scholarly journals Cl−secretion in ATP-treated renal epithelial C7-MDCK cells is mediated by activation of P2Y1receptors, phospholipase A2and protein kinase A

2005 ◽  
Vol 568 (3) ◽  
pp. 789-801 ◽  
Author(s):  
A. Olga Akimova ◽  
Nathalie Bourcier ◽  
Sebastien Taurin ◽  
Richard A. Bundey ◽  
Konrad Grygorczyk ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Mdck Cells ◽  
Cl Secretion ◽  
Kinase A

Activators of protein kinase A and of protein kinase C inhibit MDCK cell myo-inositol and betaine uptake.

1995 ◽  
Vol 6 (6) ◽  
pp. 1559-1564
Author(s):  
A S Preston ◽  
A Yamauchi ◽  
H M Kwon ◽  
J S Handler
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Translational Regulation ◽  
Mdck Cells ◽  
Relative Effect ◽  
Amino Acid Sequences ◽  
Prolonged Incubation ◽  
Kinase C ◽  
Kinase A

Amino acid sequences of the myo-inositol and betaine cotransporters that are induced in MDCK cells by hypertonicity include consensus sequences for phosphorylation by protein kinase A and by protein kinase C. To test for the effect of activation of protein kinases A and C on the activity of those cotransporters, MDCK cells were exposed to activators of each kinase and the activity of both cotransporters was assayed. Incubation with 8-bromoadenosine 3':5'-cyclic monophosphate (8Br-cAMP) or 3-isobutyl-1-methylxanthine (IBMX), activators of protein kinase A, and incubation with an active phorbol ester or with an active diacylglycerol, activators of protein kinase C, inhibited the activity of both cotransporters by about 30%. The relative effect of the activation of protein kinase A and of protein kinase C was similar in hypertonic and isotonic cells. The effects of activators of protein kinase A and of protein kinase C were not additive. The two cotransporters behaved differently when protein kinase C activity was down-regulated by prolonged incubation with a higher concentration of phorbol 12-myristate 13-acetate. There was a doubling of activity of the myo-inositol cotransporter and no change in the activity of the betaine cotransporter in hypertonic and isotonic cells. Although the mechanisms of the effects of activation of the two kinases remain to be established, it is clear that the kinases can mediate post-translational regulation of the uptake of compatible osmolytes.

scholarly journals Epac1 mediates protein kinase A–independent mechanism of forskolin-activated intestinal chloride secretion

2009 ◽  
Vol 135 (1) ◽  
pp. 43-58 ◽  
Author(s):  
Kazi Mirajul Hoque ◽  
Owen M. Woodward ◽  
Damian B. van Rossum ◽  
Nicholas C. Zachos ◽  
Linxi Chen ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Chinese Hamster Ovary Cells ◽  
Short Circuit ◽  
Basolateral Membrane ◽  
Chinese Hamster ◽  
T84 Cells ◽  
Cl Secretion ◽  
Intestinal Chloride Secretion ◽  
Kinase A

Intestinal Cl− secretion is stimulated by cyclic AMP (cAMP) and intracellular calcium ([Ca2+]i). Recent studies show that protein kinase A (PKA) and the exchange protein directly activated by cAMP (Epac) are downstream targets of cAMP. Therefore, we tested whether both PKA and Epac are involved in forskolin (FSK)/cAMP-stimulated Cl− secretion. Human intestinal T84 cells and mouse small intestine were used for short circuit current (Isc) measurement in response to agonist-stimulated Cl− secretion. FSK-stimulated Cl− secretion was completely inhibited by the additive effects of the PKA inhibitor, H89 (1 µM), and the [Ca2+]i chelator, 1,2-bis-(o-aminophenoxy)-ethane-N,N,N’,N’-tetraacetic acid, tetraacetoxymethyl ester (BAPTA-AM; 25 µM). Both FSK and the Epac activator 8-pCPT-2’-O-Me-cAMP (50 µM) elevated [Ca2+]i, activated Ras-related protein 2, and induced Cl− secretion in intact or basolateral membrane–permeabilized T84 cells and mouse ileal sheets. The effects of 8-pCPT-2’-O-Me-cAMP were completely abolished by BAPTA-AM, but not by H89. In contrast, T84 cells with silenced Epac1 had a reduced Isc response to FSK, and this response was completely inhibited by H89, but not by the phospholipase C inhibitor U73122 or BAPTA-AM. The stimulatory effect of 8-pCPT-2’-O-Me-cAMP on Cl− secretion was not abolished by cystic fibrosis transmembrane conductance (CFTR) inhibitor 172 or glibenclamide, suggesting that CFTR channels are not involved. This was confirmed by lack of effect of 8-pCPT-2’-O-Me-cAMP on whole cell patch clamp recordings of CFTR currents in Chinese hamster ovary cells transiently expressing the human CFTR channel. Furthermore, biophysical characterization of the Epac1-dependent Cl− conductance of T84 cells mounted in Ussing chambers suggested that this conductance was hyperpolarization activated, inwardly rectifying, and displayed a Cl−>Br−>I− permeability sequence. These results led us to conclude that the Epac-Rap-PLC-[Ca2+]i signaling pathway is involved in cAMP-stimulated Cl− secretion, which is carried by a novel, previously undescribed Cl− channel.

