Molecular analysis of the promoter region of the gene encoding the beta-amyloid precursor protein.

ABSTRACT

The amyloid beta-peptide (approximately 4 kDa-M(r)) is generated by the proteolytic cleavage of a larger beta-amyloid precursor protein (beta APP) encoded by a gene on chromosome 21. The abnormality in gene regulation of beta APP may be an important factor in the neuropathology of Alzheimer's disease. The control of transcription is mediated by different DNA regulatory elements (cis-acting) present in the promoter of the gene. There are about 26 DNA motifs, present in the immediate 5'-flanking region of the beta APP gene, through which various cell/tissue-specific factors (trans-acting) can exert their influence on transcription. Here, the effects of nerve growth factor (NGF), basic fibroblast growth factor (bFGF), phorbol 12-myristate 13-acetate (PMA), interleukin-1 (IL-1) and retinoic acid (RA) on promoter activity of the beta APP gene were analyzed. To investigate the effect of these factors on promoter activity, a recombinant plasmid which contained sequences of -489 base pairs (bp) from the 5'-flanking region of the beta APP gene was used. The truncated region of the promoter was linked upstream to a reporter gene, chloramphenicol acetyl transferase (CAT). Promoter activity was tested by transient transfection of the fusion plasmid in PC12 cells using the electroporation method (960 microF at 350 V). The treatment of PC12 cells with either NGF, bFGF, PMA, IL-1 or RA stimulated the activity of the beta APP promoter. Treatment of cells with either NGF or bFGF resulted in a higher level of stimulation in the basal level of promoter activity than when cells were treated with either PMA, IL-1 or RA. The pre-treatment of cells with these factors for a duration of 4 days prior to transfection with the promoter plasmid was necessary for the stimulatory effect. The cells that were treated with either of these factors after transfection showed no significant change in the basal level of promoter activity. Thus, certain growth factors and a cytokine could enhance the basal level of promoter activity of the beta APP gene, suggesting a possible participation of a growth-factor (s)-mediated transcriptional element in the control of gene expression of beta APP.