J Biol Chem. 2002 Sep 13;277(37):34375-82. Epub 2002 Jul 11.
Erythropoietin modulates calcium influx through TRPC2.
Chu, X., Cheung, J. Y., Barber, D. L., Birnbaumer, L., Rothblum, L. I., Conrad, K., Abrasonis, V., Chan, Y. M., Stahl, R., Carey, D. J., Miller, B. A.,
["Henry Hood Research Program, The Sigfried and Janet Weis Center for Research, Geisinger Clinic, 100 N. Academy Avenue, Danville, PA 17822, USA."]
["Henry Hood Research Program, The Sigfried and Janet Weis Center for Research, Geisinger Clinic, 100 N. Academy Avenue, Danville, PA 17822, USA."]
Mammalian isoforms of calcium-permeable Drosophila transient receptor potential channels (TRPC) are involved in the sustained phase of calcium entry in nonexcitable cells. Erythropoietin (Epo) stimulates a rise in intracellular calcium ([Ca](i)) via activation of voltage-independent calcium channel(s) in erythroid cells. Here, involvement of murine orthologs of classical TRPC in the Epo-modulated increase in [Ca](i) was examined. RT-PCR of TRPC 1-6 revealed high expression of only TRPC2 in Epo-dependent cell lines HCD-57 and Ba/F3 Epo-R, in which Epo stimulates a rise in [Ca](i). Using RT-PCR, Western blotting, and immunolocalization, expression of the longest isoform of mTRPC2, clone 14, was demonstrated in HCD-57 cells, Ba/F3 Epo-R cells, and primary murine erythroblasts. To determine whether erythropoietin is capable of modulating calcium influx through TRPC2, CHO cells were cotransfected with Epo-R subcloned into pTracer-CMV and either murine TRPC2 clone 14 or TRPC6, a negative control, into pQBI50. Successful transfection of Epo-R was verified in single cells by detection of green fluorescent protein from pTracer-CMV using digital video imaging, and successful transfection of TRPC was confirmed by detection of blue fluorescent protein fused through a flexible linker to TRPC. [Ca](i) changes were simultaneously monitored in cells loaded with Rhod-2 or Fura Red. Epo stimulation of CHO cells cotransfected with Epo-R and TRPC2 resulted in a rise in [Ca](i) above base line (372 +/- 71%), which was significantly greater (p < or = 0.0007) than that seen in cells transfected with TRPC6 or empty pQBI50 vector. This rise in [Ca](i) required Epo and extracellular calcium. These results identify a calcium-permeable channel, TRPC2, in erythroid cells and demonstrate modulation of calcium influx through this channel by erythropoietin.
PMID: 12167663
 Screening
Validation: In vitro validation
Validation: In vivo validation
Characterization
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Screening
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| Experimental screening | Non-experimental screening | Reference | ||||||||
| TRP channel construct | Interactor source | |||||||||
| TRP channel | Interactor | Method | Species | Region | Species | Organ/tissue | Sample type | |||
| TRPC2 |
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Epo-R | Inference | Prediction | 12167663 | |||||
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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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| 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 | ||||
| TRPC2 |
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Epo-R | Co-immunofluorescence staining | CHO | Mouse | Full-length | Mouse | Full-length | 12167663 | |
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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)
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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 | ||||||
| TRPC2 |
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Epo-R | Calcium measurement | Activation | 12167663 | |||||||
(:
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)