Abstract
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-neuronal cells and neurons of rat. Both APP isoforms differ in regard to functional properties like post-translational modification, APP secretion, and proteolytic production of Abeta peptide from APP molecules. Since Abeta generation is an important factor in the development of Alzheimer's disease, one could anticipate that these major APP isoforms might contribute differentially to the mechanisms underlying neurodegeneration in Alzheimer's disease. In this study, we established an APP minigene system in a murine cell system to identify cis-acting elements controlling exon 15 recognition. A 12. 5-kilobase pair genomic fragment of the murine APP gene contained all cis-regulatory elements to reproduce the splicing pattern of the endogenous APP transcripts. By using this approach, two intronic cis-elements flanking exon 15 were identified that block the inclusion of exon 15 in APP transcripts of non-neuronal cells. Point mutation analysis of these intronic regions indicated 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 non-neuronal splicing system.
Highlights
Amyloid plaques represent characteristic neuropathological lesions in brains of patients with Alzheimer’s disease
Since in the present study we used the mouse system to identify cis-acting elements involved in the splicing regulation of exon 15, we first examined the expression pattern of exon 15-containing (APPex15) and lacking (L-amyloid precursor protein (APP)) APP mRNA isoforms in a number of organs of three adult female mice
Cell Type-specific Recognition of Exon 15 Is Conserved between Mouse and Rat—Consistent with previously published data on the distribution of leukocyte-derived APP (L-APP) transcripts in rat [11], our expression analysis revealed the ubiquitous occurrence of LAPP in mouse tissues
Summary
Amyloid plaques represent characteristic neuropathological lesions in brains of patients with Alzheimer’s disease. The expression levels of L-APP in the central nervous system, including hippocampus, cerebellum, and cerebral cortex, is low (Ͻ10%), and no significant L-APP amounts were detected in cultured primary rat neurons. These findings suggested that exon 15 is recognized by a cell type-specific mechanism in rat. Fusion of exon 14 with exon 16 creates a new post-translational modification motif ENEGSG This motif is recognized by xylosyltransferase that attaches a chondroitin sulfate glycosaminoglycan chain via xylose to the serine residue of the ENEGSG sequence. In order to get some insight into the splicing regulation of exon 15, in the present study we performed a molecular anal-
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have