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Activation of Amyloid Precursor Protein Processing by Growth Factors is Dependent on Ras GTPase Activity

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Abstract

The β-amyloid peptide is generated by the proteolysis of the amyloid precursor protein (APP) by the action of β- and γ-secretase. The mechanisms underlying this process are poorly understood. Using a cell-based reporter gene assay we analysed the possible signals and pathways that could be involved in APP cleavage. We used the stable cell line HeLa AG that expresses the human APP695 fused with the yeast transcription factor Gal4. This fusion protein is normally translocated into the plasma membrane and after APP-Gal4 cleavage, the AICD-Gal4 fragment released can activate the transcription of a luciferase reporter gene. Through this reporter system, we demonstrated that Ras GTPase, but not Ral and Rap, could promote APP-Gal4 cleavage. In addition HeLa AG cells stimulated with EGF or PDGF or overexpressing EGFR exhibit increased APP proteolysis in a Ras-dependent way. This process is also dependent on γ-secretase activity, being abolished by the γ-secretase inhibitor DAPT.

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Acknowledgments

This work was supported by a FAR (ex60%) grant to E.M. and by PRIN 2005 grant. We would like to thank L.A. Quilliam for providing pFLAG-CMV2-RalA and pMT2-HA-Rlf vectors; PG Traber for pRC-CMV-Gal4 vector; N. Zambrano for constitutively active Src (SrcYF) vector; R. Zippel for pc3myc-H-Ras plasmid; A. Wittinghofer for providing pCDNA3-Rap1 and pHSG-C3G -HA vectors; J.L. Bos for pRSV-rasN17 and pRSV-rasL61 constructs; L. Beguinot for pMMTV-EGFR vectors and M. Pierotti for pRC/CMV-TrkA plasmid.

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Authors and Affiliations

  1. Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza Della Scienza 2, 20126, Milan, Italy

    Loredana Amigoni, Michela Ceriani, Fiorella Belotti & Enzo Martegani

  2. CEINGE Biotecnologie Avanzate, 80145, Naples, Italy

    Giuseppina Minopoli

  3. Department of Biochemistry and Medical Biotechnology, University of Napoli Federico II, 80131, Naples, Italy

    Giuseppina Minopoli

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  1. Loredana Amigoni
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  2. Michela Ceriani
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  3. Fiorella Belotti
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  4. Giuseppina Minopoli
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  5. Enzo Martegani
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Correspondence to Enzo Martegani.

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Amigoni, L., Ceriani, M., Belotti, F. et al. Activation of Amyloid Precursor Protein Processing by Growth Factors is Dependent on Ras GTPase Activity. Neurochem Res 36, 392–398 (2011). https://doi.org/10.1007/s11064-010-0343-8

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  • DOI: https://doi.org/10.1007/s11064-010-0343-8

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