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. 2013 Dec 26;289(7):4219–4232. doi: 10.1074/jbc.M113.518514

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FIGURE 5. — Kinase activity, but not phosphatase activity, is required for HRas^V12-dependent disruption of the PP5·ERK2 complex. A, HEK-293FT cells were transfected with wild type HA-ERK1 or wild type HA-ERK2 together with (+) or without (−) phosphatase-dead FLAG-PP5 (FLAG-PP5^PD) and HRas^V12. FLAG immune complexes (FLAG IPs) and cell lysates were analyzed by Western blotting using antibodies recognizing the indicated proteins. B, wild type HA-ERK1 (HA-ERK1^WT), kinase-dead HA-ERK1 (HA-ERK1^KD), wild type HA-ERK2 (HA-ERK2^WT), and kinase-dead HA-ERK2 (HA-ERK2^KD) were transfected into HEK-293FT cells alone or together with wild type FLAG-PP5 in the absence or presence of HRas^V12. FLAG IPs and cell lysates were analyzed as described in A. *, IgG heavy chain. The data are representative of experiments performed three (A) and five (B) independent times with similar results.