Signaling from the Living Plasma Membrane
2011; Cell Press; Volume: 144; Issue: 6 Linguagem: Inglês
10.1016/j.cell.2011.01.029
ISSN1097-4172
AutoresHernán E. Grecco, Malte Schmick, Philippe I. H. Bastiaens,
Tópico(s)Photoreceptor and optogenetics research
ResumoOur understanding of the plasma membrane, once viewed simply as a static barrier, has been revolutionized to encompass a complex, dynamic organelle that integrates the cell with its extracellular environment. Here, we discuss how bidirectional signaling across the plasma membrane is achieved by striking a delicate balance between restriction and propagation of information over different scales of time and space and how underlying dynamic mechanisms give rise to rich, context-dependent signaling responses. In this Review, we show how computer simulations can generate counterintuitive predictions about the spatial organization of these complex processes. Our understanding of the plasma membrane, once viewed simply as a static barrier, has been revolutionized to encompass a complex, dynamic organelle that integrates the cell with its extracellular environment. Here, we discuss how bidirectional signaling across the plasma membrane is achieved by striking a delicate balance between restriction and propagation of information over different scales of time and space and how underlying dynamic mechanisms give rise to rich, context-dependent signaling responses. In this Review, we show how computer simulations can generate counterintuitive predictions about the spatial organization of these complex processes. Biological systems operate within a carefully tailored balance of opposing tendencies, favoring one or the other in response to internal and external cues. Such duality between robustness and adaptability or between exploration of possibilities and commitment to a decision, for example, permeates every level of organization. The function of the plasma membrane is an excellent example of this duality, as it defines the cell by isolating it from the extracellular environment while at the same time integrating the cell with its surroundings by transferring messenger molecules or initiating reaction cascades within it. Isolation versus communication is therefore the precarious balance that the plasma membrane must continuously maintain, separating the outside from the inside while presenting each a representation of the other. To the inside, the plasma membrane summarizes the cell's "social" context while projecting the cell's state to the outside. For this reason, plasma membrane function is fundamental not only to keep a single cell alive, but also to maintain its proper behavior in the organismal collective. The plasma membrane is composed of a bilayer of lipids and incorporated proteins, whose interactions as an ensemble enable it to receive, remember, process, and relay information along and across it. These interactions form a signal transduction hierarchy of interconnected time- and lengthscales bridging more than three orders of magnitude, from nanometer-sized proteins to the micrometer scale of the cell. Within each level of the hierarchy, the lengthscales (how far information will spread) are coupled with the timescales (how fast information will spread) through underlying physical-chemical mechanisms such as free diffusion, reaction-diffusion, or active transport. In its most primitive state, the plasma membrane forms a spherical shell, 5 nm thick, and is permeable only to small nonpolar molecules such as oxygen and nitrogen. In a water-based environment, lipids shield their hydrophobic tails from the surrounding polar fluid, exposing their more hydrophilic heads. This arrangement minimizes the free energy of the water-lipid system and therefore occurs spontaneously. This property of self-assembly provided a convenient evolutionary path for the generation of a relatively stable supramolecular structure that shields its contents from the dissipative effects of diffusion (Griffiths, 2007Griffiths G. Cell evolution and the problem of membrane topology.Nat. Rev. Mol. Cell