Identification of cis-elements regulating exon 15 splicing of the amyloid precursor protein pre-mRNA

Alternative splicing of exon 15 of the amyloid precursor protein (APP) pre- mRNA generates two APP isoform groups App(ex15) (containing exon 15) and L- APP (without exon 15), which show a cell-specific distribution in non-neuro nal cells and neurons of rat. Both APP isoforms differ in regard to functio nal properties like posttranslational modification, APP secretion, and prot eolytic production of A beta peptide from APP molecules. Since A beta gener ation is an important factor in the development of Alzheimer's disease, one could anticipate that these major APP isoforms might contribute differenti ally to the mechanisms underlying neurodegeneration in Alzheimer's disease, In this study, we established an APP minigene system in a murine cell syst em to identify cis-acting elements controlling exon 15 recognition. A 12.5- kilobase pair genomic fragment of the murine APP gene contained all cis-reg ulatory elements to reproduce the splicing pattern of the endogenous APP tr anscripts. By using this approach, two intronic cis-elements flanking exon 15 were identified that block the inclusion of exon 15 in APP transcripts o f non-neuronal cells. Point mutation analysis of these intronic regions ind icated that pyrimidine-rich sequences are involved in the splice repressor function. Finally, grafting experiments demonstrated that these regulatory regions cell-specifically enhance the blockage of a chimeric exon in the no n-neuronal splicing system.