title: Assaying Activation and Subcellular Localization of ERK in Cells and Tissues.
abstract: The extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) are the focus of many studies due to their involvement in numerous physiological and pathological processes, such as cell proliferation and differentiation, and oncogenic transformation, respectively. ERK1/2 belong to the mitogen-activated protein kinase (MAPKs) family, which are serine/threonine kinases that participate in signal transduction and are activated by dual phosphorylation in the Thr-X-Tyr motif located in their activation loop. In addition, ERK activation induces its dimerization and translocation into the nucleus. On the basis of this knowledge, different assays and tools have been developed to determine ERK activity or monitor its activation. In this chapter, we describe methods to assay ERK activity based on the ability of ERK immunocomplexes to phosphorylate a substrate, as well as on immunoblot analysis using antibodies that recognize ERK1/2 phosphorylated in the Thr-X-Tyr motif. In addition, we describe an immunocytochemistry procedure to reveal stimuli-induced nuclear translocation of ERK1/2.
authors.affiliation: Department of Biochemistry and Physiology, School of Pharmacy, University of Barcelona, Av. Joan XXIII, 27-31, 08028, Barcelona, Spain. ccaelles@ub.edu.
authors.forename: Carme
authors.initials: C
authors.lastname: Caelles
authors.affiliation: Department of Biochemistry and Physiology, School of Pharmacy, University of Barcelona, Av. Joan XXIII, 27-31, 08028, Barcelona, Spain.
authors.forename: Carles
authors.initials: C
authors.lastname: Bayod
authors.affiliation: Department of Biochemistry and Physiology, School of Pharmacy, University of Barcelona, Av. Joan XXIII, 27-31, 08028, Barcelona, Spain.
authors.forename: Melisa
authors.initials: M
authors.lastname: Morcillo
mesh_terms: D000818:Animals; D015153:Blotting, Western; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D042541:Intracellular Space / Q000378:metabolism; D021381:Protein Transport
chemical_list: D048049:Extracellular Signal-Regulated MAP Kinases
keywords: ERK activity; ERK nuclear translocation; ERK1; ERK2; Immunoblot analysis; Immunocomplex assay; Immunocytochemistry analysis; Phospho-ERK