Biol. 2007; 8: 1018-1024Crossref PubMed Scopus (37) Google Scholar). However, the plasma membrane of the modern cell is not a static, self-assembled system but is continuously renewed to preserve its nonequilibrium state. For example, its lipid composition is dynamically maintained by a combination of lipid synthesis and chemical conversion, vesicular fusion and fission events that tie into intracellular transport and sorting processes (van Meer et al., 2008van Meer G. Voelker D.R. Feigenson G.W. Membrane lipids: where they are and how they behave.Nat. Rev. Mol. Cell Biol. 2008; 9: 112-124Crossref PubMed Scopus (3810) Google Scholar). The lipids, which were previously thought to serve only a structural function, are themselves subject to chemical modification and can thereby relay signals. The resulting axial and lateral asymmetry of the membrane can be rapidly modulated to allow for bidirectional information transfer. Lipids also provide a fluid matrix in which proteins reside and diffuse laterally (Zimmerberg and Gawrisch, 2006Zimmerberg J. Gawrisch K. The physical chemistry of biological membranes.Nat. Chem. Biol. 2006; 2: 564-567Crossref PubMed Scopus (74) Google Scholar). These membrane proteins, which represent more than 50% of the cross-sectional area of the membrane in some cell types (Janmey and Kinnunen, 2006Janmey P.A. Kinnunen P.K. Biophysical properties of lipids and dynamic membranes.Trends Cell Biol. 2006; 16: 538-546Abstract Full Text Full Text PDF PubMed Scopus (257) Google Scholar), provide the machinery for most of the plasma membrane's dynamic properties. In addition to structural and sensory functions, they mediate matter exchange with the environment, enabling the membrane to actively and passively regulate transport of substances across it, even against a concentration gradient. This, for example, can generate ion gradients across the membrane that have important physiological functions such as water homeostasis and electrical excitability. Experimental work with model lipid membranes has nevertheless been one of the main sources of quantitative information about the physical properties of lipid bilayers (Janmey and Kinnunen, 2006Janmey P.A. Kinnunen P.K. Biophysical properties of lipids and dynamic membranes.Trends Cell Biol. 2006; 16: 538-546Abstract Full Text Full Text PDF PubMed Scopus (257) Google Scholar). Biophysical parameters such as rigidity, tension, spontaneous curvature, and elastic moduli have thus been determined as a function of temperature, hydration, and lipid composition. Such model membranes clearly lack the dynamic features of their real-life equivalents. For example, lipid composition is generally symmetrical in model membranes (Devaux and Morris, 2004Devaux P.F. Morris R. Transmembrane asymmetry and lateral domains in biological membranes.Traffic. 2004; 5: 241-246Crossref PubMed Scopus (209) Google Scholar), and phenomena such as membrane coupling to the cytoskeleton (Kwik et al., 2003Kwik J. Boyle S. Fooksman D. Margolis L. Sheetz M.P. Edidin M. Membrane cholesterol, lateral mobility, and the phosphatidylinositol 4,5-bisphosphate-dependent organization of cell actin.Proc. Natl. Acad. Sci. USA. 2003; 100: 13964-13969Crossref PubMed Scopus (387) Google Scholar) are difficult to reproduce in vitro. Almost 40 years have passed since Singer and Nicholson wrote their seminal work detailing the fluid mosaic model of the plasma membrane (Singer and Nicolson, 1972Singer S.J. Nicolson G.L. The fluid mosaic model of the structure of cell membranes.Science. 