Molecular basis for zinc transporter 1 action as an endogenous inhibitor of L-type calcium channels.
The L-type calcium channel (LTCC) has a variety of physiological roles that are critical for the proper function of many cell types and organs. Recently, a member of the zinc-regulating family of proteins, ZnT-1, was recognized as an endogenous inhibitor of the LTCC, but its mechanism of action has not been elucidated. In the present study, using two-electrode voltage clamp recordings in Xenopus oocytes, we demonstrate that ZnT-1-mediated inhibition of the LTCC critically depends on the presence of the LTCC regulatory beta-subunit. Moreover, the ZnT-1-induced inhibition of the LTCC current is also abolished by excess levels of the beta-subunit. An interaction between ZnT-1 and the beta-subunit, as demonstrated by co-immunoprecipitation and by fluorescence resonance energy transfer, is consistent with this result. Using surface biotinylation and total internal reflection fluorescence microscopy in HEK293 cells, we show a ZnT-1-dependent decrease in the surface expression of the pore-forming alpha(1)-subunit of the LTCC. Similarly, a decrease in the surface expression of the alpha(1)-subunit is observed following up-regulation of the expression of endogenous ZnT-1 in rapidly paced cultured cardiomyocytes. We conclude that ZnT-1-mediated inhibition of the LTCC is mediated through a functional interaction of ZnT-1 with the LTCC beta-subunit and that it involves a decrease in the trafficking of the LTCC alpha(1)-subunit to the surface membrane.
Levy
Shiri
S
Department of Physiology, Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva 84101, Israel.
Beharier
Ofer
O
Etzion
Yoram
Y
Mor
Merav
M
Buzaglo
Liat
L
Shaltiel
Lior
L
Gheber
Levi A
LA
Kahn
Joy
J
Muslin
Anthony J
AJ
Katz
Amos
A
Gitler
Daniel
D
Moran
Arie
A
D000818:Animals; D016466:CHO Cells; D020746:Calcium Channels, L-Type / Q000737:chemistry*; D002352:Carrier Proteins / Q000378:metabolism / Q000502:physiology*; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008954:Models, Biological; D032383:Myocytes, Cardiac / Q000166:cytology; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D051381:Rats; D017207:Rats, Sprague-Dawley; D014981:Xenopus
D020746:Calcium Channels, L-Type; D002352:Carrier Proteins; C017800:zinc-binding protein