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

  • Marzia Indrigo
  • Alessandro Papale
  • Daniel Orellana
  • Riccardo BrambillaEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 661)


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.

Key words

ERK1 ERK2 MEK1 Lentiviral vector RNAi microRNA Gene knockdown Cell proliferation Mouse embryo fibroblast NIH 3T3 



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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Copyright information

© Springer Science+Business Meida, LLC 2010

Authors and Affiliations

  • Marzia Indrigo
    • 1
  • Alessandro Papale
    • 1
  • Daniel Orellana
    • 1
  • Riccardo Brambilla
    • 1
    Email author
  1. 1.Division of Neuroscience, Institute of Experimental NeurologySan Raffaele Foundation and UniversityMilanoItaly

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