1972; 175: 720-731Crossref PubMed Scopus (5764) Google Scholar). In this work, which elegantly integrated the experimental and theoretical knowledge of the time, the authors stated:Biological membranes play a crucial role in almost all cellular phenomena, yet our understanding of the molecular organization of membranes is still rudimentary. Experience has taught us, however, that in order to achieve a satisfactory understanding of how any biological system functions, the detailed molecular composition and structure of that system must be known. In spite of the enormous amount of knowledge about the structure and composition of the plasma membrane gathered in recent decades, our understanding of it can still be considered "rudimentary" in light of the complexity of its dynamics that have become apparent since then. A major challenge will therefore be to animate our rather static view of the plasma membrane by bringing our model membrane systems to life in the test tube. Here, we will discuss the impact that the dynamic, "living" membrane has on cellular information processing. From the extensive range of research available, we focus on examples that represent canonical mechanisms to constrain information within the cell, relying on the plasma membrane as a dynamically maintained supramolecular structure. The bidirectional transduction of signals by the plasma membrane is modulated by its state, and the cell's historical context therefore determines its response to incoming signals. However, incoming signals also modify the state of the membrane, and in this way, the transducing medium becomes the message. How fast and to what extent this signal is propagated across the membrane must be tightly regulated to allow information to spread on a scale that is relevant to the biological process while preventing spurious responses. This requires a balance between responding to an actual signal and resisting spurious events induced by noise, a feat that is achieved by partitioning the plasma membrane into domains that span several time- and lengthscales, corresponding to the dimensions at which the biological processes operate. The largest partitions of the plasma membrane occur at a micrometer scale. For example, the partitioning of epithelial cells into apical and basolateral domains generates a cellular polarity that enables transcytotic vectorial transport between two distinct extracellular environments (Mellman and Nelson, 2008Mellman I. Nelson W.J. Coordinated protein sorting, targeting and distribution in polarized cells.Nat. Rev. Mol. 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The resulting apical and basolateral domains maintain their different lipid and protein compositions dynamically through the life cycle of the cell (Muth and Caplan, 2003Muth T.R. Caplan M.J. Transport protein trafficking in polarized cells.Annu. Rev. Cell Dev. Biol. 2003; 19: 333-366Crossref PubMed Scopus (97) Google Scholar, Simons and van Meer, 1988Simons K. van Meer G. Lipid sorting in epithelial cells.Biochemistry. 1988; 27: 6197-6202Crossref PubMed Scopus (1051) Google Scholar). The living plasma membrane, together with PAR proteins, forms a self-referencing system that establishes polarity by mechanochemically restructuring the cell. Though such a coarse-grained partitioning provides the cell with a stable polarized structure on a timescale of days, it does not supply a sufficiently rapid response mechanism that is appropriate for localized cues such as those that occur during chemotaxis, for which a short-term, fine-grained spatial memory is needed. In model membranes, protein diffusion is fast enough (D ≈5 μm2/s) to equilibrate across microns within seconds (Ramadurai et al., 2010Ramadurai S. Duurkens R. Krasnikov V.V. Poolman B. Lateral diffusion of membrane proteins: consequences of hydrophobic mismatch and lipid composition.Biophys. J. 2010; 99: 1482-1489Abstract Full Text Full Text PDF PubMed Scopus (39) Google Scholar). Under these conditions, localized signals such as activated receptors would redistribute across an area equivalent to the cell surface in a few minutes. In such homogeneous membrane systems, the diffusion coefficient scales logarithmically with the inverse of the diffusant radius (Saffman and Delbrück, 1975Saffman P.G. Delbrück M. Brownian motion in biological membranes.Proc. Natl. Acad. Sci. USA. 1975; 72: 3111-3113Crossref PubMed Scopus (1283) Google Scholar). This implies that slowing down diffusion through oligomerization of receptors is not enough to constrain mobility. For example, an oligomer of a hundred monomers will