Dynamic interaction of hTRPC6 with the Orai1-STIM1 complex or hTRPC3 mediates its role in capacitative or non-capacitative Ca2+ entry pathways

TRPC (canonical transient receptor potential) channel subunits have been shown to assemble into homo- or hetero-meric channel complexes, including different Ca2+-handling proteins, required for the activation of CCE (capacitative Ca2+ entry) or NCCE (non-CCE) pathways. In the present study we found evidence for the dynamic interaction between endogenously expressed hTRPC6 (human TRPC6) with either both Orai 1 and STIMI (stromal interaction molecule I) or hTRPC3 to participate in CCE or NCCE. Electrotransjection of cells with all anti-hTRPC6 antibody, directed towards the C-terminal region, reduces CCE induced by TPEN [N,N,',N'-tetrakis-(2-pyridylmethyl)-ethylenediamine], which reduces the intraluminal free Ca2+ concentration. Cell stimulation with thrombin or extensive Ca2+-store depletion by TG (thapsigargin) + ionomycin enhanced the interaction between hTRPC6 and the CCE proteins Oral I and STIM1 I. In contrast, stimulation with the diacylglycerol analogue OAG (1-oleoyl-2-acetyl-sn-glycerol) displaces hTRPC6 from Orai1 and STIM1 and enhances the association between hTRPC6 and hTRPC3. The interaction between hTRPC6 and hTRPC3 was abolished by dinlethyl-BAP"I'A [1.2-bis(o-aminophenoxy)ethane-N,N,N'.N'-tetra-acetic acid) loading, which indicates that this phenomenon is Ca2+-dependent. These findings support the hypothesis that hTRPC6 participates both in CC E and NCCE through its interaction with tire Orai1-STIM1 complex or hTRPC3 respectively.