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. 2016 Dec 21;292(4):1449–1461. doi: 10.1074/jbc.M116.768986

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FIGURE 5. — Both Rap1a and Rap1b contain putative 14-3-3 binding sites. A, HEK293 cells transfected with Myc-14-3-3 and either GFP-Rap1b (WT) or the phosphorylation mutant Rap1bAA (AA), constitutively active mutant Rap1bE63 (E63), or Rap1bE63AA (E63AA), were treated with F/I, as indicated. The levels of Myc-14-3-3 and GFP-Rap1 following GFP immunoprecipitation (IP) are shown. The levels of Myc-14-3-3 in the TCL are shown in the bottom panel. B, cells transfected with the constitutively active mutants Rap1bE63 or Rap1bE63AA were treated with F/I, as indicated. The levels of endogenous 14-3-3 and FLAG-Rap1 following FLAG IP are shown. The levels of endogenous 14-3-3 in the TCL are shown in the bottom panel. C, KSR^−/− MEF cells transfected with FLAG-Rap1b or vector and GFP-KSR were treated with F/I, as indicated. Endogenous 14-3-3 and FLAG-Rap1b following FLAG IP are shown. The levels of endogenous 14-3-3 and FLAG-RapE63, and GFP-KSR in the TCL are shown. D, HEK293 cells transfected with FLAG-RapE63 and vector and HA-14-3-3γ, HA-14-3-3β, and HA-14-3-3σ, and were treated with F/I, as indicated. HA-14-3-3 and GFP-Rap1b following FLAG IP are shown. The levels of HA-14-3-3 and GFP-RapE63 in the TCL are shown.