TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells
Bibliographic record
Abstract
Erythropoietin (Epo) stimulates a significant increase in the intracellular calcium concentration ([Ca(2+)](i)) through activation of the murine transient receptor potential channel TRPC2, but TRPC2 is a pseudogene in humans. TRPC3 expression increases on normal human erythroid progenitors during differentiation. Here, we determined that erythropoietin regulates calcium influx through TRPC3. Epo stimulation of HEK 293T cells transfected with Epo receptor and TRPC3 resulted in a dose-dependent increase in [Ca(2+)](i), which required extracellular calcium influx. Treatment with the phospholipase C (PLC) inhibitor U-73122 or down-regulation of PLCgamma1 by RNA interference inhibited the Epo-stimulated increase in [Ca(2+)](i) in TRPC3-transfected HEK 293T cells and in primary human erythroid precursors, demonstrating a requirement for PLC. TRPC3 associated with PLCgamma, and substitution of predicted PLCgamma Src homology 2 binding sites (Y226F, Y555F, Y648F, and Y674F) on TRPC3 reduced the interaction of TRPC3 with PLCgamma and inhibited the rise in [Ca(2+)](i). Substitution of Tyr(226) alone with phenylalanine significantly reduced the Epo-stimulated increase in [Ca(2+)](i) but not the association of PLCgamma with TRPC3. PLC activation results in production of inositol 1,4,5-trisphosphate (IP(3)). To determine whether IP(3) is involved in Epo activation of TRPC3, TRPC3 mutants were prepared with substitution or deletion of COOH-terminal IP(3) receptor (IP(3)R) binding domains. In cells expressing TRPC3 with mutant IP(3)R binding sites and Epo receptor, interaction of IP(3)R with TRPC3 was abolished, and Epo-modulated increase in [Ca(2+)](i) was reduced. Our data demonstrate that Epo modulates TRPC3 activation through a PLCgamma-mediated process that requires interaction of PLCgamma and IP(3)R with TRPC3. They also show that TRPC3 Tyr(226) is critical in Epo-dependent activation of TRPC3. These data demonstrate a redundancy of TRPC channel activation mechanisms by widely different agonists.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.001 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".