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Lentiviral Vectors to Study the Differential Function of ERK1 and ERK2 MAP Kinases

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MAP Kinase Signaling Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 661))

Abstract

Accumulating evidence indicates that p44ERK1 and p42ERK2 mitogen-activated protein kinases (MAPKs) have distinct quantitative roles in cell signaling. In our recently proposed model of regulation of ERK1 and ERK2, p42 plays a major role in delivering signals from the cell membrane to the nucleus, while p44 acts as a partial agonist of ERK2 toward effectors and downstream activators, thus providing a fine tuning system of the global signaling output. Here, we describe systems to modulate MAPK signaling in vitro and in vivo via lentiviral vector (LV)-mediated gene transfer, using three systems: RNAi with small hairpin RNAs, microRNA-mediated gene knockdown, and expression of signaling-interfering mutants of MEK1. We show, by using proliferation assays in mouse embryo fibroblasts (MEF) and NIH 3T3 cells, that gene knockdown of ERK1 promotes cell proliferation in a manner indistinguishable from a constitutively active MEK1 construct, while ERK2 RNAi causes a significant growth arrest, similar to that observed with the ectopic expression of a dominant negative MEK1 mutant.

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Acknowledgments

This work was supported by the Michael J Fox Foundation for Parkinson’s Research and the Parkinson’s Disease Society of the UK, as well as by the Italian Ministry of Health (to RB).

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Authors and Affiliations

  1. Division of Neuroscience, Institute of Experimental Neurology, San Raffaele Foundation and University, Milano, Italy

    Marzia Indrigo, Alessandro Papale, Daniel Orellana & Riccardo Brambilla

Authors
  1. Marzia Indrigo
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  2. Alessandro Papale
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  3. Daniel Orellana
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  4. Riccardo Brambilla
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Corresponding author

Correspondence to Riccardo Brambilla .

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Editors and Affiliations

  1. Dept. Biological Regulation, Weizmann Institute of Science, Rehovot, Israel

    Rony Seger

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Indrigo, M., Papale, A., Orellana, D., Brambilla, R. (2010). Lentiviral Vectors to Study the Differential Function of ERK1 and ERK2 MAP Kinases. In: Seger, R. (eds) MAP Kinase Signaling Protocols. Methods in Molecular Biology, vol 661. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-795-2_12

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  • DOI: https://doi.org/10.1007/978-1-60761-795-2_12

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-60761-794-5

  • Online ISBN: 978-1-60761-795-2

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