Physiological significance of functional coordination by Ca²⁺ channels in signalsome

Abstract

Mammalian homologues of Drosophira transient receptor potential (trp) proteins (TRP) form Ca²⁺ permeable cation channels, which are activated in response to stimulation of G-protein-coupled receptors. We have previously demonstrated that TRPC5 permeates Ca²⁺ in response to stimulation through direct cysteine oxidation by reactive oxygen species (ROS) and reactive nitrogen species (RNS). We have examined activity of TRPC5 channel recombinantly expressed in HEK293 cells using patch-clamp technique and the fluorescent Ca²⁺ indicator fura-2 for measurements of intracellular free calcium concentration ([Ca²⁺]_i). In bovine aortic endothelial cells (BAEC) treated with all-trans-retinoic acid (RA) or recombinantly expressing TRPC5, ATP receptor stimulation caused significant production of nitric oxide (NO) derived from endothelial nitric oxide synthase (eNOS) and further [Ca²⁺]_i rises. RNA interference targeting bovine TRPC5 attenuated ATP-induced [Ca²⁺]_i rises and NO production in BAEC treated with RA. Furthermore, physical association between TRPC5 and eNOS at the caveolin-rich plasma membrane area was revealed by immunocytochemistry and co-immuneoprecipitation experiments. These results suggest that receptor-activated TRPC5 Ca²⁺ influx activity by coexpressed eNOS leads to an idea that NO generated by nearby eNOS activates TRPC5 in caveolae. There is a positive feedback cycle between TRPC5 and eNOS that amplifies NO production and [Ca²⁺]_i rises through TRPC5 nitrosylation. [Jpn J Physiol 55 Suppl:S6 (2005)]