Regulation of the Germ Cell‐Specific Mouse ALF Gene

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 ALF gene. Two of the protein‐DNA complexes formed with liver extracts (C4 and C5) are due to the Zn‐finger proteins Sp1 and Sp3, respectively, while 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 concensus. Studies on the activity of the ALF promoter in somatic 293 cells revealed mutations which result in increased reporter activity. In addition, RNAi‐mediated downregulation 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.