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. 2019 Jun 21;8:e47300. doi: 10.7554/eLife.47300

Figure 1. Frizzled CRD specificity for Reck-Gpr124-Wnt7a binding and signaling.

Figure 1.

(A) Reck(CC1-5)-AP binding to live HEK293T cells transfected with Gpr124, Wnt7a, and full-length Fz (top) or Gpr124, Wnt7a, and FzCRD-Myc-GPI (bottom). (B) Reck(CC1-5)-AP binding as in (A), with Gpr124, Wnt7a, and the indicated FzCRD-Myc-GPI targets, together with anti-Myc and anti-1D4 controls. (C) Summary of the AP binding assay in (A) and (B). (D) Reck(CC1-5)-AP binding as in (A), with WT or chimeric Frizzleds. (E) Beta-catenin signaling assay using STF cells transfected with Wnt7a, Gpr124, Reck, and Lrp5 (left) or Wnt7a and Lrp5 (right), together with WT or chimeric Frizzleds. Inset: summary of AP binding (D) and STF signaling (E). In this and subsequent figures, bars represent mean ± SD. Statistical significance, determined by the unpaired t-test, is represented by * (p<0.05), ** (p<0.01), *** (p<0.001), and **** (p<0.0001). The statistical comparisons in (E), (G), and (I) are to the ‘No Fz’ control. (F) AP-3xMyc-Norrin binding assay as in (A), with WT or chimeric Frizzleds. (G) Beta-catenin signaling assay using STF cells transfected with Tspan12 and Norrin (left) or Norrin (right), together with WT or chimeric Frizzleds. Inset: summary of AP binding (F) and STF signaling (G). (H) Schematic of the FzCRD-Myc-GPI competition experiment in (I). (I) The effect of FzCRD-Myc-GPI competition on beta-catenin signaling by Reck, Gpr124, and Wnt7a. Inset: summary of STF signaling.

Figure 1—source data 1. Relative luciferase activity for STF experiments in Figure 1 panels E, G, and I.
elife-47300-fig1-data1.xlsx (53.9KB, xlsx)
DOI: 10.7554/eLife.47300.003