scholarly journals Protein kinase A-regulated membrane trafficking of a green fluorescent protein-aquaporin 5 chimera in MDCK cells

2006 ◽  
Vol 1763 (4) ◽  
pp. 337-344 ◽  
Author(s):  
Chisato Kosugi-Tanaka ◽  
Xuefei Li ◽  
Chenjuan Yao ◽  
Tetsuya Akamatsu ◽  
Norio Kanamori ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Membrane Trafficking ◽  
Fluorescent Protein ◽  
Mdck Cells ◽  
Aquaporin 5 ◽  
Green Fluorescent ◽  
Kinase A

scholarly journals Prostaglandin E 2 ‐Stimulated Cl − Secretion in Airway Epithelium Involves Protein Kinase A and Calcium Crosstalk

2019 ◽  
Vol 33 (S1) ◽  
Author(s):  
Shayda Abazari ◽  
Gopika Hari ◽  
Elizabeth Crowther ◽  
Franny Kiles ◽  
Annie Trumbull ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Airway Epithelium ◽  
Prostaglandin E ◽  
Cl Secretion ◽  
Kinase A

scholarly journals Gs alpha stimulates transcytosis and apical secretion in MDCK cells through cAMP and protein kinase A.

1994 ◽  
Vol 126 (3) ◽  
pp. 677-687 ◽  
Author(s):  
S H Hansen ◽  
J E Casanova
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Adenylyl Cyclase ◽  
Mdck Cells ◽  
Transport Processes ◽  
Apical Plasma Membrane ◽  
Wild Type ◽  
Early Endosomes ◽  
Gs Alpha ◽  
Kinase A

Recent evidence suggests a role for heterotrimeric G proteins in vesicular transport. Cholera toxin, which activates Gs alpha by ADP-ribosylation, has been reported to stimulate both apical secretion (Pimplikar, S.W., and K. Simons. 1993. Nature (Lond.). 352:456-458) and apically directed transcytosis (Bomsel, M., and K.E. Mostov. 1993. J. Biol. Chem. 268:25824-25835) in MDCK cells, via a cAMP-independent mechanism. Here, we demonstrate that apical secretion and apically directed transcytosis are significantly stimulated by agents that elevate cellular cAMP. Forskolin, which activates adenylyl cyclase directly, and 8BrcAMP augment both transport processes in MDCK cells. The increase is not limited to receptor-mediated transport (polymeric Ig receptor), since transcytosis of ricin, a galactose-binding lectin, is similarly stimulated. The effects of elevated cellular cAMP on apical secretion and transcytosis are apparently mediated via protein kinase A (PKA), as they are inhibited by H-89, a selective PKA inhibitor. Experiments employing a 17 degrees C temperature block indicate that cAMP/PKA acts at a late, possibly rate-limiting stage in the transcytotic pathway, after translocation of internalized markers into the apical cytoplasm. However, no significant stimulus of apical recycling was observed in the presence of FSK, suggesting that cAMP/PKA either affects transcytosis at a level proximal to apical early endosomes and/or specifically increases the efficiency by which transcytosing molecules are delivered to the apical plasma membrane. Finally, we overexpressed wild-type Gs alpha and a mutant, Q227L, which constitutively activates adenylyl cyclase, in MDCK cells. Although Q227L increased transcytosis more than wild-type Gs alpha, neither construct was as effective as FSK in stimulating transcytosis, arguing against a significant role of Gs alpha in transcytosis independent of cAMP and PKA.

