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Lentiviral vectors to study the differential function of ERK1 and ERK2 MAP kinases

Indrigo, Marzia, Papale, Alessandro, Orellana, Daniel and Brambilla, Riccardo 2010. Lentiviral vectors to study the differential function of ERK1 and ERK2 MAP kinases. In: Seger, Rony ed. MAP Kinase Signaling Protocols, Methods in Molecular Biology, vol. 2. Springer, pp. 205-220. (10.1007/978-1-60761-795-2_12)

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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.

Item Type: Book Section
Date Type: Publication
Status: Published
Schools: Biosciences
Subjects: Q Science > Q Science (General)
Uncontrolled Keywords: ERK1; ERK2; MEK1; lentiviral vector; RNAi; microRNA; gene knockdown; cell proliferation; mouse embryo fibroblast; NIH 3T3
Publisher: Springer
ISBN: 9781607617945
ISSN: 1064-3745
Last Modified: 04 Jun 2017 03:47
URI: http://orca.cf.ac.uk/id/eprint/26550

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