scholarly journals A novel role for FGF and extracellular signal–regulated kinase in gap junction–mediated intercellular communication in the lens

2001 ◽  
Vol 154 (1) ◽  
pp. 197-216 ◽  
Author(s):  
Anh-Chi N. Le ◽  
Linda S. Musil
Keyword(s):  
Growth Factor ◽  
Gap Junction ◽  
Intercellular Communication ◽  
Equatorial Region ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Junctional Coupling ◽  
Gap Junctional ◽  
Gap Junctional Coupling

Gap junction–mediated intercellular coupling is higher in the equatorial region of the lens than at either pole, a property believed to be essential for lens transparency. We show that fibroblast growth factor (FGF) upregulates gap junctional intercellular dye transfer in primary cultures of embryonic chick lens cells without detectably increasing either gap junction protein (connexin) synthesis or assembly. Insulin and insulin-like growth factor 1, as potent as FGF in inducing lens cell differentiation, had no effect on gap junctions. FGF induced sustained activation of extracellular signal–regulated kinase (ERK) in lens cells, an event necessary and sufficient to increase gap junctional coupling. We also identify vitreous humor as an in vivo source of an FGF-like intercellular communication-promoting activity and show that FGF-induced ERK activation in the intact lens is higher in the equatorial region than in polar and core fibers. These findings support a model in which regional differences in FGF signaling through the ERK pathway lead to the asymmetry in gap junctional coupling required for proper lens function. Our results also identify upregulation of intercellular communication as a new function for sustained ERK activation and change the current paradigm that ERKs only negatively regulate gap junction channel activity.

scholarly journals Role of Phosphoinositide 3-Kinase and Endocytosis in Nerve Growth Factor-Induced Extracellular Signal-Regulated Kinase Activation via Ras and Rap1

2000 ◽  
Vol 20 (21) ◽  
pp. 8069-8083 ◽  
Author(s):  
Randall D. York ◽  
Derek C. Molliver ◽  
Savraj S. Grewal ◽  
Paula E. Stenberg ◽  
Edwin W. McCleskey ◽  
...  
Keyword(s):  
Nerve Growth Factor ◽  
Growth Factor ◽  
Map Kinase ◽  
Pc12 Cells ◽  
Erk Signaling ◽  
Nerve Growth ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Phosphoinositide 3 Kinase

ABSTRACT Neurotrophins promote multiple actions on neuronal cells including cell survival and differentiation. The best-studied neurotrophin, nerve growth factor (NGF), is a major survival factor in sympathetic and sensory neurons and promotes differentiation in a well-studied model system, PC12 cells. To mediate these actions, NGF binds to the TrkA receptor to trigger intracellular signaling cascades. Two kinases whose activities mediate these processes include the mitogen-activated protein (MAP) kinase (or extracellular signal-regulated kinase [ERK]) and phosphoinositide 3-kinase (PI3-K). To examine potential interactions between the ERK and PI3-K pathways, we studied the requirement of PI3-K for NGF activation of the ERK signaling cascade in dorsal root ganglion cells and PC12 cells. We show that PI3-K is required for TrkA internalization and participates in NGF signaling to ERKs via distinct actions on the small G proteins Ras and Rap1. In PC12 cells, NGF activates Ras and Rap1 to elicit the rapid and sustained activation of ERKs respectively. We show here that Rap1 activation requires both TrkA internalization and PI3-K, whereas Ras activation requires neither TrkA internalization nor PI3-K. Both inhibitors of PI3-K and inhibitors of endocytosis prevent GTP loading of Rap1 and block sustained ERK activation by NGF. PI3-K and endocytosis may also regulate ERK signaling at a second site downstream of Ras, since both rapid ERK activation and the Ras-dependent activation of the MAP kinase kinase kinase B-Raf are blocked by inhibition of either PI3-K or endocytosis. The results of this study suggest that PI3-K may be required for the signals initiated by TrkA internalization and demonstrate that specific endocytic events may distinguish ERK signaling via Rap1 and Ras.

