Signaling to extracellular signal-regulated kinase from ErbB1 kinase and protein kinase C: feedback, heterogeneity, and gating

Rebecca M. Perrett, Robert C. Fowkes, Christopher J. Caunt, Krasimira Tsaneva-Atanasova, Clive G. Bowsher, Craig A. McArdle*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

14 Citations (Scopus)

Abstract

Many extracellular signals act via the Raf/MEK/ERK cascade in which kinetics, cell-cell variability, and sensitivity of the ERK response can all influence cell fate. Here we used automated microscopy to explore the effects of ERK-mediated negative feedback on these attributes in cells expressing endogenous ERK or ERK2-GFP reporters. We studied acute rather than chronic stimulation with either epidermal growth factor (ErbB1 activation) or phorbol 12,13-dibutyrate (PKC activation). In unstimulated cells, ERK-mediated negative feedback reduced the population-average and cell-cell variability of the level of activated ppERK and increased its robustness to changes in ERK expression. In stimulated cells, negative feedback (evident between 5 min and 4 h) also reduced average levels and variability of phosphorylated ERK (ppERK) without altering the "gradedness" or sensitivity of the response. Binning cells according to total ERK expression revealed, strikingly, that maximal ppERK responses initially occur at submaximal ERK levels and that this non-monotonic relationship changes to an increasing, monotonic one within 15 min. These phenomena occur in HeLa cells and MCF7 breast cancer cells and in the presence and absence of ERK-mediated negative feedback. They were best modeled assuming distributive (rather than processive) activation. Thus, we have uncovered a novel, time-dependent change in the relationship between total ERK and ppERK levels that persists without negative feedback. This change makes acute response kinetics dependent on ERK level and provides a "gating" or control mechanism in which the interplay between stimulus duration and the distribution of ERK expression across cells could modulate the proportion of cells that respond to stimulation.

Original languageEnglish
Pages (from-to)21001-21014
Number of pages14
JournalJournal of Biological Chemistry
Volume288
Issue number29
Early online date10 Jun 2013
DOIs
Publication statusPublished - 19 Jul 2013

Keywords

  • Enzyme Activation
  • Epidermal Growth Factor
  • Extracellular Signal-Regulated MAP Kinases
  • Feedback, Physiological
  • HeLa Cells
  • Humans
  • Kinetics
  • MAP Kinase Signaling System
  • MCF-7 Cells
  • Microscopy, Fluorescence
  • Models, Biological
  • Phorbol 12,13-Dibutyrate
  • Phosphorylation
  • Protein Kinase C
  • Receptor, Epidermal Growth Factor
  • Time Factors
  • GONADOTROPIN-RELEASING-HORMONE
  • DUAL-SPECIFICITY PHOSPHATASES
  • ERK MAP KINASE
  • NEGATIVE FEEDBACK
  • SPATIOTEMPORAL REGULATION
  • NUCLEAR TRANSLOCATION
  • MAMMALIAN-CELLS
  • PHOSPHORYLATION
  • RECEPTOR

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