AMP-activated protein kinase induces actin cytoskeleton reorganization in epithelial cells

2010 ◽  
Vol 396 (3) ◽  
pp. 656-661 ◽  
Author(s):  
Lisa Miranda ◽  
Sarah Carpentier ◽  
Anna Platek ◽  
Nusrat Hussain ◽  
Marie-Agnès Gueuning ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Actin Cytoskeleton ◽  
Cytoskeleton Reorganization ◽  
Amp Activated Protein Kinase

scholarly journals AMP-activated Protein Kinase Impairs Endothelial Actin Cytoskeleton Assembly by Phosphorylating Vasodilator-stimulated Phosphoprotein

2006 ◽  
Vol 282 (7) ◽  
pp. 4601-4612 ◽  
Author(s):  
Constanze Blume ◽  
Peter M. Benz ◽  
Ulrich Walter ◽  
Joohun Ha ◽  
Bruce E. Kemp ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Actin Cytoskeleton ◽  
Energy Homeostasis ◽  
Serum Response Factor ◽  
Regulatory Protein ◽  
Fiber Formation ◽  
Actin Stress Fiber ◽  
Zdf Rat ◽  
Vasp Phosphorylation ◽  
Amp Activated Protein Kinase

Vasodilator-stimulated phosphoprotein (VASP) is an actin regulatory protein that links signaling pathways to remodeling of the cytoskeleton. VASP functions are modulated by protein kinases, which phosphorylate the sites Ser-157, Ser-239, and Thr-278. The kinase responsible for Thr-278 phosphorylation, biological functions of the phosphorylation, and association with disease states have remained enigmatic. Using VASP phosphorylation status-specific antibodies, we identified AMP-activated protein kinase (AMPK), a serine-threonine kinase and fundamental sensor of energy homeostasis, in a screen for kinases that phosphorylate the Thr-278 site of VASP in endothelial cells. Pharmacological AMPK inhibitors and activators and AMPK mutants revealed that the kinase specifically targets residue Thr-278 but not Ser-157 or Ser-239. Quantitative fluorescence-activated cell sorter analysis and serum response factor transcriptional reporter assays, which quantify the cellular F-/G-actin equilibrium, indicated that AMPK-mediated VASP phosphorylation impaired actin stress fiber formation and altered cell morphology. In the Zucker Diabetic Fatty (ZDF) rat model for type II diabetes, AMPK activity and Thr-278 phosphorylation were substantially reduced in arterial vessel walls. These findings suggest that VASP is a new AMPK substrate, that VASP Thr-278 phosphorylation translates metabolic signals into actin cytoskeleton rearrangements, and that this signaling system becomes down-regulated in diabetic vessels.

TGF beta 1 promotes actin cytoskeleton reorganization and migratory phenotype in epithelial tracheal cells in primary culture

1996 ◽  
Vol 109 (9) ◽  
pp. 2207-2219 ◽  
Author(s):  
S. Boland ◽  
E. Boisvieux-Ulrich ◽  
O. Houcine ◽  
A. Baeza-Squiban ◽  
M. Pouchelet ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Cell Proliferation ◽  
Cell Migration ◽  
Epithelial Cells ◽  
Actin Cytoskeleton ◽  
Primary Culture ◽  
Tgf Beta ◽  
Cytoskeleton Reorganization ◽  
Extracellular Matrix Components ◽  
Migratory Phenotype

In the present study we have investigated the effects of transforming growth factor beta (TGF beta 1) on rabbit tracheal epithelial cells in primary culture, with respect to cell proliferation and differentiation. Epithelial tracheal cells derived from an explant plated on an extracellular matrix, formed an outgrowth resulting from cell division and cell migration. TGF beta 1 treatment produced a negative effect on cell proliferation, but in contrast, promoted a marked enhancement of cell migration and increase in outgrowth surface. TGF beta 1 induced marked cell shape changes, including cell spreading and lack of stratification, associated with reduced cell-cell contacts and increased cell-substratum anchorage, as seen by electron microscopic observations. Immunocytological studies demonstrated major TGF beta 1-induced actin cytoskeleton reorganization, corresponding to the development of a basal stress fiber network and decrease of the annular cell border, without affecting the tight junctions. The migratory phenotype was approached by microcinematography which clearly showed that TGF beta 1 triggered a stimulatory effect on migration of epithelial cells, determined using an image analyzing system. Present findings suggest a beneficial role for TGF beta 1 during wound healing in providing the acquisition of a migratory phenotype, with a higher capacity to migrate either on collagen or on different extracellular matrix components including laminin and fibronectin. Conversely, present data are not consistent with a squamous response to TGF beta 1, since metaplastic differentiation did not occur, as characterized by cytokeratin expression and cross-linked envelopes formation.