Regulation of MEK/ERK pathway output by subcellular localization of B-Raf

2012 ◽  
Vol 40 (1) ◽  
pp. 67-72 ◽  
Author(s):  
Catherine Andreadi ◽  
Catherine Noble ◽  
Bipin Patel ◽  
Hong Jin ◽  
Maria M. Aguilar Hernandez ◽  
...  
Keyword(s):  
Growth Factor ◽  
Subcellular Localization ◽  
Signalling Pathway ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Intracellular Signalling Pathway ◽  
Pathway Activation ◽  
Extracellular Signal ◽  
A Cell ◽  
Phase Entry

The strength and duration of intracellular signalling pathway activation is a key determinant of the biological outcome of cells in response to extracellular cues. This has been particularly elucidated for the Ras/Raf/MEK [mitogen-activated growth factor/ERK (extracellular-signal-regulated kinase) kinase]/ERK signalling pathway with a number of studies in fibroblasts showing that sustained ERK signalling is a requirement for S-phase entry, whereas transient ERK signalling does not have this capability. A major unanswered question, however, is how a cell can sustain ERK activation, particularly when ERK-specific phosphatases are transcriptionally up-regulated by the pathway itself. A major point of ERK regulation is at the level of Raf, and, to sustain ERK activation in the presence of ERK phosphatases, sustained Raf activation is a requirement. Three Raf proteins exist in mammals, and the activity of all three is induced following growth factor stimulation of cells, but only B-Raf activity is maintained at later time points. This observation points to B-Raf as a regulator of sustained ERK activation. In the present review, we consider evidence for a link between B-Raf and sustained ERK activation, focusing on a potential role for the subcellular localization of B-Raf in this key physiological event.

scholarly journals The Tyrosine Phosphatase SHP-2 Is Required for Sustained Activation of Extracellular Signal-Regulated Kinase and Epithelial Morphogenesis Downstream from the Met Receptor Tyrosine Kinase

2000 ◽  
Vol 20 (22) ◽  
pp. 8513-8525 ◽  
Author(s):  
Christiane R. Maroun ◽  
Monica A. Naujokas ◽  
Marina Holgado-Madruga ◽  
Albert J. Wong ◽  
Morag Park
Keyword(s):  
Growth Factor ◽  
Tyrosine Phosphatase ◽  
Epithelial Morphogenesis ◽  
Met Receptor ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Transient Activation ◽  
Extracellular Signal ◽  
Positive Modulator ◽  
Sustained Activation

ABSTRACT Epithelial morphogenesis is critical during development and wound healing, and alterations in this program contribute to neoplasia. Met, the hepatocyte growth factor (HGF) receptor, promotes a morphogenic program in epithelial cell lines in matrix cultures. Previous studies have identified Gab1, the major phosphorylated protein following Met activation, as important for the morphogenic response. Gab1 is a docking protein that couples the Met receptor with multiple signaling proteins, including phosphatidylinositol-3 kinase, phospholipase Cγ, the adapter protein Crk, and the tyrosine specific phosphatase SHP-2. HGF induces sustained phosphorylation of Gab1 and sustained activation of extracellular signal-regulated kinase (Erk) in epithelial Madin-Darby canine kidney cells. In contrast, epidermal growth factor fails to promote a morphogenic program and induces transient Gab1 phosphorylation and Erk activation. To elucidate the Gab1-dependent signals required for epithelial morphogenesis, we undertook a structure-function approach and demonstrate that association of Gab1 with the tyrosine phosphatase SHP-2 is required for sustained Erk activation and for epithelial morphogenesis downstream from the Met receptor. Epithelial cells expressing a Gab1 mutant protein unable to recruit SHP-2 elicit a transient activation of Erk in response to HGF. Moreover, SHP-2 catalytic activity is required, since the expression of a catalytically inactive SHP-2 mutant, C/S, abrogates sustained activation of Erk and epithelial morphogenesis by the Met receptor. These data identify SHP-2 as a positive modulator of Erk activity and epithelial morphogenesis downstream from the Met receptor.

scholarly journals Gap junctional coupling and connexin immunoreactivity in rabbit retinal glia

2006 ◽  
Vol 23 (1) ◽  
pp. 1-10 ◽  
Author(s):  
KATHLEEN R. ZAHS ◽  
PAUL W. CEELEN
Keyword(s):  
Gap Junctions ◽  
Gap Junction ◽  
Lucifer Yellow ◽  
Müller Cells ◽  
Müller Cell ◽  
Muller Cells ◽  
Junctional Coupling ◽  
Muller Cell ◽  
Gap Junctional ◽  
Gap Junctional Coupling

