Fig 1: Western blotting detects EMT-related protein expression levels after ARPC2 knockdown or overexpression. (A) MCF-7 cells were stably transfected with NC or pcDNA3.1-ARPC2, and the levels of p-Smad2 and Smad2/3 were assessed by western blotting. *P<0.05. (B) MCF-7 cells were treated with 10 ng/l SB43 or DMSO for 48 h, and the protein expression levels of EMT markers, p-Smad2 and total Smad2/3 protein were assessed by western blotting. *P<0.05. (C) Western blotting was performed to examine ARPC2, E-cadherin, and vimentin expression after 48 h of treatment with TGF-β in siCtrl and siARPC2. *P<0.05. (D) Effect of ARPC2 downregulation, analysis of the expression of ARPC2, E-cadherin, vimentin, MMP-9, MMP-3 and ZEB-1 by western blotting. *P<0.05. (E) Effect of ARPC2 overexpression. Analysis of the level of ARPC2, E-cadherin, vimentin, MMP-9, MMP-3 and ZEB-1 by western blotting. *P<0.05. (F) ARPC2 mRNA expression levels in MDA-MB-231 cells treated with different concentrations of TGF-β at 48 h and (10 ng/ml) TGF-β at different time-points. EMT, epithelial-mesenchymal transition; ARPC2, actin-related protein 2/3 complex. *P<0.05, **P<0.01.
Fig 2: ARPC2 is upregulated in breast cancer cell lines and specimens from patients with breast cancer. (A) The level of ARPC2 mRNA was higher in breast cancer cell lines than MCF-10A non-malignant cells through qRT-PCR. **P<0.01. (B) ARPC2 protein expression in breast cancer cell lines was higher than in MCF-10A non-malignant cells. (C) Relative ARPC2 expression in breast cancer and normal breast tissues analyzed though IHC. Breast cancer cells feature strong-moderate cytoplasmic staining; normal tissues exhibit weak staining. (D) Kaplan-Meier overall survival curves generated in accordance to the ARPC2 protein levels of patients with breast cancer (low vs. high); ARPC2 was associated with worse overall survival. (E) Ratio of ARPC2 mRNA in different BrCa types was analyzed using TCGA database. ARPC2, actin-related protein 2/3 complex; TCGA, The Cancer Genome Atlas. The dots mean extreme outlier.
Fig 3: Effect of ARPC2 on MCF-7 cells in vivo. (A) The subcutaneous tumor growth curves. **P<0.01. (B) The quantification of the tumor weight. **P<0.01. (C) The representative images of subcutaneous implanted tumor cells. (D) Lung and lymphonodus sections were counterstained with H&E. (E) Immunohistochemical index Ki-67 and ARPC2-positively stained cells in the tissue sections of a xenograft model compared to the control group. ARPC2, actin-related protein 2/3 complex; H&E, hematoxylin and eosin.
Fig 4: Effect of ARPC2 on MCF-7 or MDA-MB-231 cells which were grown and transfected with siRNA-ARPC2 or pcDNA3.1-ARPC2. (A) ARPC2 expression level was confirmed at the transcription level by western blotting. Cells were transfected with siRNAs or plasmid targeting ARPC2 to knock down or induce ARPC2 expression. (B) ARPC2 mRNA expression level was confirmed by RT-PCR. *P<0.05. (C) Cell colony formation assay was performed in MDA-MB-231 or MCF-7 cells after transfection. **P<0.01. (D) A wound healing assay; the migrated cells were fixed and counted. Data represent the average of three experiments. **P<0.01. (E) Cell migration was examined by Boyden chamber assay and type I collagen-coated Transwell. **P<0.01. Cell invasion was examined using Matrigel-coated Transwell cell culture chambers. Ability of migration and invasion of cells were quantified by counting the number of cells that migrated or invaded the underside of the porous polycarbonate membrane under a phase-contrast microscope. ARPC2, actin-related protein 2/3 complex.
Supplier Page from Abcam for Anti-ARPC2 antibody