Fig. 5: STAT3 governs the transcriptional switch in the TFF3-stimulated oncogenic transformation of immortalized-HMECs. | Cell Death & Disease

Fig. 5: STAT3 governs the transcriptional switch in the TFF3-stimulated oncogenic transformation of immortalized-HMECs.

From: Expression of two non-mutated genetic elements is sufficient to stimulate oncogenic transformation of human mammary epithelial cells

Fig. 5

a Luciferase activity of the CCND1 promoter in immortalized-HMECs with forced expression of TFF3 and their vector control cells after transient-transfection of STAT3 DN or on exposure to JSI-124 (0.2 µM) or Stattic (2 µM) inhibitor. b Luciferase activity of the BCL2 promoter in immortalized-HMECs with forced expression of TFF3 and their vector control cells after transient-transfection of STAT3 DN or on exposure to JSI-124 (0.2 µM) or Stattic (2 µM) inhibitor. c Western blot analysis was used to assess the protein levels of CCND1, CDK4, CCNE1, CDK2, and BCL2 in immortalized-HMECs with forced expression of TFF3 and their vector control cells, after inhibition of STAT3. Inhibition of STAT3 executed using transient-transfection of STAT3 DN or on exposure to JSI-124 (0.2 µM) or Stattic (2 µM) inhibitor. Densitometric analysis demonstrated that HMEC-hTERT-TFF3 cells exhibited increased protein levels of CCND1 (49 ± 8%), CCNE1 (95 ± 18%), and BCL2 (114 ± 17%) compared to HMEC-hTERT-vector cells. MCF10A-TFF3 cells exhibited increased protein levels of CCND1 (4902 ± 263%), CCNE1 (125 ± 27%), and BCL2 (79 ± 16%) compared to MCF10A-vector cells. MCF12A-TFF3 cells exhibited increased protein levels of CCND1 (92 ± 25%), CCNE1 (47 ± 12%), and BCL2 (137 ± 25%) compared to MCF12A-vector cells. Soluble whole cellular extracts were run on an SDS-PAGE and immunoblotted as described in materials and methods. β-actin was used as an input control for cell lysate. The sizes of detected protein bands in kDa are shown on the right side. d Correlation between mRNA levels of TFF3 and CCND1 or BCL2 in the MC cohort. Pearson’s χ2-test was used to compare the differences between groups. Statistical significance was assessed by using an unpaired two-tailed Student’s t-test (P < 0.05 was considered as significant) using GraphPad Prism 5. The luciferase assay was performed as described in Material and Methods. The column is mean of triplicate experiments; bars, ±SD. **P < 0.001, *P < 0.05

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