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J. Biol. Chem., Vol. 281, Issue 45, 34288-34298, November 10, 2006

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Regulatory Factor Interactions and Somatic Silencing of the Germ Cell-specific ALF Gene^*

From the Department of Molecular and Cell Biology, University of Texas at Dallas, Richardson, Texas 75080

Germ cell-specific genes are active in oocytes and spermatocytes but are silent in all other cell types. To understand the basis for this seemingly simple pattern of regulation, we characterized factors that recognize the promoter-proximal region of the germ cell-specific TFIIA/-like factor (ALF) gene. Two of the protein-DNA complexes formed with liver extracts (C4 and C5) are due to the zinc finger proteins Sp1 and Sp3, respectively, whereas another complex (C6) is due to the transcription factor RFX1. Two additional complexes (C1 and C3) are due to the multivalent zinc finger protein CTCF, a factor that plays a role in gene silencing and chromatin insulation. An investigation of CTCF binding revealed a recognition site of only 17 bp that overlaps with the Sp1/Sp3 site. This site is predictive of other genomic CTCF sites and can be aligned to create a functional consensus. Studies on the activity of the ALF promoter in somatic 293 cells revealed mutations that result in increased reporter activity. In addition, RNAi-mediated down-regulation of CTCF is associated with activation of the endogenous ALF gene, and both CTCF and Sp3 repress the promoter in transient transfection assays. Overall, the results suggest a role for several factors, including the multivalent zinc finger chromatin insulator protein CTCF, in mediating somatic repression of the ALF gene. Release of such repression, perhaps in conjunction with other members of the CTCF, RFX, and Sp1 families of transcription factors, could be an important aspect of germ cell gene activation.

Received for publication, July 27, 2006 , and in revised form, September 11, 2006.

^* This work was supported by grants from the National Institutes of Health and The Welch Foundation (to J. D.). 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.

¹ To whom correspondence should be addressed: Dept. of Molecular and Cell Biology, University of Texas at Dallas, 2601 N. Floyd Rd., Richardson, TX 75080. Tel.: 972-883-6882; E-mail: dejong{at}utdallas.edu.

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