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. 2021 Feb 19;40(9):e106113. doi: 10.15252/embj.2020106113

Figure 5. VE‐cadherin‐Y731 becomes phosphorylated before it reaches the cell surface and is masked by catenins.

Figure 5

  1. HUVEC were transduced with increasing titers of adenovirus expressing human SHP2. VE‐cadherin was immunoprecipitated from cell lysates and precipitates as well as total cell lysates were analyzed by immunoblots for indicated antigens.
  2. bEnd.3 cells were lysed, and VE‐cadherin or immature VE‐cadherin was immunoprecipitated, respectively, with C5 or VD47 antibodies. Precipitates were blotted against pY731‐VE‐cadherin, β‐catenin, plakoglobin, p120, α‐catenin, and VE‐cadherin.
  3. VE‐cadherin–catenin complex was immunoprecipitated from HUVEC lysates and was either left untreated (no SDS) or treated with 0.2% SDS for 30 min at room temperature, followed by removal of SDS and addition of 2 μg recombinant human SHP2 for 30 min at 30°C. Precipitates were analyzed in immunoblots for pY731‐VE‐cadherin, plakoglobin, β‐catenin, and VE‐cadherin.
  4. Quantification of pY731 blot signals of three independent experiments (from C), with the signal intensity for untreated VE‐cadherin set as 1. Molecular weight markers are indicated in kDa.

Data information: Immunoblots are representative of at least three (A, B‐right panel, C) or five (B‐left panel) independent experiments (mean ± SEM). Statistical significance was tested with unpaired two‐tailed t‐test, *P < 0.05, n.s., not significant.

Source data are available online for this figure.