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Vilardo E.,CNR Institute of Neurobiology and Molecular Medicine | Barbato C.,CNR Institute of Neurobiology and Molecular Medicine | Barbato C.,EBRI European Brain Research Institute Fondazione EBRI Rita Levi Montalcini | Ciotti M.,CNR Institute of Neurobiology and Molecular Medicine | And 3 more authors.
Journal of Biological Chemistry | Year: 2010

The amyloid precursor protein (APP) and its proteolytic product amyloid beta (Aβ) are associated with both familial and sporadic forms of Alzheimer disease (AD). Aberrant expression and function of microRNAs has been observed in AD. Here, we show that in rat hippocampal neurons cultured in vitro, the down-regulation of Argonaute-2, a key component of the RNA-induced silencing complex, produced an increase in APP levels. Using site-directed mutagenesis, a microRNA responsive element (RE) for miR-101 was identified in the 3′-untranslated region (UTR) of APP. The inhibition of endogenous miR-101 increased APP levels, whereas lentiviral-mediated miR-101 overexpression significantly reduced APP and Aβ load in hippocampal neurons. In addition, miR-101 contributed to the regulation of APP in response to the proinflammatory cytokine interleukin-1β (IL-1β). Thus, miR-101 is a negative regulator of APP expression and affects the accumulation of Aβ, suggesting a possible role for miR-101 in neuropathological conditions. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.


Ruberti F.,CNR Institute of Neurobiology and Molecular Medicine | Barbato C.,CNR Institute of Neurobiology and Molecular Medicine | Barbato C.,EBRI European Brain Research Institute Fondazione EBRI Rita Levi Montalcini | Cogoni C.,EBRI European Brain Research Institute Fondazione EBRI Rita Levi Montalcini | Cogoni C.,University of Rome La Sapienza
Communicative and Integrative Biology | Year: 2010

Amyloid Precursor Protein (APP) and its proteolytic product amyloid beta (Aβ) are critical in the pathogenesis of Alzheimer's Disease (AD). APP gene duplication and transcriptional upregulation are linked to AD. In addition, normal levels of APP appear to be required for some physiological functions in the developing brain. Several studies in mammalian cell lines and primary neuron cultures indicate that RNA binding proteins and microRNAs interacting with regulatory regions of the APP mRNA modulate expression of APP post-transcriptionally. However, when the various mechanisms of APP posttranscriptional regulation are recruited and which of them are acting in a synergistic fashion to balance APP protein levels, is unclear. Recent studies suggest that further investigation of the molecules and pathways involved in APP post-transcriptional regulation are warranted. © 2010 Landes Bioscience.


PubMed | EBRI European Brain Research Institute Fondazione EBRI Rita Levi Montalcini
Type: Journal Article | Journal: Journal of neurochemistry | Year: 2010

MicroRNAs have been associated to fine-tuning spatial and temporal control of gene expression during neuronal development. The neuronal Cl(-) extruding, K(+)Cl(-) co-transporter 2 (KCC2) is known to play an important role in neuronal Cl(-) homeostasis and in determining the physiological response to activation of anion selective GABA receptors. Here we show that microRNA-92 is developmentally down-regulated during maturation of rat cerebellar granule neurons (CGNs) in vitro. Computational predictions suggest several high-ranking targets for microRNA-92 including the KCC2 gene. Consistently, the KCC2 protein levels were up-regulated in mature CGN in vitro and a functional association between microRNA-92 and KCC2 3 untranslated region was established using luciferase assays. The generation of an inward directed Cl(-) electrochemical gradient, necessary for the hyperpolarizing effect of GABA, requires robust KCC2 expression in several neuronal types. Here we show that lentiviral-mediated microRNA-92 over-expression reduced KCC2 protein levels and positively shifted reversal potential of GABA induced Cl(-) currents in CGNs. In addition KCC2 re-expression reversed microRNA-92 electrophysiological phenotype. Consistently microRNA-92 inhibition induced both an increase of the level of KCC2 and a negative shift in GABA reversal potential. These findings introduce a new player in the developmental change of GABA from depolarization to hyperpolarization.

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Loading EBRI European Brain Research Institute Fondazione EBRI Rita Levi Montalcini collaborators