Proc Natl Acad Sci U S A. 2008 Oct 14;105(41):15920-5. doi: 10.1073/pnas.0805062105. Epub 2008 Oct 3.
Syntaxin 5 regulates the endoplasmic reticulum channel-release properties of polycystin-2.
Geng, L., Boehmerle, W., Maeda, Y., Okuhara, D. Y., Tian, X., Yu, Z., Choe, C. U., Anyatonwu, G. I., Ehrlich, B. E., Somlo, S.,
["Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06510, USA."]
["Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06510, USA."]
Polycystin-2 (PC2), the gene product of one of two genes mutated in dominant polycystic kidney disease, is a member of the transient receptor potential cation channel family and can function as intracellular calcium (Ca(2+)) release channel. We performed a yeast two-hybrid screen by using the NH(2) terminus of PC2 and identified syntaxin-5 (Stx5) as a putative interacting partner. Coimmunoprecipitation studies in cell lines and kidney tissues confirmed interaction of PC2 with Stx5 in vivo. In vitro binding assays showed that the interaction between Stx5 and PC2 is direct and defined the respective interaction domains as the t-SNARE region of Stx5 and amino acids 5 to 72 of PC2. Single channel studies showed that interaction with Stx5 specifically reduces PC2 channel activity. Epithelial cells overexpressing mutant PC2 that does not bind Stx5 had increased baseline cytosolic Ca(2+) levels, decreased endoplasmic reticulum (ER) Ca(2+) stores, and reduced Ca(2+) release from ER stores in response to vasopressin stimulation. Cells lacking PC2 altogether had reduced cytosolic Ca(2+) levels. Our data suggest that PC2 in the ER plays a role in cellular Ca(2+) homeostasis and that Stx5 functions to inactivate PC2 and prevent leaking of Ca(2+) from ER stores. Modulation of the PC2/Stx5 interaction may be a useful target for impacting dysregulated intracellular Ca(2+) signaling associated with polycystic kidney disease.
PMID: 18836075
 Screening
Validation: In vitro validation
Validation: In vivo validation
Characterization
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Screening
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| TRP channel | Interactor | Method | Species | Region | Species | Organ/tissue | Sample type | |||
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Syntaxin5 | Yeast two-hybrid | Human | 3-213 | Mouse | Embryonic | cDNA library | 18836075 | |
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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 | ||
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Syntaxin5 | Fusion protein-pull down assay | Human | 3-213 | E. coli | Mouse | Full-length | Not specified | 18836075 |
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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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| 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 | ||||
| TRPP1 |
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Syntaxin5 | Co-immunoprecipitation | LLC-PK1 | Human | Full-length | Mouse | Full-length | 18836075 | |
| TRPP1 |
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Syntaxin5 | Co-immunofluorescence staining | LLC-PK1 | Human | Full-length | Mouse | Full-length | 18836075 | |
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Screening
Validation: In vitro validation
Validation: In vivo validation
Characterization
Functional consequence
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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 | ||||
| TRPP1 |
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Syntaxin5 | Fusion protein-pull down assay | Human | Not determined | Mouse | 256-333 | 18836075 | ||
| TRPP1 |
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Syntaxin5 | Co-immunoprecipitation | Human | 5-72 | Mouse | Not determined | 18836075 | ||
(:
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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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| Functional consequence
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| TRP channel | Interactor | Method | Post-translational modification | Subcellular trafficking | Activity | Reference | ||||||
| TRPP1 |
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Syntaxin5 | Patch clamp | Inhibition | 18836075 | |||||||
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:
click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)