RGS2 inhibits the epithelial Ca2+ channel TRPV6.
The epithelial Ca(2+) channels TRPV5 and TRPV6 constitute the apical Ca(2+) entry pathway in the process of active Ca(2+) (re)absorption. By yeast two-hybrid and glutathione S-transferase pulldown analysis we identified RGS2 as a novel TRPV6-associated protein. RGS proteins determine the inactivation kinetics of heterotrimeric G-protein-coupled receptor (GPCR) signaling by regulating the GTPase activity of G(alpha) subunits. Here we demonstrate that TRPV6 interacts with the NH(2)-terminal domain of RGS2 in a Ca(2+)-independent fashion and that overexpression of RGS2 reduces the Na(+) and Ca(2+) current of TRPV6 but not that of TRPV5-transfected human embryonic kidney 293 (HEK293) cells. In contrast, overexpression of the deletion mutant DeltaN-RGS2, lacking the NH(2)-terminal domain of RGS2, in TRPV6-expressing HEK293 cells did not show this inhibition. Furthermore, cell surface biotinylation indicated that the inhibitory effect of RGS2 on TRPV6 activity is not mediated by differences in trafficking or retrieval of TRPV6 from the plasma membrane. This effect probably results from the direct interaction between RGS2 and TRPV6, affecting the gating properties of the channel. Finally, the scaffolding protein spinophilin, shown to recruit RGS2 and regulate GPCR-signaling via G(alpha), did not affect RGS2 binding and electrophysiological properties of TRPV6, indicating a GPCR-independent mechanism of TRPV6 regulation by RGS2.
Schoeber
Joost P
JP
Department of Physiology, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen 6500 HB, The Netherlands.
Topala
Catalin N
CN
Wang
Xinhua
X
Diepens
Robin J
RJ
Lambers
Tim T
TT
Hoenderop
Joost G
JG
Bindels
René J
RJ
D000818:Animals; D015220:Calcium Channels / Q000378:metabolism*; D002460:Cell Line; D004848:Epithelium / Q000378:metabolism*; D006801:Humans; D051379:Mice; D020710:RGS Proteins / Q000502:physiology*; D050916:TRPV Cation Channels / Q000037:antagonists & inhibitors* / Q000378:metabolism*
D015220:Calcium Channels; D020710:RGS Proteins; C087182:RGS2 protein, human; D050916:TRPV Cation Channels; C422827:TRPV6 protein, human