title: Modulation of voltage- and Ca2+-dependent gating of CaV1.3 L-type calcium channels by alternative splicing of a C-terminal regulatory domain.
abstract: Low voltage activation of Ca(V)1.3 L-type Ca(2+) channels controls excitability in sensory cells and central neurons as well as sinoatrial node pacemaking. Ca(V)1.3-mediated pacemaking determines neuronal vulnerability of dopaminergic striatal neurons affected in Parkinson disease. We have previously found that in Ca(V)1.4 L-type Ca(2+) channels, activation, voltage, and calcium-dependent inactivation are controlled by an intrinsic distal C-terminal modulator. Because alternative splicing in the Ca(V)1.3 alpha1 subunit C terminus gives rise to a long (Ca(V)1.3(42)) and a short form (Ca(V)1.3(42A)), we investigated if a C-terminal modulatory mechanism also controls Ca(V)1.3 gating. The biophysical properties of both splice variants were compared after heterologous expression together with beta3 and alpha2delta1 subunits in HEK-293 cells. Activation of calcium current through Ca(V)1.3(42A) channels was more pronounced at negative voltages, and inactivation was faster because of enhanced calcium-dependent inactivation. By investigating several Ca(V)1.3 channel truncations, we restricted the modulator activity to the last 116 amino acids of the C terminus. The resulting Ca(V)1.3(DeltaC116) channels showed gating properties similar to Ca(V)1.3(42A) that were reverted by co-expression of the corresponding C-terminal peptide C(116). Fluorescence resonance energy transfer experiments confirmed an intramolecular protein interaction in the C terminus of Ca(V)1.3 channels that also modulates calmodulin binding. These experiments revealed a novel mechanism of channel modulation enabling cells to tightly control Ca(V)1.3 channel activity by alternative splicing. The absence of the C-terminal modulator in short splice forms facilitates Ca(V)1.3 channel activation at lower voltages expected to favor Ca(V)1.3 activity at threshold voltages as required for modulation of neuronal firing behavior and sinoatrial node pacemaking.
authors.affiliation: Institute of Pharmacy, Pharmacology, and Toxicology, and Center for Molecular Biosciences, University of Innsbruck, Peter-Mayr-Strasse 1/I, Innsbruck, Austria.
authors.forename: Anamika
authors.initials: A
authors.lastname: Singh
authors.forename: Mathias
authors.initials: M
authors.lastname: Gebhart
authors.forename: Reinhard
authors.initials: R
authors.lastname: Fritsch
authors.forename: Martina J
authors.initials: MJ
authors.lastname: Sinnegger-Brauns
authors.forename: Chiara
authors.initials: C
authors.lastname: Poggiani
authors.forename: Jean-Charles
authors.initials: JC
authors.lastname: Hoda
authors.forename: Jutta
authors.initials: J
authors.lastname: Engel
authors.forename: Christoph
authors.initials: C
authors.lastname: Romanin
authors.forename: Jörg
authors.initials: J
authors.lastname: Striessnig
authors.forename: Alexandra
authors.initials: A
authors.lastname: Koschak
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000737:chemistry / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008297:Male; D051379:Mice; D008954:Models, Biological; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary
chemical_list: C472073:CACNA1D protein, human; C485995:Cacna1d protein, mouse; D020746:Calcium Channels, L-Type; D002118:Calcium