
Hiroaki
H
Kojima
Daisuke
D
Fukada
Yoshitaka
Y
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D001665:Binding Sites; D004524:Eels / Q000235:genetics; D005136:Eye Proteins / Q000096:biosynthesis / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics*; D005801:Genes, Homeobox / Q000235:genetics; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D018398:Homeodomain Proteins / Q000378:metabolism; D008164:Luminescent Proteins / Q000096:biosynthesis; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics; D009928:Organ Specificity; D051857:Otx Transcription Factors; D010870:Pineal Gland / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D012160:Retina / Q000378:metabolism; D012243:Rhodopsin / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D015534:Trans-Activators / Q000378:metabolism; D015027:Zebrafish / Q000235:genetics*
D005136:Eye Proteins; D018398:Homeodomain Proteins; D008164:Luminescent Proteins; D009419:Nerve Tissue Proteins; D051857:Otx Transcription Factors; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C109288:cone rod homeobox protein; C403706:exorhodopsin; D049452:Green Fluorescent Proteins; D012243:Rhodopsin