title: Signaling to extracellular signal-regulated kinase from ErbB1 kinase and protein kinase C: feedback, heterogeneity, and gating.
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.
authors.affiliation: From the Laboratories for Integrative Neuroscience and Endocrinology, School of Clinical Sciences, University of Bristol, Whitson Street, Bristol BS13NY, United Kingdom.
authors.forename: Rebecca M
authors.initials: RM
authors.lastname: Perrett
authors.affiliation: Endocrine Signaling Group, Royal Veterinary College, Royal College St., London NW10TU, United Kingdom.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Fowkes
authors.affiliation: Department of Biology and Biochemistry, University of Bath, Claverton Down, Bath BA27AY, United Kingdom.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Caunt
authors.affiliation: Bristol Centre for Applied Nonlinear Mathematics, Department of Engineering Mathematics, University of Bristol, Bristol BS81TR, United Kingdom, and.
authors.forename: Krasimira
authors.initials: K
authors.lastname: Tsaneva-Atanasova
authors.affiliation: School of Mathematics, University of Bristol, Bristol BS81TW, United Kingdom.
authors.forename: Clive G
authors.initials: CG
authors.lastname: Bowsher
authors.affiliation: From the Laboratories for Integrative Neuroscience and Endocrinology, School of Clinical Sciences, University of Bristol, Whitson Street, Bristol BS13NY, United Kingdom,. Electronic address: craig.mcardle@bristol.ac.uk.
authors.forename: Craig A
authors.initials: CA
authors.lastname: McArdle
mesh_terms: D004789:Enzyme Activation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D025461:Feedback, Physiological / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D020935:MAP Kinase Signaling System / Q000187:drug effects; D061986:MCF-7 Cells; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D013997:Time Factors
chemical_list: D015240:Phorbol 12,13-Dibutyrate; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D011493:Protein Kinase C; D048049:Extracellular Signal-Regulated MAP Kinases
keywords: Cell Signaling; ERK; Epidermal Growth Factor (EGF); ErbB1; Feedback; MAP Kinases (MAPKs); Protein Kinase C (PKC)