RhoC–ROCK but not RhoA are required for Hic-5–mediated matrix degradation. (A) TGF-β–stimulated MCF10A GFP cells or unstimulated mouse GFP–Hic-5–expressing MCF10A cells treated with the ROCK inhibitor Y27632 demonstrate inhibited matrix degradation. (B) Quantitation of the percentage of cells associated with matrix degradation. (C) RhoA and RhoC were efficiently depleted from the TGF-β–treated MCF10A GFP cells with two independent siRNAs. (D) RhoA and RhoC knockdown of TGF-β–treated MCF10A GFP cells plated on fluorescent gelatin. RhoC-depleted cells have inhibited ability to degrade matrix. (E) Quantitation of the percentage of cells associated with matrix degradation. (F) The GST-RBD pull-down assay demonstrates that RhoC activity is increased in TGF-β–treated MCF10A GFP cells, which is suppressed by depletion of TGF-β–induced endogenous Hic-5 using human siRNA (hHic-5). (G) RhoA and RhoC were depleted from the mouse GFP–Hic-5–expressing MCF10A cells. Molecular mass standards are indicated next to the gel blots in kilodaltons. (H) GFP–Hic-5 expressing cells treated with RhoA and RhoC RNAi plated on fluorescent gelatin. RhoC knockdown inhibits matrix degradation. (I) Quantitation of the percentage of cells associated with matrix degradation. Bars, 20 µm. Error bars represent the standard error of the mean. **, P < 0.005; ***, P < 0.0005.