diffuse at only half the speed of a monomer. Therefore, diffusion of proteins must be contained in order to maintain spatial memory with micrometer precision over a timescale of minutes. Single-molecule experiments have shown that the plasma membrane is partitioned into 50–300 nm wide domains by the combined action of actin-based membrane skeleton "fences" and anchored-transmembrane protein "pickets" (Kusumi and Sako, 1996Kusumi A. Sako Y. Cell surface organization by the membrane skeleton.Curr. Opin. Cell Biol. 1996; 8: 566-574Crossref PubMed Scopus (311) Google Scholar). Within these membrane domains, proteins and lipids are highly mobile, with nanoscopic diffusion coefficients in the order of those measured for model membranes (Kusumi et al., 2005Kusumi A. Nakada C. Ritchie K. Murase K. Suzuki K. Murakoshi H. Kasai R.S. Kondo J. Fujiwara T. Paradigm shift of the plasma membrane concept from the two-dimensional continuum fluid to the partitioned fluid: high-speed single-molecule tracking of membrane molecules.Annu. Rev. Biophys. Biomol. Struct. 2005; 34: 351-378Crossref PubMed Scopus (865) Google Scholar). The hopping of signaling proteins across the fences occurs with low probability and thereby becomes the rate-limiting factor in lateral information transfer. In contrast to diffusion, the hopping rate is strongly dependent on the size of proteins, and therefore ligand-induced oligomerization of activated receptors traps the signal within these domains (Nelson et al., 1999Nelson S. Horvat R.D. Malvey J. Roess D.A. Barisas B.G. Clay C.M. Characterization of an intrinsically fluorescent gonadotropin-releasing hormone receptor and effects of ligand binding on receptor lateral diffusion.Endocrinology. 1999; 140: 950-957Crossref PubMed Scopus (45) Google Scholar). Such oligomerization-induced trapping thus provides a mechanism for the maintenance of spatial memory. Conversely, the confinement of monomers due to their low hopping rate facilitates oligomerization within domains. These domains can be considered as well-mixed protein reaction vessels because the time needed to diffuse through them is two orders of magnitude smaller (150 μs) than the residence time within them (15 ms) (Kusumi et al., 2005Kusumi A. Nakada C. Ritchie K. Murase K. Suzuki K. Murakoshi H. Kasai R.S. Kondo J. Fujiwara T. Paradigm shift of the plasma membrane concept from the two-dimensional continuum fluid to the partitioned fluid: high-speed single-molecule tracking of membrane molecules.Annu. Rev. Biophys. Biomol. Struct. 2005; 34: 351-378Crossref PubMed Scopus (865) Google Scholar). Compartmentalization therefore increases the rate of interaction between receptors. This has important implications for proteins such as the epidermal growth factor receptor (EGFR), which can oligomerize even in the absence of ligand. In cells expressing moderate numbers of EGFR molecules such as BAF/3 or COS7 (5 × 104 receptors/cell), the number of receptors per membrane domain, and therefore the degree of receptor clustering, will be low (2–3 receptors per cluster, consistent with the observations of Clayton et al., 2005Clayton A.H. Walker F. Orchard S.G. Henderson C. Fuchs D. Rothacker J. Nice E.C. Burgess A.W. Ligand-induced dimer-tetramer transition during the activation of the cell surface epidermal growth factor receptor-A multidimensional microscopy analysis.J. Biol. Chem. 2005; 280: 30392-30399Crossref PubMed Scopus (195) Google Scholar). However, in cancer cell lines that express abnormally high levels of EGFR such as A431 (2 × 106 receptors/cell), the average size of transient clusters will be much higher (10–15 receptors per cluster, consistent with the observation of Zidovetzki et al., 1981Zidovetzki R. Yarden Y. Schlessinger J. Jovin T.M. Rotational diffusion of epidermal growth factor complexed to cell surface receptors reflects rapid microaggregation and endocytosis of occupied receptors.Proc. Natl. Acad. Sci. USA. 1981; 78: 6981-6985Crossref PubMed Scopus (112) Google Scholar) (Figure 1A ). These preformed clusters have a profound impact on the propagation