期刊
BIOCHEMICAL PHARMACOLOGY
卷 193, 期 -, 页码 -出版社
PERGAMON-ELSEVIER SCIENCE LTD
DOI: 10.1016/j.bcp.2021.114781
关键词
Cys-loop receptor (CLR); Pentameric ligand-gated ion channel (pLGIC); Zinc-Activated Channel (ZAC); Agonist binding; Chimeric subunits; Leu9' residue
资金
- Danish Council of Independent Research for Medical Sciences
- Carlsberg Foundation
- NIH-National Institute of Mental Health [MH097446]
- National Institute of Neurological Disorders and Stroke [NS108378, NS111064, NS111338]
The molecular mechanisms underlying ZAC signaling were explored using Xenopus oocytes, revealing that Zn2+ and protons predominantly activate ZAC through its extracellular domain. Despite differences in identified agonists and channel characteristics, the signal transduction of ZAC appears to undergo similar conformational transitions as classical CLRs.
The molecular basis for the signal transduction through the classical Cys-loop receptors (CLRs) has been delineated in great detail. The Zinc-Activated Channel (ZAC) constitutes a so far poorly elucidated fifth branch of the CLR superfamily, and in this study we explore the molecular mechanisms underlying ZAC signaling in Xenopus oocytes by two-electrode voltage clamp electrophysiology. In studies of chimeric receptors fusing either the extracellular domain (ECD) or the transmembrane/intracellular domain (TMD-ICD) of ZAC with the complementary domains of 5-HT(3)A serotonin or alpha(1) glycine receptors, serotonin and Zn2+/H+ evoked robust concentration-dependent currents in 5-HT(3)A/ZAC- and ZAC/alpha(1)-Gly-expressing oocytes, respectively, suggesting that Zn2+ and protons activate ZAC predominantly through its ECD. The molecular basis for Zn2+-mediated ZAC signaling was probed further by introduction of mutations of His, Cys, Glu and Asp residues in this domain, but as none of the mutants tested displayed substantially impaired Zn2+ functionality compared to wild-type ZAC, the location of the putative Zn2+ binding site(s) in the ECD was not identified. Finally, the functional importance of Leu(246) (Leu9') in the transmembrane M2 alpha-helix of ZAC was investigated by Ala, Val, Ile and Thr substitutions. In concordance with findings for this highly conserved residue in classical CLRs, the ZAC(L9'X) mutants exhibited left-shifted agonist concentration-response relationships, markedly higher degrees of spontaneous activity and slower desensitization kinetics compared to wild-type ZAC. In conclusion, while ZAC is an atypical CLR in terms of its (identified) agonists and channel characteristics, its signal transduction seems to undergo similar conformational transitions as those in the classical CLR.
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