Time course of β1 integrin knockdown and MMP-2 activation in MCF-7-MT1 cells. (A): MCF-7-MT1 cells were transfected with the indicated siRNAs. At the indicated time points, cell lysates were collected for analysis of β1 integrin levels by Western blot. (B): Zymogram: MCF-7 cells were transfected with the indicated siRNAs. rMMP-2 was added at serial time points after β1 integrin siRNA transfection and cells were treated where indicated with Col I. Conditioned medium was collected and MMP-2 activation was assessed by zymography. Semiquantitative densitometry was performed and is expressed as percent area. (C): MT1-MMP levels were reduced in β1 integrin - abrogated cells. MCF-7-MT1 cells were transfected with the indicated siRNAs, and protein levels of β1 integrin and MT1-MMP were examined by Western blot at 72 hours after transfection. (D): MT1-MMP densitometry was analyzed and is expressed in arbitrary units. (E): The indicated siRNAs were transfected into MCF-7-MT1 cells, incubated for 6 hours cells were then mounted and viewed by light microscopy. (F): Immunofluorescence analysis for β1 integrin and MT1-MMP at the MCF-7-MT1 cell surface. After 72 hours transfection, cells were plated on Teflon printed glass slides and treated with Col I where indicated. Cells were subjected to immunofluorescence and then viewed by confocal microscopy. (G): Ectopic MT1-MMP expression rescued suppression of Col I-induced MMP-2 activation by β1/6 integrin siRNA. MCF-7-MT1 cells were transfected with the indicated siRNAs, and subsequently transfected with MT1-MMP. Cells were then left untreated or treated with Col I. Efficiency of MT1-MMP inductions were determined by Western Blot. (H): MMP-2 activation was analyzed by zymography. Semi-quantitative densitometry is expressed in percent of area.