Linoleic acid induces opening of connexin26 hemichannels through a PI3K/Akt/Ca2+-dependent pathway

Figueroa, Vania; Saéz, Pablo; Salas, José D.; Salas, Daniela; Jara, Oscar; Martı́nez, Agustı́n D.; Sáez, Juan C.; Retamal, Mauricio A.

doi:10.1016/j.bbamem.2012.12.006

Cited by 28 publications

(40 citation statements)

References 47 publications

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“…Using a different experimental approach to deplete intracellular ATP consisting in hypoxia, nutrient deprivation, high K + and acidic pH to mimic the extracellular milieu of ischemic cells, our results showed that Akt-mediated regulation of Cx43 HCs was also present in this experimental model. Considering that linoleic acid, a pro-inflammatory unsaturated fatty acid, also increase Cx43 HC activity by a Ca 2+/ Akt dependent pathway [29], we propose that this mechanism is activated in general inflammatory states regardless the stimuli. Considering our results and those reported previously, Akt-dependent regulation of Cx43 HCs and gap junctions seems to be a general mechanism operating in physiological (mechanic stress) [26] as well as pathologic conditions (hypoxia/reoxygenation, MI) [25,45], but further experiments should be done to test this hypothesis.…”

Section: Discussionmentioning

confidence: 99%

“…Images were acquired by using an Olympus BX 51W1I upright microscope and the imaging system described above. Data were recorded every 1 min and analyzed with METAFLUOR software (Universal Imaging, Downingtown, PA) as described previously [29]. For experiments with Akti, cells were first incubated for 30 min with 10 μM Akti and then loaded with Fura2 by incubating them for additional 30 min in saline solution containing 5 μM Fura2-AM.…”

Section: Methodsmentioning

confidence: 99%

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Role of Akt and Ca2+ on cell permeabilization via connexin43 hemichannels induced by metabolic inhibition

Salas

Puebla

Lampe

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Basis of Diseas

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Connexin hemichannels are regulated under physiological and pathological conditions. Metabolic inhibition, a model of ischemia, promotes surface hemichannel activation associated, in part, with increased surface hemichannel levels, but little is known about its underlying mechanism. Here, we investigated the role of Akt on the connexin43 hemichannel's response induced by metabolic inhibition. In HeLa cells stably transfected with rat connexin43 fused to EGFP (HeLa43 cells), metabolic inhibition induced a transient Akt activation necessary to increase the amount of surface connexin43. The increase in levels of surface connexin43 was also found to depend on an intracellular Ca2+ signal increase that was partially mediated by Akt activation. However, the metabolic inhibition-induced Akt activation was not significantly affected by intracellular Ca2+ chelation. The Akt-dependent increase in connexin43 hemichannel activity in HeLa43 cells also occurred after oxygen-glucose deprivation, another ischemia-like condition, and in cultured cortical astrocytes (endogenous connexin43 expression system) under metabolic inhibition. Since opening of hemichannels has been shown to accelerate cell death, inhibition of Akt-dependent phosphorylation of connexin43 hemichannels could reduce cell death induced by ischemia/reperfusion.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Role of Akt and Ca2+ on cell permeabilization via connexin43 hemichannels induced by metabolic inhibition

Salas

Puebla

Lampe

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Basis of Diseas

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“…CaM was recognized to play a critical role in regulating gap junction channels by uncoupling gap junction channels and inhibiting gap junction permeability (19). Although extracellular Ca 2+ inhibits hemichannel opening, intracellular Ca 2+ is reported to positively regulate Cx43 and Cx32 hemichannel activity (20)(21)(22). Additionally, increased intracellular Ca 2+ enhances the electrical conductance of Cx46 hemichannels, contrary to the reduced gap junction channels (20).…”

Section: Discussionmentioning

confidence: 99%

“…Although extracellular Ca 2+ inhibits hemichannel opening, intracellular Ca 2+ is reported to positively regulate Cx43 and Cx32 hemichannel activity (20)(21)(22). Additionally, increased intracellular Ca 2+ enhances the electrical conductance of Cx46 hemichannels, contrary to the reduced gap junction channels (20). Concurring with our finding, another paper reports that G45 residue in Cx26 is a Ca 2+ sensor and mutation of this residue to E in the Keratitisichthyosis-deafness (KID) syndrome leads to "leaky" hemichannels and cell death (23).…”

Section: Discussionmentioning

confidence: 99%

Cataract-associated connexin 46 mutation alters its interaction with calmodulin and function of hemichannels

