title: The Nuclear Translocation of ERK.
abstract: The ERK1 and ERK2 (ERK1/2) cascade is a central signaling pathway activated by a wide variety of extracellular agents that transmit the messages of G Protein Coupled Receptors (GPCRs) and Receptor Tyrosine Kinases (RTKs). Being such a central pathway, the activity of the cascade is well regulated, including by dynamic changes of the subcellular localization of components of the ERK1/2 cascade. In resting cells, ERK1/2 are localized in the cytosol due to their interactions with different anchoring proteins. After stimulation, ERK1/2 are phosphorylated by MEK1/2 on their regulatory TEY motif, which permits their detachment from the anchoring proteins. This detachment exposes ERK1/2 to additional phosphorylation on two serine residues (SPS motif) within the nuclear translocation signal (NTS) of the kinases. This additional phosphorylation allows ERK1/2 to interact with importin7, which consequently promotes their translocation to the nucleus. More studies are still required in order to better understand the mechanism and consequence of the nuclear translocation of ERK1/2. In this chapter, we describe some of the techniques used to study nuclear translocation of ERK1/2 in mammalian cells. We briefly mention methods such as digitonin permeabilization and cellular fractionation, as well as overexpression of reporter constructs. More thoroughly, we describe immunofluorescence, immunoprecipitation, and proximity ligation assay (PLA) approaches that are routinely used in our laboratory. Hopefully, the increase of knowledge based on these methods will open more opportunities for the identification of new therapeutic targets for diseases where the ERK1/2 cascade is dysregulated, such as cancer, neurodegenerative diseases, and diabetes.
authors.affiliation: Department of Biological Regulation, The Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Denise A
authors.initials: DA
authors.lastname: Berti
authors.affiliation: Department of Biological Regulation, The Weizmann Institute of Science, Rehovot, 76100, Israel. rony.seger@weizmann.ac.il.
authors.forename: Rony
authors.initials: R
authors.lastname: Seger
mesh_terms: D002352:Carrier Proteins; D002453:Cell Cycle; D002467:Cell Nucleus / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D042541:Intracellular Space; D010766:Phosphorylation; D011485:Protein Binding; D021381:Protein Transport; D015398:Signal Transduction
chemical_list: D002352:Carrier Proteins; D048049:Extracellular Signal-Regulated MAP Kinases
keywords: Co-immunoprecipitation; ERK; Immunoflouresence; MAPK; Nuclear translocation; Proximity ligation assay