Characterization of the APP proximal promoter and 5'-untranslated regions: identification of cell type-specific domains and implications in APP gene expression and Alzheimer's disease.

Alzheimer's disease is characterized by brain deposition of toxic amyloid beta-peptide (Abeta), generated from the Abeta precursor protein (APP). APP gene expression is regulated via the proximal promoter region (PPR; -46/-1 in the human sequence; +1 transcription start) and the 5'-untranslated region (5'-UTR; +1/+147). We have recently identified a unique CAGA sequence, "amyloid" (+83/+86) present only in the APP gene from amyloid plaque-forming species, absent in all APP-like-proteins' (APLP1 and APLP2) genes. To assay functional activity of PPR + UTR and 5'-UTR regions that either contain or lack the "amyloid" box, we tested nine constructs in transient transfection studies. We observed significantly high reporter gene activity with -46/144, -46/100, -46/54, and 54/144 constructs. The 54/100 fragment, which contains a transforming growth factor-beta/ "amyloid"/interleukin-1 acute box cassette, showed different activity depending on cell type. Electrophoretic mobility shift assay (EMSA) showed distinct DNA-nuclear protein interaction in all fragments, differing among both cell types and specific fragment. Reporter gene expression corroborates with the DNA-protein binding pattern. To directly examine the "amyloid" box, we generated oligomers for CAGA mutants or mutated adjacent nucleotides. EMSA results showed that altering "amyloid" or adjacent sequence alters specific DNA-nuclear protein interaction in both mutation- and cell-type-specific manners. Reporter gene assay reveals mutant-specific expression effects. Therefore, the -46/54 region appears to be essential for basal expression of the APP gene, the 54/100 and 100/144 regions may have tissue-specific activity, and the "amyloid" CAGA box plays a role in APP gene regulation.