Activity of the yeast vacuolar TRP channel TRPY1 is inhibited by Ca2+-calmodulin binding.

Abstract Transient receptor (TRP) cation channels, which are conserved across mammals, flies, fish, sea squirts, worms, and fungi essentially contribute to cellular Ca2+ signalling. The activity of the unique TRP channel in yeast, TRPY1, relies on the vacuolar and cytoplasmic Ca2+ concentration. However, the mechanism(s) of Ca2+-dependent regulation of TRPY1 and possible contribution(s) of Ca2+-binding proteins are yet not well understood. Our results demonstrate a Ca2+-dependent binding of yeast calmodulin to TRPY1. TRPY1 activity was increased in the cmd1-6 yeast strain, carrying a non-Ca2+-binding calmodulin mutant, compared to the parent strain expressing wild-type calmodulin (Cmd1). Expression of Cmd1 in cmd1-6 yeast rescued the wild-type phenotype. In addition, in HEK-293 cells hypertonic shock-induced TRPY1-dependent Ca2+ influx and Ca2+ release were increased by the calmodulin antagonist ophiobolin A. We found that co-expression of mammalian calmodulin impeded TRPY1´s activity by reinforcing effects of endogenous calmodulin. Finally, inhibition of TRPY1 by Ca2+-calmodulin required the cytoplasmic amino acid stretch E33-Y92. In summary, our results show that TRPY1 is under inhibitory control of Ca2+-calmodulin and that mammalian calmodulin can replace yeast calmodulin for this inhibition. These findings add TRPY1 to the innumerable cellular proteins, which include a variety of ion channels, that use calmodulin (CaM) as a constitutive or dissociable Ca2+-sensing subunit, and contribute to a better understanding of the modulatory mechanisms of Ca2+-calmodulin.