title: Differential regulation of the Na+-Ca2+exchanger 3 (NCX3) by protein kinase PKC and PKA.
abstract: Isoform 3 of the Na+-Ca2+exchanger (NCX3) participates in the Ca2+fluxes across the plasma membrane. Among the NCX family, NCX3 carries out a peculiar role due to its specific functions in skeletal muscle and the immune system and to its neuroprotective effect under stress exposure. In this context, proper understanding of the regulation of NCX3 is primordial to consider its potential use as a drug target. In this study, we demonstrated the regulation of NCX3 by protein kinase A (PKA) and C (PKC). Disparity in regulation has been previously reported among the splice variants of NCX3 therefore the activity of Ca2+uptake and extrusion of the two murine variants was measured using fura-2-based Ca2+imaging and revealed that both variants are similarly regulated. PKC stimulation diminished the Ca2+uptake performed by NCX3 in the reverse mode, triggered by a rise in [Ca2+]ior [Na+]i, whereas an opposite response was observed upon PKA stimulation, with a significant increase of the Ca2+uptake after a rise in [Ca2+]i. The latter stimulation affected similarly the efflux capacity of NCX3 whereas Ca2+extrusion capacity remained unaffected under activation of PKC. Next, using site-directed mutagenesis, the sensitivity of NCX3 to PKC was abolished by singly mutating its predicted phosphorylation sites T529 or S695. The sensitivity to PKC might be due to the influence of T529 phosphorylation on the Ca2+-binding domain 1. Additionally, we showed that stimulation of NCX3 by PKA occurred through residue S524. This effect may well participate in the fight-or-flight response in skeletal muscle and the long-term potentiation in hippocampus.
authors.affiliation: Department of Physiology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, The Netherlands; Centre for System Biology and Bioenergetics, Radboud University Medical Center, The Netherlands.
authors.forename: Lauriane Y M
authors.initials: LYM
authors.lastname: Michel
authors.affiliation: Department of Physiology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, The Netherlands.
authors.forename: Sjoerd
authors.initials: S
authors.lastname: Verkaart
authors.affiliation: Department of Physiology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, The Netherlands.
authors.forename: Femke
authors.initials: F
authors.lastname: Latta
authors.affiliation: Department of Physiology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, The Netherlands.
authors.forename: Joost G J
authors.initials: JGJ
authors.lastname: Hoenderop
authors.affiliation: Department of Physiology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, The Netherlands; Centre for System Biology and Bioenergetics, Radboud University Medical Center, The Netherlands. Electronic address: Rene.Bindels@radboudumc.nl.
authors.forename: René J M
authors.initials: RJM
authors.lastname: Bindels
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation / Q000235:genetics; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000235:genetics / Q000378:metabolism*
chemical_list: C101832:SLC38A3 protein, human; D019831:Sodium-Calcium Exchanger; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D002118:Calcium
keywords: Ca(2+) transport; Ca(2+)-binding domain; Long-term potentiation; Phosphorylation; Skeletal muscle; Sodium-calcium exchange