Gap junctions provide a pathway for the direct intercellular exchange of ions and small signaling molecules. Gap junctional coupling between retinal astrocytes and between astrocytes and Müller cells, the principal glia of vertebrate retinas, has been previously demonstrated by the intercellular transfer of gap-junction permeant tracers. However, functional gap junctions have yet to be demonstrated between mammalian Müller cells. In the present study, when the gap-junction permeant tracers Neurobiotin and Lucifer yellow were injected into a Müller cellviaa patch pipette, the tracers transferred to at least one additional cell in more than half of the cases examined. Simultaneous whole-cell recordings from pairs of Müller cells in the isolated rabbit retina revealed electrical coupling between closely neighboring cells, confirming the presence of functional gap junctions between rabbit Müller cells. The limited degree of this coupling suggests that Müller cell–Müller cell gap junctions may coordinate the functions of small ensembles of these glial cells. Immunohistochemistry and immunoblotting were used to identify the connexins in rabbit retinal glia. Connexin30 (Cx30) and connexin43 (Cx43) immunoreactivities were associated with astrocytes in the medullary ray region of the retinas of both pigmented and albino rabbits. Connexin43 was also found in Müller cells, but antibody recognition differed between astrocytic and Müller cell connexin43.

scholarly journals GP78 Cooperates with Dual-Specificity Phosphatase 1 To Stimulate Epidermal Growth Factor Receptor-Mediated Extracellular Signal-Regulated Kinase Signaling

2019 ◽  
Vol 39 (11) ◽  
Author(s):  
Dhong Hyo Kho ◽  
Mohammed Hafiz Uddin ◽  
Madhumita Chatterjee ◽  
Andreas Vogt ◽  
Avraham Raz ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Growth Factor Receptor ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Dual Specificity ◽  
Extracellular Signal ◽  
Epidermal Growth ◽  
Proliferation And Invasion

ABSTRACT GP78 is an autocrine motility factor (AMF) receptor (AMFR) with E3 ubiquitin ligase activity that plays a significant role in tumor cell proliferation, motility, and metastasis. Aberrant extracellular signal-regulated kinase (ERK) activation via receptor tyrosine kinases promotes tumor proliferation and invasion. The activation of GP78 leads to ERK activation, but its underlying mechanism is not fully understood. Here, we show that GP78 is required for epidermal growth factor receptor (EGFR)-mediated ERK activation. On one hand, GP78 interacts with and promotes the ubiquitination and subsequent degradation of dual-specificity phosphatase 1 (DUSP1), an endogenous negative regulator of mitogen-activated protein kinases (MAPKs), resulting in ERK activation. On the other hand, GP78 maintains the activation status of EGFR, as evidenced by the fact that EGF fails to induce EGFR phosphorylation in GP78-deficient cells. By the regulation of both EGFR and ERK activation, GP78 promotes cell proliferation, motility, and invasion. Therefore, this study identifies a previously unknown signaling pathway by which GP78 stimulates ERK activation via DUSP1 degradation to mediate EGFR-dependent cancer cell proliferation and invasion.

scholarly journals Signal Transmission Between Gap-Junctionally Coupled Passive Cables Is Most Effective at an Optimal Diameter

2006 ◽  
Vol 95 (6) ◽  
pp. 3831-3843 ◽  
Author(s):  
Farzan Nadim ◽  
Jorge Golowasch
Keyword(s):  
Gap Junction ◽  
Current Flow ◽  
Signal Transmission ◽  
Electrical Signal ◽  
Neuronal Processes ◽  
Junctional Coupling ◽  
Length Constant ◽  
Gap Junctional ◽  
Gap Junctional Coupling ◽  
Optimal Diameter

We analyze simple morphological configurations that represent gap-junctional coupling between neuronal processes or between muscle fibers. Specifically, we use cable theory and simulations to examine the consequences of current flow from one cable to other gap-junctionally coupled passive cables. When the proximal end of the first cable is voltage clamped, the amplitude of the electrical signal in distal portions of the second cable depends on the cable diameter. However, this amplitude does not simply increase if cable diameter is increased, as expected from the larger length constant; instead, an optimal diameter exists. The optimal diameter arises because the dependency of voltage attenuation along the second cable on cable diameter follows two opposing rules. As cable diameter increases, the attenuation decreases because of a larger length constant yet increases because of a reduction in current density arising from the limiting effect of the gap junction on current flow into the second cable. The optimal diameter depends on the gap junction resistance and cable parameters. In branched cables, dependency on diameter is local and thus may serve to functionally compartmentalize branches that are coupled to other cells. Such compartmentalization may be important when periodic signals or action potentials cause the current flow across gap junctions.

