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The Journal of Neuroscience, June 1, 2002, 22(11):4357-4363

A Negative Regulatory Element Required for Light-Dependent pinopsin Gene Expression

Yoko Takanaka, Toshiyuki Okano, Kazuyuki Yamamoto, and Yoshitaka Fukada

Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, and Japan Science and Technology, Core Research for Evolutional Science and Technology, Tokyo 113-0033, Japan

In vertebrates, a variety of light-stimulated genes are distributed in the retina, the pineal gland, and the suprachiasmatic nucleus, but a cis-element(s) responsible for the light-dependent transcriptional regulation is left unexplored. Focusing on the pinopsin gene, a light-stimulated gene in the chick pineal gland, we performed a transcriptional analysis in the primary culture of the chick pineal cells that were transiently transfected with a luciferase reporter gene fused with various lengths of the 5' upstream region of the pinopsin gene. Light-dependent enhancer activity was detectable in the construct with the upstream region between 1156 and +31. Introduction of mutations within the 18 bp sequence at positions 1103 to 1086 (TGGCACGTGGGGTTCCTC), including a CACGTG E-box sequence, elevated the transcriptional activity in the dark and thereby abrogated the light dependency, suggesting that the 18 bp sequence is essential for a reduction of the transcriptional activity in the dark. In an electrophoretic mobility-shift assay, we identified a pineal nuclear factor(s) capable of binding to the 18 bp element in a sequence-specific manner. When a 49 bp fragment (1122 to 1074) including the 18 bp sequence was placed upstream of the simian virus 40 promoter, the transcriptional activity was dramatically suppressed regardless of light conditions in the chick pineal cells, and a more pronounced repression was observed in nonpineal/nonphotosensory LMH and NIH 3T3 cells. These results suggest that the 18 bp element in the pinopsin promoter constitutes the binding site of a ubiquitous factor that serves for the transcriptional repression that is required, although not sufficient, for the light-dependent expression of pinopsin gene in the chick pinealocytes.

Key words: chicken; gene expression; light induction; pineal gland; pinopsin gene; transcriptional regulation

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Copyright © 2002 Society for Neuroscience  0270-6474/02/22114357-07$05.00/0

This article has been cited by other articles:

				T. Okano and Y. Fukada Chicktacking Pineal Clock J. Biochem., December 1, 2003; 134(6): 791 - 797. [Abstract] [Full Text] [PDF]

				Y. Asaoka, H. Mano, D. Kojima, and Y. Fukada Pineal expression-promoting element (PIPE), a cis-acting element, directs pineal-specific gene expression in zebrafish PNAS, November 26, 2002; 99(24): 15456 - 15461. [Abstract] [Full Text] [PDF]

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