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Novel reporter for faithful monitoring of ERK2 dynamics in living cells and model organisms

(2015) PLOS ONE. 10(10).
Author
Organization
Abstract
Uncoupling of ERK1/2 phosphorylation from subcellular localization is essential towards the understanding of molecular mechanisms that control ERK1/2-mediated cell-fate decision. ERK1/2 non-catalytic functions and discoveries of new specific anchors responsible of the subcellular compartmentalization of ERK1/2 signaling pathway have been proposed as regulation mechanisms for which dynamic monitoring of ERK1/2 localization is necessary. However, studying the spatiotemporal features of ERK2, for instance, in different cellular processes in living cells and tissues requires a tool that can faithfully report on its subcellular distribution. We developed a novel molecular tool, ERK2-LOC, based on the T2A-mediated coexpression of strictly equimolar levels of eGFP-ERK2 and MEK1, to faithfully visualize ERK2 localization patterns. MEK1 and eGFP-ERK2 were expressed reliably and functionally both in vitro and in single living cells. We then assessed the subcellular distribution and mobility of ERK2-LOC using fluorescence microscopy in non-stimulated conditions and after activation/inhibition of the MAPK/ERK1/2 signaling pathway. Finally, we used our coexpression system in Xenopus laevis embryos during the early stages of development. This is the first report on MEK1/ERK2 T2A-mediated coexpression in living embryos, and we show that there is a strong correlation between the spatiotemporal subcellular distribution of ERK2-LOC and the phosphorylation patterns of ERK1/2. Our approach can be used to study the spatiotemporal localization of ERK2 and its dynamics in a variety of processes in living cells and embryonic tissues.
Keywords
MAP KINASE, NUCLEAR TRANSLOCATION, SIGNAL-REGULATED KINASE, ACTIVATED PROTEIN-KINASE, MESODERM INDUCTION, XENOPUS EMBRYOS, GROWTH-FACTOR, 2A PEPTIDE, IN-VIVO, CASCADE

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Citation

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MLA
Sipieter, François et al. “Novel Reporter for Faithful Monitoring of ERK2 Dynamics in Living Cells and Model Organisms.” PLOS ONE 10.10 (2015): n. pag. Print.
APA
Sipieter, F., Cappe, B., Pisfil, M. G., Spriet, C., Bodart, J.-F., Cailliau-Maggio, K., Vandenabeele, P., et al. (2015). Novel reporter for faithful monitoring of ERK2 dynamics in living cells and model organisms. PLOS ONE, 10(10).
Chicago author-date
Sipieter, François, Benjamin Cappe, Mariano Gonzalez Pisfil, Corentin Spriet, Jean-François Bodart, Katia Cailliau-Maggio, Peter Vandenabeele, Laurent Héliot, and Franck Riquet. 2015. “Novel Reporter for Faithful Monitoring of ERK2 Dynamics in Living Cells and Model Organisms.” Plos One 10 (10).
Chicago author-date (all authors)
Sipieter, François, Benjamin Cappe, Mariano Gonzalez Pisfil, Corentin Spriet, Jean-François Bodart, Katia Cailliau-Maggio, Peter Vandenabeele, Laurent Héliot, and Franck Riquet. 2015. “Novel Reporter for Faithful Monitoring of ERK2 Dynamics in Living Cells and Model Organisms.” Plos One 10 (10).
Vancouver
1.
Sipieter F, Cappe B, Pisfil MG, Spriet C, Bodart J-F, Cailliau-Maggio K, et al. Novel reporter for faithful monitoring of ERK2 dynamics in living cells and model organisms. PLOS ONE. 2015;10(10).
IEEE
[1]
F. Sipieter et al., “Novel reporter for faithful monitoring of ERK2 dynamics in living cells and model organisms,” PLOS ONE, vol. 10, no. 10, 2015.
@article{6995610,
  abstract     = {Uncoupling of ERK1/2 phosphorylation from subcellular localization is essential towards the understanding of molecular mechanisms that control ERK1/2-mediated cell-fate decision. ERK1/2 non-catalytic functions and discoveries of new specific anchors responsible of the subcellular compartmentalization of ERK1/2 signaling pathway have been proposed as regulation mechanisms for which dynamic monitoring of ERK1/2 localization is necessary. However, studying the spatiotemporal features of ERK2, for instance, in different cellular processes in living cells and tissues requires a tool that can faithfully report on its subcellular distribution. We developed a novel molecular tool, ERK2-LOC, based on the T2A-mediated coexpression of strictly equimolar levels of eGFP-ERK2 and MEK1, to faithfully visualize ERK2 localization patterns. MEK1 and eGFP-ERK2 were expressed reliably and functionally both in vitro and in single living cells. We then assessed the subcellular distribution and mobility of ERK2-LOC using fluorescence microscopy in non-stimulated conditions and after activation/inhibition of the MAPK/ERK1/2 signaling pathway. Finally, we used our coexpression system in Xenopus laevis embryos during the early stages of development. This is the first report on MEK1/ERK2 T2A-mediated coexpression in living embryos, and we show that there is a strong correlation between the spatiotemporal subcellular distribution of ERK2-LOC and the phosphorylation patterns of ERK1/2. Our approach can be used to study the spatiotemporal localization of ERK2 and its dynamics in a variety of processes in living cells and embryonic tissues.},
  articleno    = {e0140924},
  author       = {Sipieter, François and Cappe, Benjamin and Pisfil, Mariano Gonzalez and Spriet, Corentin and Bodart, Jean-François and Cailliau-Maggio, Katia and Vandenabeele, Peter and Héliot, Laurent and Riquet, Franck},
  issn         = {1932-6203},
  journal      = {PLOS ONE},
  keywords     = {MAP KINASE,NUCLEAR TRANSLOCATION,SIGNAL-REGULATED KINASE,ACTIVATED PROTEIN-KINASE,MESODERM INDUCTION,XENOPUS EMBRYOS,GROWTH-FACTOR,2A PEPTIDE,IN-VIVO,CASCADE},
  language     = {eng},
  number       = {10},
  pages        = {30},
  title        = {Novel reporter for faithful monitoring of ERK2 dynamics in living cells and model organisms},
  url          = {http://dx.doi.org/10.1371/journal.pone.0140924},
  volume       = {10},
  year         = {2015},
}

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