Normal development of preimplantation mouse embryos deficient in gap junctional coupling

1997 ◽  
Vol 110 (15) ◽  
pp. 1751-1758 ◽  
Author(s):  
P.A. De Sousa ◽  
S.C. Juneja ◽  
S. Caveney ◽  
F.D. Houghton ◽  
T.C. Davies ◽  
...  
Keyword(s):  
Gap Junction ◽  
Preimplantation Development ◽  
Null Mutation ◽  
Dye Coupling ◽  
Permeability Characteristics ◽  
Gap Junction Channels ◽  
Junctional Coupling ◽  
Preimplantation Mouse ◽  
Gap Junctional ◽  
Gap Junctional Coupling

The connexin multigene family (13 characterized members in rodents) encodes the subunits of gap junction channels. Gap junctional intercellular coupling, established during compaction of the preimplantation mouse embryo, is assumed to be necessary for development of the blastocyst. One member of the connexin family, connexin43, has been shown to contribute to the gap junctions that form during compaction, yet embryos homozygous for a connexin43 null mutation develop normally, at least until implantation. We show that this can be explained by contributions from one or more additional connexin genes that are normally expressed along with connexin43 in preimplantation development. Immunogold electron microscopy confirmed that roughly 30% of gap junctions in compacted morulae contain little or no connexin43 and therefore are likely to be composed of another connexin(s). Confocal immunofluorescence microscopy was then used to demonstrate that connexin45 is also assembled into membrane plaques, beginning at the time of compaction. Correspondingly, embryos homozygous for the connexin43 null mutation were found to retain the capacity for cell-to-cell transfer of fluorescent dye (dye coupling), but at a severely reduced level and with altered permeability characteristics. Whereas mutant morulae showed no evidence of dye coupling when tested with 6-carboxyfluorescein, dye coupling could be demonstrated using 2′,7′-dichlorofluorescein, revealing permeability characteristics previously established for connexin45 channels. We conclude that preimplantation development in the mouse can proceed normally even though both the extent and nature of gap junctional coupling have been perturbed. Despite the distinctive properties of connexin43 channels, their role in preimplantation development can be fulfilled by one or more other types of gap junction channels.

scholarly journals Raf-1 Serine 338 Phosphorylation Plays a Key Role in Adhesion-Dependent Activation of Extracellular Signal-Regulated Kinase by Epidermal Growth Factor

2005 ◽  
Vol 25 (11) ◽  
pp. 4466-4475 ◽  
Author(s):  
Matthew L. Edin ◽  
Rudy L. Juliano
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Highly Active ◽  
Extracellular Signal ◽  
Tightly Coupled ◽  
Epidermal Growth ◽  
P21 Activated Kinase ◽  
Suspended Cells

ABSTRACT Activation of the extracellular signal-regulated kinase (ERK) 1/2 cascade by polypeptide growth factors is tightly coupled to adhesion to extracellular matrix in nontransformed cells. Raf-1, the initial kinase in this cascade, is intricately regulated by phosphorylation, localization, and molecular interactions. We investigated the complex interactions between Raf-1, protein kinase A (PKA), and p21-activated kinase (PAK) to determine their roles in the adhesion dependence of signaling from epidermal growth factor (EGF) to ERK. We conclude that Raf-1 phosphorylation on serine 338 (S338) is a critical step that is inhibited in suspended cells. Restoration of phosphorylation at S338, either by expression of highly active PAK or by expression of an S338 phospho-mimetic Raf-1 mutation, led to a partial rescue of ERK activation in suspended cells. Raf-1 inhibition in suspension was not due to excessive negative regulation on inhibitory sites S43 and S259, as these serines were largely dephosphorylated in suspended cells. Finally, strong phosphorylation of Raf-1 S338 provided resistance to PKA-mediated inhibition of ERK activation. Phosphorylation at Raf-1 S43 and S259 by PKA only weakly inhibited EGF activation of Raf-1 and ERK when cells maintained high Raf-1 S338 phosphorylation.

