Figure 1.
Figure 1. TpoR contains a cytoplasmic KWQFP motif that maintains the unliganded receptor inactive. (A) The TpoR JM domain contains 5 additional residues that show no homology to the EpoR (shown in red italics). To construct theΔ5TpoR, the KWQFP motif in the TpoR cytosolic JM domain was deleted by PCR mutagenesis. (B) Proliferation assays performed in Ba/F3 cells expressing the wtTpoR or Δ5TpoR at equal GFP levels in the absence of any cytokines or stimulated with 5 ng/mL Tpo as indicated. Cell numbers (averages of triplicates ± SD) were counted at day 12. (C) Total protein levels of the wtTpoR or Δ5TpoR in Ba/F3 cells infected to GFP levels at 40% to 50%, as revealed by Western blotting using anti-HA antibodies. (D) HA staining for cell-surface levels of the wtTpoR (red) or Δ5TpoR (blue) in Ba/F3 or hematopoietic progenitor cells infected to GFP levels at 40%. Green indicates Ba/F3 cells (negative control).

TpoR contains a cytoplasmic KWQFP motif that maintains the unliganded receptor inactive. (A) The TpoR JM domain contains 5 additional residues that show no homology to the EpoR (shown in red italics). To construct theΔ5TpoR, the KWQFP motif in the TpoR cytosolic JM domain was deleted by PCR mutagenesis. (B) Proliferation assays performed in Ba/F3 cells expressing the wtTpoR or Δ5TpoR at equal GFP levels in the absence of any cytokines or stimulated with 5 ng/mL Tpo as indicated. Cell numbers (averages of triplicates ± SD) were counted at day 12. (C) Total protein levels of the wtTpoR or Δ5TpoR in Ba/F3 cells infected to GFP levels at 40% to 50%, as revealed by Western blotting using anti-HA antibodies. (D) HA staining for cell-surface levels of the wtTpoR (red) or Δ5TpoR (blue) in Ba/F3 or hematopoietic progenitor cells infected to GFP levels at 40%. Green indicates Ba/F3 cells (negative control).

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