DYSREGULATION OF THE Aβ DEGRADING ENZYME NEPRILYSIN (MME) COULD CONTRIBUTE TO ALZHEIMER’S DISEASE

Abstract

Alzheimer's disease (AD) is the foremost world-wide cause of age-related dementia. Its global prevalence constantly on the increase due to an aging population. Among the dementias, AD is characterized by accumulation of amyloid β (Aβ) plaques and aggregates of hyper-phosphorylated microtubule-associated protein tau (p-τ) in brains. Nevertheless, the underlying pathogenesis of AD is presently unknown, particularly the most common form of AD, late-onset. Even though genetic, epigenetic, and environmental factors play critical roles in AD etiology (Maloney and Lahiri, Lancet Neurol-2016), specific triggers are still unknown. We hypothesize that AD results from dysregulation of key biochemical pathways, particularly through regulatory molecules, such as non-coding RNAs, including microRNA (Long, Maloney, Rogers and Lahiri, Mol. Psychiatry-2018). We further posit that disruption of miRNA regulation of membrane metallo-endopeptidase (MME), or neprilysin could contribute to Aβ accumulation and hence AD pathogenesis, since MME is an important Aβ-clearing enzyme. Using the bioinformatics tools we predicted miRNA binding sites on the MME mRNA 3’-untranslated region (3’-UTR). We made neuronally-differentiated human neuroblastoma (SK-N-SH) cells with all-trans retinoic acid (ATRA). We transfected the differentiated cells with either siRNA against MME or candidate miRNAs. We assayed levels of MME mRNA and its protein by qRT-PCR and Western blotting, respectively. Upon identifying miR-181d as a functional candidate, we tested sequence specificity and miR-181d activity with reporter gene- MME 3’-UTR constructs. We also measured MME protein levels in several human fetal tissues. While several miRNAs were predicted to target the MME 3’-UTR, including miR-9, miR-181d, miR-128 and miR-216, only miR-181d treatment produced significant reduction of MME mRNA and protein levels in neuronal cultures. The functional assays further elucidated miR181d activity. MME is expressed in a variety of human organs, with the highest expression in kidney. Notably, miR-181d can down-regulate MME mRNA and protein through the specific target site within MME 3’-UTR. MME is a key Aβ-degrading enzyme involved in AD. Dysregulation by (presumably excess) miR-181d could contribute to AD pathogenesis. Supported by the NIA/NIH grant to DKL.