The interaction of beta-amyloid with alpha1-chimaerin involved in the pathogenesis of Alzheimer's disease

Abstract

5’-UTR reporter construct (pGAL). MiR-346 mimic was co-transfected into mammalian cells along with the reporter construct to validate target predictions. APP mRNA and protein levels were assessed by RT-qPCR and Western blotting. Target protectors were used to disrupt the interaction between miR-346 and reporter mRNA. Results: Bioinformatic tools successfully predicted a miR-346 target site in APP 5’-UTR. Reporter assays in HeLa cells revealed that miR-346 significantly stimulated reporter expression via APP 5’-UTR. This stimulatory effect on reporter expression was reversed with co-transfection of a target protector designed against the predictedmiR-346 target site in APP 5’-UTR. To confirm the effect of miR-346 on native protein expression, miR-346 mimic was transfected into HeLa cells. In response, APP protein levels were dramatically increased relative to controls. This effect was replicated in human astrocytic U373 and rat neuronal-like PC12 cells. APP mRNA levels in HeLa were only slightly increased following miR-346 transfection, suggesting a post-transcriptional mechanism. In a comparison across cell lines, miR-346 levels were highest in CNS-like NT2 neurons as compared to non-neuronal cell lines. Conclusions: Our results reveal a novel, non-canonical regulatory interaction between miR-346 and the APP 5’-UTR mediated primarily through a post-transcriptional mechanism. Therefore, miR-346 may represent a novel drug target in AD.