Fig 5. G-quadruplex regulation of APP translation.

(A) Western blot analysis of cells transfected with reporter constructs containing APP ₆₉₅ coding sequence containing a C-terminal myc tag with either wild type (WT G-quad) or mutant (Mut G-quad) G-quadruplex sequence. Mock transfection was used to confirm that these constructs were over-expressed. Antibody 9B11 (anti-Myc) was used to detect APP-Myc. (B) Schematic representation of methods taken to identify newly synthesized APP (See methods). (C) Western blot analysis for immunoprecipitation demonstrating the successful pull down of Myc-tagged APP constructs (IP: using 9B11 anti myc mAb, IB: C1/6.1 anti APP). (D) Western blot analysis of total APP (top panel, IB: C1/6.1 anti APP) and newly synthesized APP (bottom panel, IB: Streptavidin). Statistical analysis was performed by normalizing the newly synthesized APP/Total APP (Streptavidin/C1/6.1) using a ratio paired t-test using Prism 6.0g for Mac.