scholarly journals Prostaglandin E2 Enhances Gap Junctional Intercellular Communication in Clonal Epithelial Cells

2021 ◽  
Vol 22 (11) ◽  
pp. 5813
Author(s):  
Alejandro Ogazon del Toro ◽  
Lidia Jimenez ◽  
Mauricio Serrano Rubi ◽  
Aida Castillo ◽  
Lorena Hinojosa ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Prostaglandin E2 ◽  
Protein Kinase A ◽  
Intercellular Communication ◽  
Mdck Cells ◽  
Lucifer Yellow ◽  
Gap Junctional Intercellular Communication ◽  
Gap Junctional ◽  
Kinase A

Prostaglandins are a group of lipids that produce diverse physiological and pathological effects. Among them, prostaglandin E2 (PGE2) stands out for the wide variety of functions in which it participates. To date, there is little information about the influence of PGE2 on gap junctional intercellular communication (GJIC) in any type of tissue, including epithelia. In this work, we set out to determine whether PGE2 influences GJIC in epithelial cells (MDCK cells). To this end, we performed dye (Lucifer yellow) transfer assays to compare GJIC of MDCK cells treated with PGE2 and untreated cells. Our results indicated that (1) PGE2 induces a statistically significant increase in GJIC from 100 nM and from 15 min after its addition to the medium, (2) such effect does not require the synthesis of new mRNA or proteins subunits but rather trafficking of subunits already synthesized, and (3) such effect is mediated by the E2 receptor, which, in turn, triggers a signaling pathway that includes activation of adenylyl cyclase and protein kinase A (PKA). These results widen the knowledge regarding modulation of gap junctional intercellular communication by prostaglandins.

scholarly journals Epac1 mediates Protein Kinase A (PKA) independent mechanism of forskolin (FSK) stimulated Cl secretion in T84 cells

2007 ◽  
Vol 21 (5) ◽  
Author(s):  
Mirajul Hoque Kazi ◽  
Damian B Rossum ◽  
Linxi Chen ◽  
Chung‐Ming Tse
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
T84 Cells ◽  
Cl Secretion ◽  
Independent Mechanism ◽  
Kinase A

Regulation of UT-A1-mediated transepithelial urea flux in MDCK cells

2006 ◽  
Vol 291 (4) ◽  
pp. C600-C606 ◽  
Author(s):  
Otto Fröhlich ◽  
Janet D. Klein ◽  
Pauline M. Smith ◽  
Jeff M. Sands ◽  
Robert B. Gunn
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Mdck Cells ◽  
Phosphodiesterase Inhibitor ◽  
Rapid Onset ◽  
Strong Increase ◽  
Urea Transport ◽  
Slow Increase ◽  
Basolateral Side ◽  
Kinase A

Transepithelial [14C]urea fluxes were measured across cultured Madin-Darby canine kidney (MDCK) cells permanently transfected to express the urea transport protein UT-A1. The urea fluxes were typically increased from a basal rate of 2 to 10 and 25 nmol·cm−2·min−1 in the presence of vasopressin and forskolin, respectively. Flux activation consisted of a rapid-onset component of small amplitude that leveled off within ∼10 min and at times even decreased again, followed by a delayed, strong increase over the next 30–40 min. Forskolin activated urea transport through activation of adenylyl cyclase; dideoxyforskolin was inactive. Vasopressin activated urea transport only from the basolateral side and was blocked by OPC-31260, indicating that its action was mediated by basolateral V2 receptors. In the presence of the phosphodiesterase inhibitor IBMX, vasopressin activated as strongly as forskolin. By itself, IBMX caused a slow increase over 50 min to ∼5 nmol·cm−2·min−1. 8-Bromoadenosine 3′,5′-cyclic monophosphate (8-BrcAMP; 300 μM) activated urea flux only when added basolaterally. IBMX augmented the activation by basolateral 8-BrcAMP. Urea flux activation by vasopressin and forskolin were only partially blocked by the protein kinase A inhibitor H-89. Even at concentrations >10 μM, urea flux after 60 min of stimulation was reduced by <50%. The rapid-onset component appeared unaffected by the presence of H-89. These data suggest that activation of transepithelial urea transport across MDCK-UT-A1 cells by forskolin and vasopressin involves cAMP as a second messenger and that it is mediated by one or more signaling pathways separate from and in addition to protein kinase A.

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