Figure 5.

Interaction with TIMP3 is essential for ITGA7-mediated cell growth suppression. A: Mutant ITGA7 that does not interact with TIMP3 failed to suppress TNF-α and cyclin D1 expression and had no impact on NF-κB nuclear distribution. Lanes 1–8: Immunoblots were generated from the protein extracts from PITT1, PITT2, or pCDNA4-ITGA7^Δ1037-1103/pCDNA6TO transfected PC3 clones (C5 and C8), induced with or without tetracycline (Tet), using antibodies specific for ITGA7, TIMP3, TNF-α, cyclin D, or β-actin. ΔITGA7-FLAG denotes ITGA7-FLAG mutant that lacks amino acids 1037 to 1103 in ITGA7 protein. Lanes 9–24: Chromatin association of p65 NF-κB subunit. PITT1 and PITT2 cells were fractionated into chromatin (Chr) and non-chromatin (NC) fractions. Immunoblots were generated using antibodies specific for p65, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), or histone 3. B: Colony formation assays of replica samples of A. Triplicate experiments were performed on each assay. C: BrdU labeling and cell cycle analyses of replica samples of A. Triplicate experiments were performed on each assay. ^∗∗P < 0.01 (t-test).