of receptor signals, as they spatially modulate the basal activity and reactivity of the plasma membrane. On a mechanistic level, the transmission of signals by receptor tyrosine kinases (RTKs) such as EGFR is relatively well understood. Binding of the cognate ligand to a receptor promotes their dimerization and thereby enables their phosphorylation in trans via their intrinsic tyrosine kinase activity (Lemmon and Schlessinger, 2010Lemmon M.A. Schlessinger J. Cell signaling by receptor tyrosine kinases.Cell. 2010; 141: 1117-1134Abstract Full Text Full Text PDF PubMed Scopus (2656) Google Scholar). The resulting phosphorylated tyrosine residues, exposed to the cytoplasm, act as docking sites for proteins that contain specialized domains, such as SH2 or PTB domains (Lim and Pawson, 2010Lim W.A. Pawson T. Phosphotyrosine signaling: evolving a new cellular communication system.Cell. 2010; 142: 661-667Abstract Full Text Full Text PDF PubMed Scopus (226) Google Scholar). Their recruitment induces allosteric changes in enzymatic activity or binding affinity on another module of the docked molecule, conveying signals deeper into the cytoplasm (Deribe et al., 2010Deribe Y.L. Pawson T. Dikic I. Post-translational modifications in signal integration.Nat. Struct. Mol. Biol. 2010; 17: 666-672Crossref PubMed Scopus (435) Google Scholar). However, though these sequences of reaction events provide insight into how signals are transferred across the membrane into the cell, they do not provide information on how these signals are regulated in space and time. To achieve this, we must consider the collective behavior of the ensemble of signaling molecules in the plasma membrane that have an influence on receptor phosphorylation state. Even at a low hopping rate of clustered receptors, the basal kinase activity of RTKs will eventually result in their full phosphorylation in the absence of a countering phosphatase activity (Reynolds et al., 2003Reynolds A.R. Tischer C. Verveer P.J. Rocks O. Bastiaens P.I. EGFR activation coupled to inhibition of tyrosine phosphatases causes lateral signal propagation.Nat. Cell Biol. 2003; 5: 447-453Crossref PubMed Scopus (178) Google Scholar). The degree of receptor phosphorylation within the plasma membrane is therefore determined by a continuous cycle of phosphorylation and dephosphorylation. Growth factor binding increases the amount of phosphorylated receptors by shifting the kinase-phosphatase balance in favor of the kinase. Though membrane-tethered and cytosolic proteins that are activated by receptors but not confined to the domains might propagate signals, their rather slow microscopic diffusion is incompatible with the timescales of minutes observed for such phenomena (Tischer and Bastiaens, 2003Tischer C. Bastiaens P.I. Lateral phosphorylation propagation: an aspect of feedback signalling?.Nat. Rev. Mol. Cell Biol. 2003; 4: 971-974Crossref PubMed Scopus (25) Google Scholar). Small molecule second messengers, such as calcium or reactive oxygen species (ROS), like hydrogen peroxide, have much larger diffusion coefficients, thereby propagating information via diffusion more quickly. Previously seen as a reaction by-product that causes oxidative stress, hydrogen peroxide has gained increasing interest as a mediator in signaling (Rhee, 2006Rhee S.G. Cell signaling. H2O2, a necessary evil for cell signaling.Science. 2006; 312: 1882-1883Crossref PubMed Scopus (1540) Google Scholar). Hydrogen peroxide is produced from the dismutation of superoxide generated by enzyme systems such as NAPDH oxidase (NOX) (Brown and Griendling, 2009Brown D.I. Griendling K.K. Nox proteins in signal transduction.Free Radic. Biol. Med. 2009; 47: 1239-1253Crossref PubMed Scopus (623) Google Scholar). Seven NOX catalytic subunits have been identified (NOX1, NOX2, NOX3, NOX4, NOX5, DUOX1, and DUOX2) that generate superoxide by transferring an electron from NADPH to molecular oxygen. The best-characterized NADPH oxidase, phagocytic NOX2, is a multisubunit enzyme complex with both transmembrane and cytosolic components. Upon stimulation, the cytosolic subunits are translocated to the membrane to bind the membrane-associated components, leading to activation of the NADPH oxidase complex. This activation process is triggered by growth factor receptor activation through the phosphorylation of two cytoplasmic subunits, P47PHOX and P67PHOX, and the conversion of GDP-bound RAC1 into GTP-bound forms through the activation of a RAC guanine nucleotide exchange factor (GEF) (Finkel, 2006Finkel T. Intracellular redox regulation by the family of small GTPases.Antioxid. Redox Signal. 