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. 2012 Feb 7;287(14):11533–11545. doi: 10.1074/jbc.M111.306340

FIGURE 2.

FIGURE 2.

MT1-MMP ubiquitination is dependent on Src-mediated phosphorylation of Tyr573 within the MT1-MMP ICD. A, MCF-7 cells were transfected as indicated with either pcDNA3.1, ubiquitin-HA, MT1-MYC, MT1-MYC-Y573A, MT1-MYC-T567A, or MT1-MYC-T567E. Cell extracts were immunoprecipitated with an anti-Myc antibody, and immunoprecipitates (IP) were immunoblotted with an anti-HA antibody. Input controls were detected with an anti-Myc, an anti-HA, and an anti-β-actin antibody. The semiquantitative analyses of band intensities of three different immunoblots are shown ± S.E. with p < 0.0001 (***) and p < 0.001 (**). ns, not significant. AU, arbitrary units. B, MCF-7 cells were transfected with ubiquitin-HA and MT1-MYC, and cells were treated for 18 h with 20 μm PP2 or for 30 min with 100 ng/ml phorbol 12-myristate 13-acetate. Cell extracts were immunoprecipitated with an anti-Myc antibody, and immunoprecipitates were immunoblotted with an anti-HA antibody. Input controls were detected with an anti-Myc, an anti-HA, or an anti-β-actin antibody. *, nonspecific band. DMSO, dimethyl sulfoxide; IB, immunoblot.