Functional interaction of auxiliary subunits and synaptic proteins with Ca(v)1.3 may impart hair cell Ca2+ current properties.
We assessed the functional determinants of the properties of L-type Ca(2+) currents in hair cells by co-expressing the pore-forming Ca(V)1.3alpha(1) subunit with the auxiliary subunits beta(1A) and/or alpha(2delta). Because Ca(2+) channels in hair cells are poised to interact with synaptic proteins, we also co-expressed the Ca(V)1.3alpha(1) subunit with syntaxin, vesicle-associated membrane protein (VAMP), and synaptosome associated protein of 25 kDa (SNAP25). Expression of the Ca(V)1.3alpha(1) subunit in human embryonic kidney cells (HEK 293) produced a dihydropyridine (DHP)-sensitive Ca(2+) current (peak current density -2.0 +/- 0.2 pA/pF; n = 11). Co-expression with beta(1A) and alpha(2delta) subunits enhanced the magnitude of the current (peak current density: Ca(V)1.3alpha(1) + beta(1A) = -4.3 +/- 0.8 pA/pF, n = 10; Ca(V)1.3alpha(1) + beta(1A) + alpha(2delta) = -4.1 +/- 0.6 pA/pF, n = 9) and produced a leftward shift of approximately 9 mV in the voltage-dependent activation of the currents. Furthermore, co-expression of Ca(V)1.3alpha(1) with syntaxin/VAMP/SNAP resulted in at least a twofold increase in the peak current density (-4.7 +/- 0.2 pA/pF; n = 11) and reduced the extent of inactivation of the Ca(2+) currents. Botulinum toxin, an inhibitor of syntaxin, accelerated the inactivation profile of Ca(2+) currents in hair cells. Immunocytochemical data also indicated that the Ca(2+) channels and syntaxin are co-localized in hair cells, suggesting there is functional interaction of the Ca(V)1.3alpha(1) with auxiliary subunits and synaptic proteins, that may contribute to the distinct properties of the DHP-sensitive channels in hair cells.
Song
Haitao
H
Center for Neuroscience, Department of Otolaryngology, University of California, Davis, California 95616, USA.
Nie
Liping
L
Rodriguez-Contreras
Adrian
A
Sheng
Zu-Hang
ZH
Yamoah
Ebenezer N
EN
D000818:Animals; D001905:Botulinum Toxins / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels; D020746:Calcium Channels, L-Type / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004095:Dihydropyridines / Q000494:pharmacology; D006198:Hair Cells, Auditory / Q000502:physiology*; D006801:Humans; D015640:Ion Channel Gating / Q000502:physiology; D007668:Kidney / Q000166:cytology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D011892:Rana catesbeiana; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013569:Synapses / Q000502:physiology*; D050825:Synaptosomal-Associated Protein 25; D014162:Transfection
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D004095:Dihydropyridines; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D011994:Recombinant Proteins; C494208:SNAP25 protein, human; D050825:Synaptosomal-Associated Protein 25; C093712:Cacna1d protein, rat; C038806:1,4-dihydropyridine; D001905:Botulinum Toxins; C096324:botulinum toxin type C; D002118:Calcium