TY - JOUR
T1 - Molecular analysis of a store-operated and 2-acetyl-sn-glycerol-sensitive non-selective cation channel
T2 - Heteromeric assembly of TRPC1-TRPC3
AU - Liu, Xibao
AU - Bandyopadhyay, Bidhan C.
AU - Singh, Brij B.
AU - Groschner, Klaus
AU - Ambudkar, Indu S.
PY - 2005/6/3
Y1 - 2005/6/3
N2 - We have reported that internal Ca2+ store depletion in HSY cells stimulates a nonselective cation current which is distinct from I CRAC in RBL cells and TRPC1-dependent ISOC in HSG cells (Liu, X., Groschner, K., and Ambudkar, I. S. (2004) J. Membr. Biol. 200, 93-104). Here we have analyzed the molecular composition of this channel. Both thapsigargin (Tg) and 2-acetyl-sra-glycerol (OAG) stimulated similar non-selective cation currents and Ca2+ entry in HSY cells. The effects of Tg and OAG were not additive. HSY cells endogenously expressed TRPC1, TRPC3, and TRPC4 but not TRPC5 or TRPC6. Immunoprecipitation of TRPC1 pulled down TRPC3 but not TRPC4. Conversely, TRPC1 co-immunoprecipitated with TRPC3. Expression of antisense TRPC1 decreased (i) Tg- and OAG-stimulated currents and Ca2+ entry and (ii) the level of endogenous TRPC1 but not TRPC4. Antisense TRPC3 similarly reduced Ca2+ entry and endogenous TRPC3. Yeast two-hybrid analysis revealed an interaction between NTRPC1 and NTRPC3 (CTRPC1-CTRPC3, CTRPC3-CTRPC1, or CTRPC1-NTRPC3 did not interact), which was confirmed by glutathione S-transferase (GST) pull-down assays (GST-NTRPC3 pulled down TRPC1 and vice versa). Expression of NTRPC1 or NTRPC3 induced similar dominant suppression of Tg- and OAG-stimulated Ca2+ entry. NTRPC3 did not alter surface expression of TRPC1 or TRPC3 but disrupted TRPC1-TRPC3 association. In aggregate, our data demonstrate that TRPC1 and TRPC3 co-assemble, via N-terminal interactions, to form a heteromeric store-operated non-selective cation channel in HSY cells. Thus selective association between TRPCs generate distinct store-operated channels. Diversity of store-operated channels might be related to the physiology of the different cell types.
AB - We have reported that internal Ca2+ store depletion in HSY cells stimulates a nonselective cation current which is distinct from I CRAC in RBL cells and TRPC1-dependent ISOC in HSG cells (Liu, X., Groschner, K., and Ambudkar, I. S. (2004) J. Membr. Biol. 200, 93-104). Here we have analyzed the molecular composition of this channel. Both thapsigargin (Tg) and 2-acetyl-sra-glycerol (OAG) stimulated similar non-selective cation currents and Ca2+ entry in HSY cells. The effects of Tg and OAG were not additive. HSY cells endogenously expressed TRPC1, TRPC3, and TRPC4 but not TRPC5 or TRPC6. Immunoprecipitation of TRPC1 pulled down TRPC3 but not TRPC4. Conversely, TRPC1 co-immunoprecipitated with TRPC3. Expression of antisense TRPC1 decreased (i) Tg- and OAG-stimulated currents and Ca2+ entry and (ii) the level of endogenous TRPC1 but not TRPC4. Antisense TRPC3 similarly reduced Ca2+ entry and endogenous TRPC3. Yeast two-hybrid analysis revealed an interaction between NTRPC1 and NTRPC3 (CTRPC1-CTRPC3, CTRPC3-CTRPC1, or CTRPC1-NTRPC3 did not interact), which was confirmed by glutathione S-transferase (GST) pull-down assays (GST-NTRPC3 pulled down TRPC1 and vice versa). Expression of NTRPC1 or NTRPC3 induced similar dominant suppression of Tg- and OAG-stimulated Ca2+ entry. NTRPC3 did not alter surface expression of TRPC1 or TRPC3 but disrupted TRPC1-TRPC3 association. In aggregate, our data demonstrate that TRPC1 and TRPC3 co-assemble, via N-terminal interactions, to form a heteromeric store-operated non-selective cation channel in HSY cells. Thus selective association between TRPCs generate distinct store-operated channels. Diversity of store-operated channels might be related to the physiology of the different cell types.
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U2 - 10.1074/jbc.C400492200
DO - 10.1074/jbc.C400492200
M3 - Article
C2 - 15834157
AN - SCOPUS:20444364145
VL - 280
SP - 21600
EP - 21606
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 22
ER -