Delineation of the functional properties exhibited by the Zinc-Activated Channel (ZAC) and its high-frequency Thr¹²⁸Ala variant (rs2257020) in Xenopus oocytes

The signalling characteristics of the Zinc-Activated Channel (ZAC), a member of the Cys-loop receptor (CLR) superfamily, are presently poorly elucidated. The ZACN polymorphism c.454G>A encoding for the Thr¹²⁸Ala variation in ZAC is found in extremely high allele frequencies across ethnicities. In this, the first study of ZAC in Xenopus oocytes by TEVC electrophysiology, ZAC^Thr128 and ZAC^Ala128 exhibited largely comparable pharmacological and signalling characteristics, but interestingly the Zn²⁺- and H⁺-evoked current amplitudes in ZAC^Ala128-oocytes were dramatically smaller than those in ZAC^Thr128-oocytes. While the variation thus appeared to impact cell surface expression and/or channel properties of ZAC, the similar expression properties exhibited by ZAC^Thr128 and ZAC^Ala128 in transfected mammalian cells indicated that their distinct functionalities could arise from the latter. In co-expression experiments, wild-type and variant ZAC subunits assembled efficiently into "heteromeric" complexes in HEK293 cells, while the concomitant presence of ZAC^Ala128 in ZAC^Thr128:ZAC^Ala128-oocytes did not exert a dominant negative effect on agonist-evoked current amplitudes compared to those in ZAC^Thr128-oocytes. Finally, the structural determinants of the functional importance of the 1-hydroxyethyl side-chain of Thr¹²⁸ appeared to be subtle, as agonist-evoked current amplitudes in ZAC^Ser128-, ZAC^Val128- and ZAC^Ile128-oocytes also were substantially lower than those in ZAC^Thr128-oocytes. In conclusion, the functional properties exhibited by ZAC in this work substantiate the notion of it being an atypical CLR. While the impact of the Thr¹²⁸Ala variation on ZAC functionality in oocytes is striking, it remains to be investigated whether and to which extent this translates into an in vivo setting and thus could constitute a source of inter-individual variation in ZAC physiology.