D.K. Lahiri and and J.M. Long


Laboratory of Molecular Neurogenetics, Department of Psychiatry, Institute of Psychiatric Research, Indiana University School of Medicine, 791 Union Drive. Indianapolis, Indiana, USA

Abstract:

The mammalian central nervous system (CNS) is a complex organ system, demanding an equally complex network of molecular pathways controlling the multitude of diverse, cellular activities.

Gene expression is a critical node at which regulatory control of molecular networks is executed. Thus, deciphering the various mechanisms employed in the physiological regulation of gene expression in the CNS is important for understanding the diseased state and for validating drug targets available for disease intervention. Our aim is to utilize the novel approach of studying the regulation of these gene products by microRNAs (miRNAs).  MiRNAs are an abundant class of small RNAs that mediate potent inhibitory effects on global gene expression. Recent advances in molecular methods allow us to study the contribution of these miRNAs to gene expression in CNS disorders, such as Alzheimer’s disease (AD) (Long and Lahiri, Experimental Neurology, 2012). Aberrations in AD are believed to result, in part, from the over-production of amyloid-β peptide (Aβ), a product of Aβ precursor protein (APP). Expression studies suggest that dysregulation of proteins involved in Aβ production, such as APP and beta-secretase, or BACE1, may contribute to excess Aβ deposition.  Elucidating how expression of these proteins is regulated will ultimately reveal new drug targets. Here we present data demonstrating miRNA-mediated regulation of APP and BACE1.

Recently, by using multiple bioinformatic tools and a series of functional studies in neuronal and glial cultures, we reported specific microRNA species (miR-101 and miR-153) regulate APP levels (Long and Lahiri, Biochem. Biophys. Res. Commun., 2011; Long, Ray and Lahiri, J. Biol. Chem., 2012). We and others have also identified additional set of miRNAs predicted to target the APP mRNA 3’-UTR and regulate APP levels (reviewed Long and Lahiri, Curr. Med. Chem., 2011). Here we report the discovery of novel BACE1-specific miRNAs. First, we prepared a chimeric BACE1 3’-UTR reporter construct (10.1kb) by inserting the 3.9 kb BACE1 3’-UTR downstream of a reporter Renilla luciferase gene and then delivered the reporter construct along with several miRNAs predicted to target the BACE1 3'-UTR into human astroglial U373 cells. Several “hits” (e.g. miR-339-5p) resulted in reduced reporter expression. We further validated the reporter expression data for miR-339-5p by Western analysis of native BACE1 levels, which were significantly reduced following miR-339-5p delivery, with a potential in reducing toxic Aβ levels. Our results reveal a novel regulatory interaction between two important AD-related genes (APP and BACE1) and specific endogenously expressed miRNA species. These regulatory interactions are likely to serve as novel therapeutic targets and should enable the development of treatment strategies that may prove beneficial in the fight against AD.

This work is supported by grants from Alzheimer’s Association and NIH to Dr. D.K. Lahiri.