THE ZINC-FINGER PROTEIN CTCF BINDS TO THE APB-BETA DOMAIN OF THE AMYLOID BETA-PROTEIN PRECURSOR PROMOTER - EVIDENCE FOR A ROLE IN TRANSCRIPTIONAL ACTIVATION

The promoter of the amyloid beta-protein precursor (APP) gene directs high levels of cell type-specific transcription with 94 base pairs 5' to the main transcriptional start site. An essential activator domain in this proximal APP promoter is a nuclear factor binding site designa ted as APB beta, The recognition domain for the APB beta binding facto r is located between position -93 and -82 relative to the main transcr iptional start site. The nuclear factor that binds to the APB beta sit e was partially purified by multiple steps of ion exchange and hydroxy apatite chromatography. Based on UV cross-linking results, a protein w ith an apparent molecular mass of 140 kDa was selected as the putative APB beta binding protein. After the final purification step consistin g of preparative SDS-polyacrylamide gel electrophoresis, partial pepti de sequences were obtained that completely matched the transcriptional factor CTCF, This protein is a known regulator of c-myc and lysozyme gene expression, and it binds to a variety of diverse DNA sequences. T he binding of CTCF to the APB beta domain was further established by c ompetition with CTCF binding oligonucleotides in mobility shift electr ophoresis. The identity was also confirmed by the observation that the APB beta binding factor is recognized by antibodies against C- and N- terminal sequences of CTCF. In addition, oligonucleotide competition d uring in vitro transcription affirmed that CTCF acts as a transcriptio nal activator in the APP gene promoter.