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. 2014 Sep 8;42(17):11011–11024. doi: 10.1093/nar/gku814

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© The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 1. — Control of uPA mRNA abundance by Fra-1 in ER⁻ breast cancer cell lines. (A) Relative abundances of Plau-001 mRNA in MCF-7, MDA-MB231, BT549, HS578T cells. mRNA abundances were assayed by RT-qPCR from total cellular RNA and normalized to S26 mRNA taken as an internal standard. The Plau-001/S26 mRNA ratio was arbitrarily set to 1 in MCF-7 cells. (B) Fra-1 protein levels in MCF-7, MDA-MB231, BT549 and HS578T cells. Fra-1 protein levels were compared by immunoblotting using a specific Fra-1 antiserum. Fra-1 is indicated by brackets. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was taken as an electrophoresis loading control. (C, E, G) RNAi-mediated depletion of Fra-1 protein in MDA-MB231 (C), BT549 (E) and HS578T cells (G). Cells were transfected with either a control siRNA (siCTL) or siRNAs directed against the Fra-1 coding sequence (siFra-1-cds) or the 3′ unstranslated region (siFra-1–3′UTR) and protein contents were analyzed by immunoblotting with the relevant antibodies 48 h later. (D, F, H) Plau-001 mRNA reduction in siRNA-transfected MDA-MB231- (D), BT549- (F) and HS578T cells (H). Plau-001 mRNA was assayed by RT-qPCR 48 h post-siRNA transfection in the same cells as in C, E and F. Plau-001 mRNA abundance was arbitrarily set to 1 in siCTL-transfected cells for calculation of mean values. In A, D, F and H, values are the means of four independent experiments and error bars indicate standard deviations. Results of the Student's paired t-test are indicated on the graphs.