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. 2016 Jun 21;7(30):48193–48205. doi: 10.18632/oncotarget.10194

Figure 1. MT2-MMP stable expression in HCT116 cells results in EMT.

Figure 1

A. Lentiviral MT2-MMP vector construction. The open reading frame (ORF) of human MT2-MMP was introduced at the AgeI/AgeI site in the GV287 vector containing Ubi-MCS-3FLAG-SV40-EGFP elements to generate stable transfectant cells. B. RT-PCR analysis of MT2-MMP mRNA levels in MT2-MMP or vector transfectant HCT116 cell lines. C. Western blot analysis of MT2-MMP protein expression in MT2-MMP transfectants, or vector control transfectants, using anti-Flag (1:1000) followed by HRP-conjugated secondary antibody. D. Immunofluorescence analysis of MT2-MMP expression (Alexa fluor 594, red; nuclei, DAPI staining, blue). Pos: 100% (the ratio of positive MT2-MMP transfectant cells). Scale bar, 50 μm. E. Morphological observations. Stable MT2-MMP transfectants had a fibroblast-like morphology, while vector control transfectants, inactive mutant E260A transfectants, and MT2-MMP transfectants treated with the MMP inhibitor GM6001 (25 μM) all showed typical epithelial morphology. Scale bar, 100 μm. F. RT-PCR analysis of MMP3, 7, 9, MT1- and MT2-MMP in HCT116 cell lines transfected with vector or MT2-MMP.