scholarly journals Intercalated disk nanoscale structure regulates cardiac conduction

2021 ◽  
Vol 153 (8) ◽  
Author(s):  
Nicolae Moise ◽  
Heather L. Struckman ◽  
Celine Dagher ◽  
Rengasayee Veeraraghavan ◽  
Seth H. Weinberg
Keyword(s):  
Gap Junction ◽  
Cardiac Tissue ◽  
Element Model ◽  
Active Role ◽  
Cardiac Conduction ◽  
Junctional Coupling ◽  
Nanoscale Structure ◽  
Gap Junctional ◽  
Gap Junctional Coupling ◽  
Intercalated Disk

The intercalated disk (ID) is a specialized subcellular region that provides electrical and mechanical connections between myocytes in the heart. The ID has a clearly defined passive role in cardiac tissue, transmitting mechanical forces and electrical currents between cells. Recent studies have shown that Na+ channels, the primary current responsible for cardiac excitation, are preferentially localized at the ID, particularly within nanodomains such as the gap junction–adjacent perinexus and mechanical junction–associated adhesion-excitability nodes, and that perturbations of ID structure alter cardiac conduction. This suggests that the ID may play an important, active role in regulating conduction. However, the structures of the ID and intercellular cleft are not well characterized and, to date, no models have incorporated the influence of ID structure on conduction in cardiac tissue. In this study, we developed an approach to generate realistic finite element model (FEM) meshes replicating nanoscale of the ID structure, based on experimental measurements from transmission electron microscopy images. We then integrated measurements of the intercellular cleft electrical conductivity, derived from the FEM meshes, into a novel cardiac tissue model formulation. FEM-based calculations predict that the distribution of cleft conductances is sensitive to regional changes in ID structure, specifically the intermembrane separation and gap junction distribution. Tissue-scale simulations predict that ID structural heterogeneity leads to significant spatial variation in electrical polarization within the intercellular cleft. Importantly, we found that this heterogeneous cleft polarization regulates conduction by desynchronizing the activation of postjunctional Na+ currents. Additionally, these heterogeneities lead to a weaker dependence of conduction velocity on gap junctional coupling, compared with prior modeling formulations that neglect or simplify ID structure. Further, we found that disruption of local ID nanodomains can either slow or enhance conduction, depending on gap junctional coupling strength. Our study therefore suggests that ID nanoscale structure can play a significant role in regulating cardiac conduction.

scholarly journals Temporal control of gap junction assembly in preimplantation mouse embryos

1995 ◽  
Vol 108 (4) ◽  
pp. 1715-1722 ◽  
Author(s):  
G. Valdimarsson ◽  
G.M. Kidder
Keyword(s):  
Dna Replication ◽  
Dna Synthesis ◽  
Gap Junction ◽  
Continuous Treatment ◽  
Mouse Development ◽  
Cell Stage ◽  
Cell Cycles ◽  
Junctional Coupling ◽  
Gap Junctional ◽  
Gap Junctional Coupling

The de novo assembly of gap junctions during compaction in the 8-cell stage of mouse development is a temporally regulated event. We have performed experiments designed to explore the relationship between this event and DNA replication in the second, third, and fourth cell cycles after fertilization. Inhibition of DNA synthesis by continuous treatment with the DNA synthesis inhibitor, aphidicolin, during the third and fourth cell cycles had no effect on the establishment of gap junctional coupling during compaction. However, a delay of 10 hours in DNA synthesis during the second cell cycle caused by a transient aphidicolin treatment resulted in the failure of gap junctional coupling at the time of compaction. Thus the timing of establishment of gap junctional coupling, like the timing of compaction itself, is linked to DNA replication in the 2-cell stage. Immunofluorescence analysis showed that the failure of gap junctional coupling after aphidicolin treatment in the 2-cell stage is correlated with the failure of nascent connexin43 to be inserted into plasma membranes. We propose that the developmental ‘clock’ that controls gap junction assembly is set in motion by events surrounding the second cycle of DNA replication, and that this ‘clock’ ultimately controls the post-translational processing of connexin43.

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