2006; 8: 1857-1863Crossref PubMed Scopus (51) Google Scholar). RAC GEFs such as βPIX are recruited via their pleckstrin homology domain, and the resulting increase in RAC activity is presumed to stimulate NOX directly (Finkel, 2006Finkel T. Intracellular redox regulation by the family of small GTPases.Antioxid. Redox Signal. 2006; 8: 1857-1863Crossref PubMed Scopus (51) Google Scholar). Importantly, NOX enzymes produce superoxide on the outer leaflet of the plasma membrane, after which it dismutates to hydrogen peroxide and diffuses back into the cell (Rhee, 2006Rhee S.G. Cell signaling. H2O2, a necessary evil for cell signaling.Science. 2006; 312: 1882-1883Crossref PubMed Scopus (1540) Google Scholar). Hydrogen peroxide has been shown to inactivate PTPs such as PTP1B by reversible oxidation of a reactive cysteine in the catalytic cleft (Janssen-Heininger et al., 2008Janssen-Heininger Y.M. Mossman B.T. Heintz N.H. Forman H.J. Kalyanaraman B. Finkel T. Stamler J.S. Rhee S.G. van der Vliet A. Redox-based regulation of signal transduction: principles, pitfalls, and promises.Free Radic. Biol. Med. 2008; 45: 1-17Crossref PubMed Scopus (585) Google Scholar). The hydrogen peroxide-mediated coupling of RTK activation with PTP inhibition (Lee et al., 1998Lee S.R. Kwon K.S. Kim S.R. Rhee S.G. Reversible inactivation of protein-tyrosine phosphatase 1B in A431 cells stimulated with epidermal growth factor.J. Biol. Chem. 1998; 273: 15366-15372Crossref PubMed Scopus (812) Google Scholar) therefore exemplifies a double-negative feedback loop, which together with the autocatalytic kinase activity of the receptor, results in a bistable system (Reynolds et al., 2003Reynolds A.R. Tischer C. Verveer P.J. Rocks O. Bastiaens P.I. EGFR activation coupled to inhibition of tyrosine phosphatases causes lateral signal propagation.Nat. Cell Biol. 2003; 5: 447-453Crossref PubMed Scopus (178) Google Scholar, Tischer and Bastiaens, 2003Tischer C. Bastiaens P.I. Lateral phosphorylation propagation: an aspect of feedback signalling?.Nat. Rev. Mol. Cell Biol. 2003; 4: 971-974Crossref PubMed Scopus (25) Google Scholar). This reaction network effectively operates in a nanoenvironment that is local to the activated receptor due to the short half-life of intracellular ROS, which are the target of very efficient antioxidant enzymes such as peroxiredoxin I (PRXI) (Woo et al., 2010Woo H.A. Yim S.H. Shin D.H. Kang D. Yu D.Y. Rhee S.G. Inactivation of peroxiredoxin I by phosphorylation allows localized H(2)O(2) accumulation for cell signaling.Cell. 2010; 140: 517-528Abstract Full Text Full Text PDF PubMed Scopus (466) Google Scholar). Although spatially constrained, the presence of ROS can still lower the excitation threshold of neighboring, inactive receptors. Receptor density then becomes the key to trigger a domino-like rapid propagation of activity at long range, whereby the RTK/PTP/H2O2 system acts as an excitable medium (Figure 1B) (Reynolds et al., 2003Reynolds A.R. Tischer C. Verveer P.J. Rocks O. Bastiaens P.I. EGFR activation coupled to inhibition of tyrosine phosphatases causes lateral signal propagation.Nat. Cell Biol. 2003; 5: 447-453Crossref PubMed Scopus (178) Google Scholar). This global activation initiated by a local source is only possible due to the tight coupling between reaction components that have opposing activities. Insight into the nature of such coupling is required to predict the spatial outcome of reaction diffusion systems. Consider the case of activated, phosphorylated RTKs that can activate their own inhibitors such as the PTP SHP1. Here, phosphotyrosines on the activated RTK bind SHP1 via its SH2 domain, which allosterically activates the phosphatase. Phosphorylation of SHP1 by the RTK then locks it into the active state, irrespective of binding to the RTK (Frank et al., 2004Frank C. Burkhardt C. Imhof D. Ringel J. Zschörnig O. Wieligmann K. Zacharias M. Böhmer F.D. Effective dephosphorylation of Src substrates by SHP-1.J. Biol. 