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. 2014 Mar 22;5(9):2596–2607. doi: 10.18632/oncotarget.1841

Figure 1. HOXC8 enhances CDH11 transcription in breast cancer cell lines.

Figure 1

(A) Total RNA was isolated from MDA-MB-231 or Hs578T cells lentivirally transduced with empty or HOXC8 expression vector, and then subjected to qRT-PCR to measure the level of CDH11 mRNA. GAPDH mRNA was used as an internal control for standardization. Columns, means; bars, SEM; n = 3. *, P < 0.05. (B) MDA-MB-231 or Hs578T cells lentivirally transduced with empty or HOXC8 expression vector were lysed and cell lysates were subjected to Western blot to detect HOXC8, CDH11, and β-actin with the respective antibodies. (C) Overnight cultured MDA-MB-231, Hs578T, MCF7 and T47D were lysed and cell lysates subjected to Western blot to detect HOXC8, CDH11 and β-actin with the respective antibodies. (D) MDA-MB-231 and Hs578T cells were transfected with scrambled or HOXC8 siRNA for 4 days and then subjected to ChIP with control IgG or RP II polyclonal antibody. The immunoprecipitated chromatin DNA was analyzed by PCR (upper panel) or qRT-PCR (lower panel) with primers amplifying region near the transcription start site (TSS) of the CDH11 promoter. Columns, means; bars, SEM; n = 3. *, P < 0.05. (E) MDA-MB-231 or Hs578T cells were transfected with scrambled or HOXC8 siRNA for 4 days followed by adding 2μg/ml actinomycin to the culture. Total RNA was isolated at varying times and then subjected to qRT-PCR to measure the level of CDH11 mRNA. GAPDH mRNA was used as an internal control. The level of CDH11 mRNA without actinomycin treatment was considered as 100%. Values are means ± SEM; n = 3. (F) MDA-MB-231 or Hs578T cells lentivirally transduced with empty or HOXC8 expression vector were subjected to ChIP with either control IgG or RP II polyclonal antibody. The immunoprecipitated chromatin DNA was analyzed by PCR (upper panel) or qRT-PCR (lower panel). Columns, means; bars, SEM; n = 3. *, P < 0.05.