Identification of a new C-terminal splice variant of CaV1.3 L-type calcium channels with unique functional properties
© Juhasz-Vedres et al; licensee BioMed Central Ltd. 2011
Published: 5 September 2011
In L-type voltage-gated calcium channels (VGCCs) the long C-terminal tail contains several sites for modulation by protein-protein interaction. CaV1.3 VGCCs (CaV1.3L) activate at negative voltages and support sinoatrial node pacemaking and hearing, and shape neuronal excitability. In CaV1.3L an intermolecular automodulatory C-terminal interaction (CTM) has been described which affects channel gating. CTM is characterized by interaction of a distal C-terminal regulatory domain (DCRD) with a more proximal regulatory domain (PCRD). If this CTM is absent as in previously described short CaV1.342A, calcium-dependent inactivation (CDI) increases and the channel activation range shifts to more negative voltages (i.e. “short” gating properties). Here we show that alternative splicing in exon 43 creates a new short splice variant CaV1.343S found in human and mouse brain. It lacks CTM, but still contains the PCRD motif, in contrast to previously described CaV1.342A. Semiquantitative PCR experiments showed that in mouse brain 39% of CaV1.3 channels contain exon 43S contrary to heart (6% 43S).
Methods and results
Biophysical analysis showed “short” gating properties for CaV1.343S in both 15 mM and 2 mM external Ca2+ when co-expressed with β3 and α2δ-1 subunits in tsA-201 cells. In 2 mM Ca2+ the inactivation rate of CaV1.343S was faster for CaV1.3L, but slower for CaV1.342A (% inactivation after 100 ms at Vmax: CaV1.342A: 64.5 ± 3.5%; CaV1.343S: 52 ± 4.5%, CaV1.3L: 37.4 ± 3%, p < 0.001) by affecting the extent of CDI. Due to a presence of PCRD, DCRD-containing C-terminal fragments from CaV1.3 or CaV1.2 channels could restore CaV1.3L gating behaviour. Indeed, co-expression of GFP-CaV1.2C349 fully restored long channel gating properties in CaV1.343S (V0.5: CaV1.343S+GFP-CaV1.2C349: 1.37 ± 1.3 mV; CaV1.3L: −2.4 ± 0.6 mV). C-terminal splicing also changed ICa kinetics during stimuli mimicking trains of action potential waveforms, revealing a lower total ICa during AP bursts in short splice variants.
Taken together, our data indicate that CaV1.3 C-terminal splicing can serve as an important mechanism to fine-tune the dynamics of calcium entry in neurons in an activity dependent manner.
Supported by FWF P-20670 JS, P-22528 AK, and SFB F44.
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