scholarly journals α1-AMP-Activated Protein Kinase Regulates Hypoxia-Induced Na,K-ATPase Endocytosis via Direct Phosphorylation of Protein Kinase Cζ

2009 ◽  
Vol 29 (13) ◽  
pp. 3455-3464 ◽  
Author(s):  
Galina A. Gusarova ◽  
Laura A. Dada ◽  
Aileen M. Kelly ◽  
Chaya Brodie ◽  
Lee A. Witters ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
A549 Cells ◽  
Dominant Negative ◽  
Alveolar Epithelial Cells ◽  
Ampk Activation ◽  
Α Subunit ◽  
Alveolar Epithelial ◽  
Mitochondrial Ros ◽  
Amp Activated Protein Kinase

ABSTRACT Hypoxia promotes Na,K-ATPase endocytosis via protein kinase Cζ (PKCζ)-mediated phosphorylation of the Na,K-ATPase α subunit. Here, we report that hypoxia leads to the phosphorylation of 5′-AMP-activated protein kinase (AMPK) at Thr172 in rat alveolar epithelial cells. The overexpression of a dominant-negative AMPK α subunit (AMPK-DN) construct prevented the hypoxia-induced endocytosis of Na,K-ATPase. The overexpression of the reactive oxygen species (ROS) scavenger catalase prevented hypoxia-induced AMPK activation. Moreover, hypoxia failed to activate AMPK in mitochondrion-deficient ρ0-A549 cells, suggesting that mitochondrial ROS play an essential role in hypoxia-induced AMPK activation. Hypoxia-induced PKCζ translocation to the plasma membrane and phosphorylation at Thr410 were prevented by the pharmacological inhibition of AMPK or by the overexpression of the AMPK-DN construct. We found that AMPK α phosphorylates PKCζ on residue Thr410 within the PKCζ activation loop. Importantly, the activation of AMPK α was necessary for hypoxia-induced AMPK-PKCζ binding in alveolar epithelial cells. The overexpression of T410A mutant PKCζ prevented hypoxia-induced Na,K-ATPase endocytosis, confirming that PKCζ Thr410 phosphorylation is essential for this process. PKCζ activation by AMPK is isoform specific, as small interfering RNA targeting the α1 but not the α2 catalytic subunit prevented PKCζ activation. Accordingly, we provide the first evidence that hypoxia-generated mitochondrial ROS lead to the activation of the AMPK α1 isoform, which binds and directly phosphorylates PKCζ at Thr410, thereby promoting Na,K-ATPase endocytosis.

scholarly journals Regulation of Megalin membrane traffic by AMP‐activated protein kinase in kidney proximal tubule epithelial cells

2021 ◽  
Vol 35 (S1) ◽  
Author(s):  
Nikol Leshchyshyn ◽  
Laura Orofiamma ◽  
Christian Delos Santos ◽  
Sadia Rahmani ◽  
Costin Antonescu
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Proximal Tubule ◽  
Membrane Traffic ◽  
Kidney Proximal Tubule ◽  
Amp Activated Protein Kinase

scholarly journals Expression of Adiponectin Receptors and Its Possible Implication in the Human Endometrium

Endocrinology ◽  
2006 ◽  
Vol 147 (7) ◽  
pp. 3203-3210 ◽  
Author(s):  
Yuri Takemura ◽  
Yutaka Osuga ◽  
Toshimasa Yamauchi ◽  
Masaki Kobayashi ◽  
Miyuki Harada ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Protein Kinase ◽  
Epithelial Cells ◽  
Stromal Cells ◽  
Adiponectin Receptors ◽  
Monocyte Chemoattractant Protein 1 ◽  
Monocyte Chemoattractant ◽  
Monocyte Chemoattractant Protein ◽  
Amp Activated Protein Kinase

