CHARACTERIZATION OF THE CHICKEN CTCF GENOMIC LOCUS, AND INITIAL STUDYOF THE CELL-CYCLE-REGULATED PROMOTER OF THE GENE

CTCF is a multifunctional transcription factor encoded by a novel cand idate tumor suppressor gene (Filippova, G. N., Lindblom, A., Meinke, L . J., Klenova, E. M., Neiman, P. E., Collins, S. J., Doggett, N. D., a nd Lobanenkov, V, V, (1998) Genes Chromosomes Cancer 22, 26-36), We ch aracterized genomic organization of the chicken CTCF (chCTCF) gene, an d studied the chCTCF promoter, Genomic locus of chCTCF contains a CC-r ich untranslated exon separated from seven coding exons by a long intr on, The 2-kilobase pair region upstream of the major transcription sta rt site contains a CpG island marked by a ''Not-knot'' that includes s equence motifs characteristic of a TATA-less promoter of housekeeping genes. When fused upstream of a reporter chloramphenicol acetyltransfe rase gene, it acts as a strong transcriptional promoter in transient t ransfection experiments. The minimal 180-base pair chCTCF promoter reg ion that is fully sufficient to confer high level transcriptional acti vity to the reporter contains high affinity binding element for the tr anscription factor YY1, This element is strictly conserved in chicken, mouse, and human CTCF genes. Mutations in the core nucleotides of the YY1 element reduce transcriptional activity of the minimal chCTCF pro moter, indicating that the conserved YY1-binding sequence is critical for transcriptional regulation of vertebrate CTCF genes, We also noted in the chCTCF promoter several elements previously characterized in c ell cycle-regulated genes, including the ''cell cycle-dependent elemen t'' and ''cell cycle gene homology region'' motifs shown to be importa nt for S/G(2)-specific up-regulation of cdc25C, cdc2, cyclin A, and Pl k (polo-like kinase) gene promoters, Presence of the cell cycle-depend ent element/cell cycle gene homology region element suggested that chC TCF expression may be cell cycle-regulated. We show that both levels o f the endogenous chCTCF mRNA, and the activity of the stably transfect ed chCTCF promoter constructs, increase in S/G(2) cells.