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. 2011 Feb;2(2):140–150. doi: 10.1177/1947601911408888

Figure 3.

Figure 3.

Low abundance of Drosha/DGCR8 leads to inefficient processing of uPA mRNA-targeted miRNAs in breast cancer cells. (A) Total RNA was isolated from MCF-7, MDA-MB-231, MDA-MB-436, and T47D cells and then subjected to qRT-PCR to measure the levels of primary, precursor, and mature forms of miR-193a, miR-193b, and miR-181a. The levels of GAPDH were also determined and serve as an internal control for standardization. Data are mean ± SE (n = 3). (B) Overnight-cultured MCF-7, MDA-MB-231, MDA-MB-436, and T47D cells were lysed, and cell lysates were subjected to immunoblotting to detect uPA, Drosha, DGCR8, Dicer, and β-actin with the respective antibodies. (C) MDA-MB-231 cells were transfected with empty vector, Drosha, and DGCR8 expressing vectors together or Dicer expression vector for 2 days. Total RNA was isolated from these cells, and qRT-PCR was performed to measure the levels of primary and mature forms of miR-193a, miR-193b, and miR-181a. The levels of GAPDH were also determined and serve as an internal control for standardization. Data are mean ± SE (n = 3). *P < 0.001 versus control. (D) Drosha/DGCR8 together or Dicer was forced to be expressed in MDA-MB-231 cells for 3 days. Cells were lysed, and cell lysates were analyzed for uPA protein levels by immunoblotting with uPA mAb.