Adiponectin, a pleiotropic cytokine, exerts its effects via the specific receptors AdipoR1 and AdipoR2. Whereas circulating adiponectin concentrations decrease in women with endometriosis and endometrial cancer, possible effects of adiponectin and the presence of the receptors in the endometrium have not been determined. In this study, we examined the expression of adiponectin receptors AdipoR1 and AdipoR2 in the human endometrium and assessed effects of adiponectin in endometrial cells. Expression of AdipoR1 and AdipoR2 in endometrial tissues was evaluated by real-time quantitative PCR, in situ hybridization, and Western blotting. The effects of adiponectin on phosphorylation of AMP-activated protein kinase, a regulator of energy homeostasis, in cultured endometrial stromal cells (ESCs) and epithelial cells (EECs) were studied by Western blotting. The effects of adiponectin on IL-1β-induced secretion of IL-6, IL-8, and monocyte chemoattractant protein 1 from cultured ESCs were determined using specific ELISAs. The expression of AdipoR1 and AdipoR2 was detected in the endometrium. The expression of both genes was increased in the midluteal phase, the period of embryo implantation. In situ hybridization revealed that both AdipoR1 and AdipoR2 appeared to be equally expressed in the epithelial cells and in the stromal cells. Adiponectin increased phosphorylation of AMP-activated protein kinase in ESCs and EECs. Adiponectin decreased IL-1β-induced secretion of IL-6, IL-8, and monocyte chemoattractant protein 1 from ESCs. These findings suggest that adiponectin exerts energy-homeostatic and antiinflammatory effects in the endometrium, and these effects might be relevant to pathological and physiological endometrium-related events such as implantation and endometriosis.

Mechanical force-activated phospholipase D is mediated by Gα12/13-Rho and calmodulin-dependent kinase in renal epithelial cells

2005 ◽  
Vol 289 (4) ◽  
pp. F826-F834 ◽  
Author(s):  
Jenny Ziembicki ◽  
Rajnish Tandon ◽  
Jeffrey R. Schelling ◽  
John R. Sedor ◽  
R. Tyler Miller ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Actin Cytoskeleton ◽  
Signaling Pathways ◽  
Phospholipase D ◽  
Mdck Cells ◽  
Mechanical Force ◽  
Rgs Proteins ◽  
Filamentous Actin ◽  
Renal Epithelial Cells ◽  
Cytoskeleton Reorganization

The renal glomerulus, the site of plasma ultrafiltration, is exposed to mechanical force in vivo arising from capillary blood pressure and fluid flow. Studies of cultured podocytes demonstrate that they respond to stretch by altering the structure of the actin cytoskeleton, but the mechanisms by which physical force triggers this architectural change and the signaling pathways that lead to generation of second messengers are not defined. In the present study, we found that in renal epithelial cells [podocytes and Madin-Darby canine kidney (MDCK) cells], application of mechanical force to the cell surface through fibronectin-coated ferric beads and exposure of the cells to magnetic force lead to Rho translocation and actin cytoskeleton reorganization. This application of force recruited Rho and filamentous actin (F-actin) to bead loci and subsequently stimulated phospholipase D (PLD), a downstream effector of Rho. Using MDCK cells that stably express regulators of G protein-signaling (RGS) proteins [RGS4 attenuates Gαi and Gαq, and the p115RhoGEF-RGS domain (p115-RGS) attenuates Gα12/13] to define the signaling pathway, we found that mechanical force induced Gα12/13-Rho activation and increased F-actin to stimulate PLD activity. The activation can be partially prevented by the C3 exoenzyme. Pretreatment of the cells with chemical inhibitors of several kinases showed that calmodulin-dependent kinase is also involved in stretch-induced PLD activation by a separate pathway. Taken together, our data demonstrate that in cultured podocytes and MDCK cells, mechanical force leads to actin cytoskeleton reorganization and PLD activation. The signaling pathways for PLD activation involve Gα12/13/Rho/F-actin and calmodulin-dependent kinase.

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