Modulation of Nav1.5 channel function by an alternatively spliced sequence in the DII/DIII linker region.
In the present study, we identified a novel splice variant of the human cardiac Na(+) channel Na(v)1.5 (Na(v)1.5d), in which a 40-amino acid sequence of the DII/DIII intracellular linker is missing due to a partial deletion of exon 17. Expression of Na(v)1.5d occurred in embryonic and adult hearts of either sex, indicating that the respective alternative splicing is neither age-dependent nor gender-specific. In contrast, Na(v)1.5d was not detected in the mouse heart, indicating that alternative splicing of Na(v)1.5 is species-dependent. In HEK293 cells, splice variant Na(v)1.5d generated voltage-dependent Na(+) currents that were markedly reduced compared with wild-type Na(v)1.5. Experiments with mexiletine and 8-bromo-cyclic AMP suggested that the trafficking of Na(v)1.5d channels was not impaired. However, single-channel recordings showed that the whole-cell current reduction was largely due to a significantly reduced open probability. Additionally, steady-state activation and inactivation were shifted to depolarized potentials by 15.9 and 5.1 mV, respectively. Systematic mutagenesis analysis of the spliced region provided evidence that a short amphiphilic region in the DII/DIII linker resembling an S4 voltage sensor of voltage-gated ion channels is an important determinant of Na(v)1.5 channel gating. Moreover, the present study identified novel short sequence motifs within this amphiphilic region that specifically affect the voltage dependence of steady-state activation and inactivation and current amplitude of human Na(v)1.5.
Institute of Physiology II, Friedrich Schiller University, 07740 Jena, Germany.
Juan A
JA
Camacho
Sabine
S
Hensellek
Jean-Sébastien
JS
Rougier
Steve
S
Blechschmidt
Hugues
H
Abriel
Klaus
K
Benndorf
Thomas
T
Zimmer
D000328:Adult; D000367:Age Factors; D000368:Aged; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D004594:Electrophysiology; D004622:Embryo, Mammalian; D005091:Exons; D005260:Female; D016368:Frameshift Mutation; D020869:Gene Expression Profiling; D006321:Heart / Q000502:physiology*; D006801:Humans; D015640:Ion Channel Gating; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000502:physiology*; D016297:Mutagenesis, Site-Directed; D062554:NAV1.5 Voltage-Gated Sodium Channel; D012313:RNA; D012737:Sex Factors; D015222:Sodium Channels / Q000502:physiology*
D009124:Muscle Proteins; D062554:NAV1.5 Voltage-Gated Sodium Channel; C568320:SCN5A protein, human; C568321:Scn5a protein, mouse; D015222:Sodium Channels; D012313:RNA