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J. Biol. Chem., Vol. 280, Issue 8, 7244-7252, February 25, 2005

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Identification of an Enhancer That Controls Up-regulation of Fibronectin during Differentiation of Embryonic Stem Cells into Extraembryonic Endoderm^*

Tetsu Shirai, Satoru Miyagi¶, Daisuke Horiuchi, Tomoko Okuda-Katayanagi, Masazumi Nishimoto, Masami Muramatsu, Yoshio Sakamoto||, Makoto Nagata, Koichi Hagiwara, and Akihiko Okuda**

From the Division of Developmental Biology, Research Center for Genomic Medicine, Saitama Medical School, 1397-1 Yamane, Hidaka, Saitama 350-1241, Japan, Department of Respiratory Medicine, Saitama Medical School, 38 Morohongo, Moroyama, Iruma-gun, Saitama 350-0495, Japan, and ^¶Rational Evolutionary Design of Advanced Biomolecules (REDS) Group, Saitama Small Enterprise Promotion Cooperation, Skip City, Kawaguchi, Saitama 333-0844, Japan

The extraembryonic endoderm is derived from inner cell mass cells of the blastocyst during early mouse embryogenesis. Formation of the extraembryonic endoderm, which later contributes to the yolk sac, appears to be a prerequisite for subsequent differentiation of the inner cell mass. While embryonic stem cells can be induced to differentiate into extraembryonic endoderm cells in vitro, the molecular mechanisms underlying this process are poorly understood. We used a promoter trap approach to search for genes that are expressed in embryonic stem cells and are highly up-regulated during differentiation to the extraembryonic endoderm fate. We showed that fibronectin fits this expression profile. Moreover we identified an enhancer in the 12th intron of the fibronectin locus that recapitulated the endogenous pattern of fibronectin expression. This enhancer carries Sox protein-binding sequences, and our analysis demonstrated that Sox7 and Sox17, which are highly expressed in the extraembryonic endoderm, were involved in enhancer activity.

Received for publication, September 17, 2004 , and in revised form, November 17, 2004.

^* This work was supported in part by the Ministry of Education, Science, Sports, and Culture and was especially supported by a Ministry grant to Saitama Medical School Research Center for Genomic Medicine. This work was also performed as a part of the REDS Project, Saitama Prefecture Collaboration of Regional Entities for the Advancement of Technological Excellence, supported by the Japan Science and Technology Agency. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^|| Present address: Kanto Central Hospital of the Mutual Aid Association of Public School, 6-25-1 Kamiyoga Setagaya-ku, Tokyo 158-0098, Japan.

^** To whom correspondence should be addressed. Tel.: 81-42-985-7268; Fax: 81-42-985-7264; E-mail: akiokuda{at}saitama-med.ac.jp.

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