Promoter activity of the beta-amyloid precursor protein gene is negativelymodulated by an upstream regulatory element

Alzheimer's disease (AD) is characterized by the aggregation of the amyloid beta-peptide (A beta) which is generated from a larger beta-amyloid precur sor protein (beta APP). An overexpression of the beta APP gene in certain a reas of the AD brain has been suggested to be an important factor in the ne uropathology of AD. Here we have further characterized an upstream regulato ry element (URE) located between - 2257 and - 2234 of the human beta APP pr omoter. In addition to its location in the promoter, BLAST search reveals t hat URE is present in several introns of the beta APP gene and is also dete cted in many other genes. For functional studies, two promoter regions were cloned upstream of the reporter gene, chloramphenicol acetyl transferase ( CAT): (i) ph beta E-B - the plasmid that contains the human (h) promoter re gion (-2832 to + 101) including URE, and (ii) prh beta E-B - the plasmid th at contains the rhesus (rh) promoter region excluding URE as it lacks a 270 bp region of the h beta APP promoter (-2435 to -2165). Transient transfect ion studies indicate that ph beta E-B displayed significantly less CAT-prom oter activity than prh beta E-B in C6, PC12 and SK-N-SH cells. To determine the role of URE in a heterologous promoter, a p beta URE construct was mad e by subcloning URE in an enhancerless promoter vector pCATP. The p beta UR E-CAT construct displayed threefold to fourfold less promoter activity than pCATP when different cell lines were transfected with the plasmids. URE in teracts with a novel protein(s) as determined by the electrophoretic mobili ty shift assay (EMSA). Although the core DNA region of URE resembles with t he NF-kB element, URE-binding protein is not related to the NF-kB transcrip tion factor. When EMSA was performed with specific competitors in different cell lines, the labeled URE probe was not competed by the oligonucleotides specific for either the AP3, NF-1 or NF-kB transcription factor. The migra tion of the URE-protein complex was different from the NF-kB-protein comple x in the EMSA gel. A distinct URE-specific nuclear factor was also detected in frontal cortex of a normal human brain. These results suggest that the URE region acts as a repressor element. that the URE-binding protein is not related to the known transcription factors tested, and that the protein is present in astrocytic, neuroblastoma, PC12 cells and in the human brain. ( C) 1999 Elsevier Science B.V. All rights reserved.