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In addition to cell-wide and submicroscopic domains with lifetimes ranging from days to seconds, even smaller (nanometer scale), shorter-lived (subsecond) domains have been proposed to transiently confine membrane proteins (Simons and Ikonen, 1997Simons K. Ikonen E. Functional rafts in cell membranes.Nature. 1997; 387: 569-572Crossref PubMed Scopus (7749) Google Scholar). These high-viscosity patches composed of cholesterol and glycosphingolipid are known as lipid rafts and have been shown to have an important role as labile platforms to which signaling components are recruited, favoring their interaction (Harding and Hancock, 2008Harding A.S. Hancock J.F. Using plasma membrane nanoclusters to build better signaling circuits.Trends Cell Biol. 2008; 18: 364-371Abstract Full Text Full Text PDF PubMed Scopus (90) Google Scholar). We refer the reader to some excellent recent reviews for more information about this extensive topic (Lingwood and Simons, 2010Lingwood D. Simons K. Lipid rafts as a membrane-organizing principle.Science. 2010; 327: 46-50Crossref PubMed Scopus (2895) Google Scholar) that goes beyond the scope of this Review. Axial signal propagation into the cytoplasm by phosphorylation of soluble substrates, like lateral signal propagation, is also tightly controlled by reaction-diffusion systems that generate a local environment of activated substrates. For example, transfer of growth factor signals from RTKs in the plasma membrane to soluble substrates in the cytoplasm also depends on cyclic reaction-diffusion systems of opposing tyrosine kinase/phosphatase activities. However, the catalytic activity of fully active PTPs is up to three orders of magnitude higher than that of tyrosine kinases (Fischer et al., 1991Fischer E.H. Charbonneau H. Tonks N.K. Protein tyrosine phosphatases: a diverse family of intracellular and transmembrane enzymes.Science. 1991; 253: 401-406Crossref PubMed Scopus (851) Google Scholar), which would preclude the effective transfer of growth factor signals via phosphorylation in the cytoplasm. On the other hand, the absence of PTP activity near the plasma membrane would allow spurious signals to be transmitted in the cell. The solution to this dilemma is the membrane-proximal, partial inactivation of PTPs by oxidation of the catalytic cysteine with hydrogen peroxide that is produced by NOX as outlined above. The reducing activity of the cytoplasm (sink) together with the source of hydrogen peroxide production at the plasma membrane generates a hydrogen peroxide gradient in the cytoplasm in which PTP activity is strongly reduced near the membrane. Thus, signal penetration via tyrosine phosphorylation is ultimately a self-referencing system in which tyrosine phosphorylation depends on the magnitude of the hydrogen peroxide gradient, which in turn depends on the balance between RTK and PTP activities. The extent of feedback in this system became even more apparent with the recent identification of PRXI as a major reducing agent that controls hydrogen peroxide levels in the cytoplasm (Woo et al., 2010Woo H.A. Yim S.H. Shin D.H. Kang D. Yu D.Y. Rhee S.G. Inactivation of peroxiredoxin I by phosphorylation allows localized H(2)O(2) accumulation for cell signaling.Cell. 2010; 140: 517-528Abstract Full Text Full Text PDF PubMed Scopus (466) Google Scholar). Importantly, its activity is inhibited by phosphorylation mediated by membrane-bound Src on tyrosine 194, thereby generating a local positive feedback loop around activated RTKs. We might the
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