title: Function and regulation of the Na+-Ca2+ exchanger NCX3 splice variants in brain and skeletal muscle.
abstract: Isoform 3 of the Na(+)-Ca(2+) exchanger (NCX3) is crucial for maintaining intracellular calcium ([Ca(2+)]i) homeostasis in excitable tissues. In this sense NCX3 plays a key role in neuronal excitotoxicity and Ca(2+) extrusion during skeletal muscle relaxation. Alternative splicing generates two variants (NCX3-AC and NCX3-B). Here, we demonstrated that NCX3 variants display a tissue-specific distribution in mice, with NCX3-B as mostly expressed in brain and NCX-AC as predominant in skeletal muscle. Using Fura-2-based Ca(2+) imaging, we measured the capacity and regulation of the two variants during Ca(2+) extrusion and uptake in different conditions. Functional studies revealed that, although both variants are activated by intracellular sodium ([Na(+)]i), NCX3-AC has a higher [Na(+)]i sensitivity, as Ca(2+) influx is observed in the presence of extracellular Na(+). This effect could be partially mimicked for NCX3-B by mutating several glutamate residues in its cytoplasmic loop. In addition, NCX3-AC displayed a higher capacity of both Ca(2+) extrusion and uptake compared with NCX3-B, together with an increased sensitivity to intracellular Ca(2+). Strikingly, substitution of Glu(580) in NCX3-B with its NCX3-AC equivalent Lys(580) recapitulated the functional properties of NCX3-AC regarding Ca(2+) sensitivity, Lys(580) presumably acting through a structure stabilization of the Ca(2+) binding site. The higher Ca(2+) uptake capacity of NCX3-AC compared with NCX3-B is in line with the necessity to restore Ca(2+) levels in the sarcoplasmic reticulum during prolonged exercise. The latter result, consistent with the high expression in the slow-twitch muscle, suggests that this variant may contribute to the Ca(2+) handling beyond that of extruding Ca(2+).
authors.affiliation: From the Departments of Physiology and Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands; Centre for System Biology and Bioenergetics, Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands.
authors.forename: Lauriane Y M
authors.initials: LYM
authors.lastname: Michel
authors.affiliation: From the Departments of Physiology and Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands.
authors.forename: Sjoerd
authors.initials: S
authors.lastname: Verkaart
authors.affiliation: Centre for System Biology and Bioenergetics, Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands; Departments of Biochemistry, Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands.
authors.forename: Werner J H
authors.initials: WJH
authors.lastname: Koopman
authors.affiliation: Centre for System Biology and Bioenergetics, Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands; Departments of Biochemistry, Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands.
authors.forename: Peter H G M
authors.initials: PHGM
authors.lastname: Willems
authors.affiliation: From the Departments of Physiology and Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands.
authors.forename: Joost G J
authors.initials: JGJ
authors.lastname: Hoenderop
authors.affiliation: From the Departments of Physiology and Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands; Centre for System Biology and Bioenergetics, Radboud University Medical Centre, 6500HB Nijmegen, The Netherlands. Electronic address: Rene.Bindels@radboudumc.nl.
authors.forename: René J M
authors.initials: RJM
authors.lastname: Bindels
mesh_terms: D017398:Alternative Splicing / Q000502:physiology; D019943:Amino Acid Substitution; D000818:Animals; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D018657:Muscle Fibers, Slow-Twitch / Q000166:cytology / Q000378:metabolism*; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D009928:Organ Specificity; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D055550:Protein Stability; D012519:Sarcoplasmic Reticulum / Q000235:genetics / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000235:genetics / Q000378:metabolism*
chemical_list: D009124:Muscle Proteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; C491101:Slc8a3 protein, mouse; D019831:Sodium-Calcium Exchanger; D002118:Calcium
keywords: Alternative Splicing; Calcium Transport; Endoplasmic Reticulum (ER); Excitotoxicity; Forward Mode; Muscle Fatigue; NCX; Plasma Membrane; Reverse Mode; Sodium Calcium Exchange