J Biol Chem. 2003 Jul 18;278(29):27208-15. Epub 2003 May 5.
Caveolin-1 contributes to assembly of store-operated Ca2+ influx channels by regulating plasma membrane localization of TRPC1.
Brazer, S. C., Singh, B. B., Liu, X., Swaim, W., Ambudkar, I. S.,
["Secretory Physiology Section, Gene Therapy and Therapeutics Branch, Department of Health and Human Services, NIDCR, National Institutes of Health, Bethesda, Maryland 20892, USA."]
["Secretory Physiology Section, Gene Therapy and Therapeutics Branch, Department of Health and Human Services, NIDCR, National Institutes of Health, Bethesda, Maryland 20892, USA."]
TRPC1, a component of store-operated Ca2+ entry (SOCE) channels, is assembled in a complex with caveolin-1 (Cav1) and key Ca2+ signaling proteins. This study examines the role of Cav1 in the function of TRPC1. TRPC1 and Cav1 were colocalized in the plasma membrane region of human submandibular gland and Madin-Darby canine kidney cells. Full-length Cav1 bound to both the N and C termini of TRPC1. Amino acids 271-349, which includes a Cav1 binding motif (amino acids 322-349), was identified as the Cav1 binding domain in the TRPC1 N terminus. Deletion of amino acids 271-349 or 322-349 prevented plasma membrane localization of TRPC1. Importantly, TRPC1Delta271-349 induced a dominant suppression of SOCE and was associated with wild-type TRPC1. Although the role of the C-terminal Cav1 binding domain is not known, its deletion did not affect localization of TRPC1 (Singh, B. B., Liu, X., and Ambudkar, I. S. (2000) J. Biol. Chem. 275, 36483-36486). Further, expression of a truncated Cav1 (Cav1Delta51-169), but not full-length Cav1, similarly disrupted plasma membrane localization of endogenously and exogenously expressed TRPC1 in human submandibular gland and Madin-Darby canine kidney cells. Cav1Delta51-169 also suppressed thapsigarginand carbachol-stimulated Ca2+ influx and increased the detergent solubility of TRPC1, although plasma membrane lipid raft domains were not disrupted. These data demonstrate that plasma membrane localization of TRPC1 depends on an interaction between its N terminus and Cav1. Thus, our data suggest that Cav1 has an important role in the assembly of SOCE channel(s).
PMID: 12732636
 Screening
Validation: In vitro validation
Validation: In vivo validation
Characterization
Functional consequence
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Validation: In vitro validation
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| Assay with recombinant proteins | Reference | |||||||||
| TRP channel construct | Interactor construct | |||||||||
| TRP channel | Interactor | Method | Species | Region | Expression system | Species | Region | Expression system | ||
| TRPC1 |
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Caveolin-1 | Fusion protein-pull down assay | Human | 1-347 | E. coli | Not specified | Full-length | HSG cell lysates | 12732636 |
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click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
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click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
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Screening
Validation: In vitro validation
Validation: In vivo validation
Characterization
Functional consequence
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top
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| Validation: In vivo validation
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| Assay with endogenous proteins | Assay with overexpressed proteins | Reference | ||||||||
| Cell or tissue | Cell or tissue | TRP channel construct | Interactor construct | |||||||
| TRP channel | Interactor | Method | Species | Region | Species | Region | ||||
| TRPC1 |
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Caveolin-1 | Co-immunofluorescence staining | HSG cell | Not used | Not specified | Full-length | 12732636 | ||
| TRPC1 |
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Caveolin-1 | Co-immunofluorescence staining | HSG cell | Human | Full-length | Not specified | Full-length | 12732636 | |
| TRPC1 |
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Caveolin-1 | Co-immunofluorescence staining | MDCK | Human | Full-length | Not specified | Full-length | 12732636 | |
| TRPC1 |
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Caveolin-1 | Co-immunoprecipitation | MDCK | Human | Full-length | Not specified | Full-length | 12732636 | |
(:
click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
:
click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
|
Screening
Validation: In vitro validation
Validation: In vivo validation
Characterization
Functional consequence
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top
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| Characterization
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| Binding region mapping | Stoichiometry | Affinity (Kd) | Reference | |||||||
| TRP channel | Interactor | |||||||||
| TRP channel | Interactor | Method | Species | Region | Species | Region | ||||
| TRPC1 |
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Caveolin-1 | Yeast two-hybrid | Human | 271-349 | Not specified | Not determined | 12732636 | ||
(:
click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
:
click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)