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Overexpression of Na+/Mg2+ exchanger SLC41A1 attenuates pro-survival signaling.

ABSTRACT: The Na+/Mg2+ exchanger SLC41A1 (A1), a key component of intracellular Mg homeostasis (IMH), is the major cellular Mg2+ efflux system, and its overexpression decreases [Mg2+]intracellular. IMH plays an important role in the regulation of many cellular processes, including cellular signaling. However, whether the overexpression of A1 and the consequent drop of [Mg2+]i impact on intracellular signaling is unknown. To examine the latter, we utilized dynamic mass redistribution (DMR) assay, PathScan® RTK signaling antibody (PRSA) array, confirmatory Western blot (WB) analyses of phosphorylation of kinases selected by PRSA, and mag-fura 2-assisted fast filter spectrometry (FFS). We demonstrate here that the overexpression of A1 quantitatively and qualitatively changes the DMR signal evoked by the application of PAR-1-selective activating peptide and/or by changing [Mg2+]extracellular in HEK293 cells. PRSA profiling of the phosphorylation of important signaling nodes followed by confirmatory WB has revealed that, in HEK293 cells, A1 overexpression significantly attenuates the phosphorylation of Akt/PKB on Thr308 and/or Ser473 and of Erk1/2 on Thr202/Tyr204 in the presence of 0 or 1 mM (physiological) Mg2+ in the bath solution. The latter is also true for SH-SY5Y and HeLa cells. Overexpression of A1 in HEK293 cells significantly lowers [Mg2+]i in the presence of [Mg2+]e = 0 or 1 mM. This correlates with the observed attenuation of prosurvival Akt/PKB - Erk1/2 signaling in these cells. Thus, A1 expression status and [Mg2+]e (and consequently also [Mg2+]i) modulate the complex physiological fingerprint of the cell and influence the activity of kinases involved in anti-apoptotic and, hence, pro-survival events in cells.

SUBMITTER: Sponder G

PROVIDER: S-EPMC5797035 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Overexpression of Na+/Mg2+ exchanger SLC41A1 attenuates pro-survival signaling.

Sponder Gerhard G Abdulhanan Nasrin N Fröhlich Nadine N Mastrototaro Lucia L Aschenbach Jörg R JR Röntgen Monika M Pilchova Ivana I Cibulka Michal M Racay Peter P Kolisek Martin M

Oncotarget 20171222 4

The Na+/Mg2+ exchanger SLC41A1 (A1), a key component of intracellular Mg homeostasis (IMH), is the major cellular Mg2+ efflux system, and its overexpression decreases [Mg2+]intracellular. IMH plays an important role in the regulation of many cellular processes, including cellular signaling. However, whether the overexpression of A1 and the consequent drop of [Mg2+]i impact on intracellular signaling is unknown. To exami ...[more]

PMID: 29435164

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Project description:Mice with smooth muscle (SM)-specific knockout of Na(+)/Ca(2+) exchanger type-1 (NCX1(SM-/-)) and the NCX inhibitor, SEA0400, were used to study the physiological role of NCX1 in mouse mesenteric arteries. NCX1 protein expression was greatly reduced in arteries from NCX1(SM-/-) mice generated with Cre recombinase. Mean blood pressure (BP) was 6-10 mmHg lower in NCX1(SM-/-) mice than in wild-type (WT) controls. Vasoconstriction was studied in isolated, pressurized mesenteric small arteries from WT and NCX1(SM-/-) mice and in heterozygotes with a global null mutation (NCX1(Fx/-)). Reduced NCX1 activity was manifested by a marked attenuation of responses to low extracellular Na(+) concentration, nanomolar ouabain, and SEA0400. Myogenic tone (MT, 70 mmHg) was reduced by approximately 15% in NCX1(SM-/-) arteries and, to a similar extent, by SEA0400 in WT arteries. MT was normal in arteries from NCX1(Fx/-) mice, which had normal BP. Vasoconstrictions to phenylephrine and elevated extracellular K(+) concentration were significantly reduced in NCX1(SM-/-) arteries. Because a high extracellular K(+) concentration-induced vasoconstriction involves the activation of L-type voltage-gated Ca(2+) channels (LVGCs), we measured LVGC-mediated currents and Ca(2+) sparklets in isolated mesenteric artery myocytes. Both the currents and the sparklets were significantly reduced in NCX1(SM-/-) (vs. WT or NCX1(Fx/-)) myocytes, but the voltage-dependent inactivation of LVGCs was not augmented. An acute application of SEA0400 in WT myocytes had no effect on LVGC current. The LVGC agonist, Bay K 8644, eliminated the differences in LVGC currents and Ca(2+) sparklets between NCX1(SM-/-) and control myocytes, suggesting that LVGC expression was normal in NCX1(SM-/-) myocytes. Bay K 8644 did not, however, eliminate the difference in myogenic constriction between WT and NCX1(SM-/-) arteries. We conclude that, under physiological conditions, NCX1-mediated Ca(2+) entry contributes significantly to the maintenance of MT. In NCX1(SM-/-) mouse artery myocytes, the reduced Ca(2+) entry via NCX1 may lower cytosolic Ca(2+) concentration and thereby reduce MT and BP. The reduced LVGC activity may be the consequence of a low cytosolic Ca(2+) concentration.

Dataset Information

Overexpression of Na+/Mg2+ exchanger SLC41A1 attenuates pro-survival signaling.

Publications

Overexpression of Na<sup>+</sup>/Mg<sup>2+</sup> exchanger SLC41A1 attenuates pro-survival signaling.

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