| RRC ID |
10925
|
| Author |
Wedel BJ, Vazquez G, McKay RR, St J Bird G, Putney JW Jr.
|
| Title |
A calmodulin/inositol 1,4,5-trisphosphate (IP3) receptor-binding region targets TRPC3 to the plasma membrane in a calmodulin/IP3 receptor-independent process.
|
| Journal |
J Biol Chem
|
| Abstract |
Conformational coupling with the inositol 1,4,5-trisphosphate (IP3) receptor has been suggested as a possible mechanism of activation of TRPC3 channels and a region in the C terminus of TRPC3 has been shown to interact with the IP3 receptor as well as calmodulin (calmodulin/IP3 receptor-binding (CIRB) region). Here we show that internal deletion of 20 amino acids corresponding to the highly conserved CIRB region results in the loss of diacylglycerol and agonist-mediated channel activation in HEK293 cells. By using confocal microscopy to examine the cellular localization of Topaz fluorescent protein fusion constructs, we demonstrate that this loss in activity is caused by faulty targeting of CIRB-deleted mutants to intracellular compartments. Wild type TRPC3 and mutants lacking a C-terminal predicted coiled coil region downstream of CIRB were targeted to the plasma membrane correctly in HEK293 cells and exhibited TRPC3-mediated calcium entry in response to agonist activation. Mutation of conserved YQ and MKR motifs to alanine within the CIRB region in TRPC3-Topaz, which would be expected to interfere with IP3 receptor and/or calmodulin binding, had no effect on channel function or targeting. Additionally, TRPC3 targets to the plasma membrane of DT40 cells lacking all three IP3 receptors and forms functional ion channels. These findings indicate that the previously identified CIRB region of TRPC3 is involved in its targeting to the plasma membrane by a mechanism that does not involve interaction with IP3 receptors.
|
| Volume |
278(28)
|
| Pages |
25758-65
|
| Published |
2003-7-11
|
| DOI |
10.1074/jbc.M303890200
|
| PII |
S0021-9258(19)75095-9
|
| PMID |
12730194
|
| MeSH |
Amino Acid Sequence
Animals
Calcium / metabolism
Calcium Channels / metabolism*
Calmodulin / metabolism
Carbachol / pharmacology
Cell Line
Chickens
Cholinergic Agonists / pharmacology
Conserved Sequence
Diglycerides / metabolism
Flow Cytometry
Gene Deletion
Humans
Inositol 1,4,5-Trisphosphate Receptors
Ion Channels / metabolism*
Kinetics
Microscopy, Confocal
Molecular Sequence Data
Mutation
Plasmids / metabolism
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Cytoplasmic and Nuclear / metabolism*
Sequence Homology, Amino Acid
TRPC Cation Channels
Time Factors
Transfection
|
| IF |
4.238
|
| Times Cited |
62
|
|
WOS Category
|
BIOCHEMISTRY & MOLECULAR BIOLOGY
|
| Resource |
| Human and Animal Cells |
DT40(RCB1464)
IP3R^(-) DT40(RCB1467) |