Riquelme²,

Wang

et al. 2018

Journal of Biological Chemistry

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Connexin channels help maintain eye lens homeostasis and transparency. The G143R missense substitution in connexin (Cx) 46 is associated with congenital Coppock cataracts; however, the underlying molecular mechanism is largely unknown. Here, we report that compared with wild type (WT) Cx46, the G143R substitution abolishes hemichannel conductance in Xenopus oocytes and in HeLa cells. Moreover, this substitution is dominant-negative and inhibits conductance of WT Cx46. CD analysis indicated that the substitution greatly reduces the α-helical structure of the intracellular Cx46 loop domain. Protein pulldown assays and isothermal titration calorimetry (ITC) revealed that this Cx46 domain directly interacts with calmodulin (CaM) in a Ca 2+ -dependent fashion, an observation confirmed by immunofluorescent co-localization of Cx46 with CaM. Interestingly, the G143R substitution enhanced the Cx46-CaM interaction and attenuated its abolishment by Ca 2+ depletion. Moreover, Cx46 increased dye influx, and the G143R substitution augmented this effect. Inhibition of Ca 2+ -mediated CaM activation blocked hemichannel permeability. The membrane potential plays a crucial role in Cx46 membrane permeability. We found that the activity of hemichannels is detectable under rest and hyperpolarization conditions, but is eliminated with depolarization. These results suggested that the G143R substitution impairs voltage-dependent electrical conductance and alters membrane permeability mediated by Cx46 hemichannels. The latter likely is caused by the substitution-induced structural changes of the intracellular loop domain associated with the increased interaction with CaM and reduced Ca 2+ sensitivity. The data suggest that the G143R-induced enhancement of the CaMCx46 interaction results in altered hemichannel activities and might be related to cataract formation.The lens is a clear part of the anterior eye that focuses light and images onto the retina. Cataracts, referring to the clouding of the lens, are responsible for 51% of world blindness according to WHO data in 2010 (1). The lens has three main parts; the capsule, the epithelial layer in the anterior portion, and fiber cells which comprise the bulk of lens (2). Lens fiber cells communicate to each other and to the surface of the lens by gap junction channels and hemichannels. These allow small molecules (< 1.2 kDa) like ions, second messenger and metabolites to pass through adjacent cells (3-5). Lens is an avascular organ and thus relies in part upon gap junction channels and hemichannels to maintain its metabolic homeostasis and transparency. Hemichannels, also called connexins, are undocked http://www.jbc.org/cgi

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“…However, knowledge about the regulation of HCs (or GJCs) by FFAs in the epithelium of the gastrointestinal tract is very limited and is incompletely understood in other cell types [21-24]. …”

Section: Introductionmentioning

confidence: 99%

Linoleic acid permeabilizes gastric epithelial cells by increasing connexin 43 levels in the cell membrane via a GPR40- and Akt-dependent mechanism

Puebla

Cisterna

Salas

et al. 2016

Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biolog

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Linoleic acid (LA) is known to activate G-protein coupled receptors and connexin hemichannels (Cx HCs) but possible interlinks between these two responses remain unexplored. Here, we evaluated the mechanism of action of LA on the membrane permeability mediated by Cx HCs in MKN28 cells. These cells were found to express connexins, GPR40, GPR120, and CD36 receptors. The Cx HC activity of these cells increased after 5 min of treatment with LA or GW9508, an agonist of GPR40/GPR120; or exposure to extracellular divalent cation-free solution (DCFS), known to increase the open probability of Cx HCs, yields an immediate increase in Cx HC of similar intensity and additive with LA-induced change. Treatment with a CD36 blocker or transfection with siRNA-GPR120 maintain the LA-induced Cx HC activity. However, cells transfected with siRNA-GPR40 did not show LA-induced Cx HC activity but activity was increased upon exposure to DCFS, confirming the presence of activatable Cx HCs in the cell membrane. Treatment with AKTi (Akt inhibitor) abrogated the LA-induced Cx HC activity. In HeLa cells transfected with Cx43 (HeLa-Cx43), LA induced phosphorylation of surface Cx43 at serine 373 (S373), site for Akt phosphorylation. HeLa-Cx43 but not HeLa-Cx43 cells with a S373A mutation showed a LA-induced Cx HC activity directly related to an increase in cell surface Cx43 levels. Thus, the increase in membrane permeability induced by LA is mediated by an intracellular signaling pathway activated by GPR40 that leads to an increase in membrane levels of Cx43 phosphorylated at serine 373 via Akt.

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Linoleic acid induces opening of connexin26 hemichannels through a PI3K/Akt/Ca2+-dependent pathway

Cited by 28 publications

References 47 publications

Role of Akt and Ca2+ on cell permeabilization via connexin43 hemichannels induced by metabolic inhibition

Role of Akt and Ca2+ on cell permeabilization via connexin43 hemichannels induced by metabolic inhibition

Cataract-associated connexin 46 mutation alters its interaction with calmodulin and function of hemichannels

Linoleic acid permeabilizes gastric epithelial cells by increasing connexin 43 levels in the cell membrane via a GPR40- and Akt-dependent mechanism

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