Molecular Basis for Zinc Transporter 1 Action as an Endogenous Inhibitor of L-type Calcium Channels

• Published in: The Journal of biological chemistry 2009-11, Vol.284 (47), p.32434-32443
• Main Authors: Levy, Shiri, Beharier, Ofer, Etzion, Yoram, Mor, Merav, Buzaglo, Liat, Shaltiel, Lior, Gheber, Levi A., Kahn, Joy, Muslin, Anthony J., Katz, Amos, Gitler, Daniel, Moran, Arie
• Format: Article
• Language: English
• Subjects: Animals; Calcium Channels, L-Type - chemistry; Carrier Proteins - metabolism; Carrier Proteins - physiology; CHO Cells; Cricetinae; Cricetulus; Female; Fluorescence Resonance Energy Transfer; Humans; Membrane Transport, Structure, Function, and Biogenesis; Models, Biological; Myocytes, Cardiac - cytology; Oocytes - metabolism; Patch-Clamp Techniques; Rats; Rats, Sprague-Dawley; Xenopus
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M109.058842

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 β-subunit. Moreover, the ZnT-1-induced inhibition of the LTCC current is also abolished by excess levels of the β-subunit. An interaction between ZnT-1 and the β-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 α1-subunit of the LTCC. Similarly, a decrease in the surface expression of the α1-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 β-subunit and that it involves a decrease in the trafficking of the LTCC α1-subunit to the surface membrane.