MiR-107 function as a tumor suppressor gene in colorectal cancer by targeting transferrin receptor 1

Abstract

Background: While microRNAs (miRNAs) are known to play a critical role in the progression of colorectal cancer, the role of miR-107 remains unknown. We evaluated its role and explored the underlying mechanism.

Materials & methods: MTT, wound-healing, transwell migration and transwell invasion assays were performed to evaluate the role of miR-107 in SW629 cell proliferation, migration and invasion. Real time-PCR and dual-luciferase reporter gene, TFR1 overexpression and western blotting assays were used to explore the underlying mechanism.

Results: MiR-107 is downregulated in colorectal cancer tissues and several human colorectal cancer cell lines. Low miR-107 expression often indicates a poor survival rate for colorectal cancer patients. MiR-107 suppresses the proliferation, migration and invasion of SW620 cells by negatively regulating transferrin receptor 1 (TFR1).

Conclusion: MiR-107 suppresses the metastasis of colorectal cancer and could be a potential therapy target in colorectal cancer patients.

Keywords: Cancer progress; Colorectal cancer; Transferrin receptor 1 (TFR1); microRNA107 (miR-107).

Fig. 1

MiR-107 is upregulated in human colorectal cancer tissues. a The results of an RT-PCR assay for miR-107 expression in human colorectal cancer tissues and the corresponding normal mucosa tissues. The quantitative data are presented as the means ± SEM, n = 50. *** p < 0.001 compared with normal tissues. b Kaplan-Meier curves for overall survival for human colorectal cancer related to miR-107 expression. c The miR-107 expression in the normal human colon epithelial cell HCoEpiC and several human colorectal cancer cell lines. GAPDH was used for normalization of the miR-107 levels. Quantitative data are presented as the means ± SEM, n = 3. *** p < 0.001 compared with the HCoEpiC

Fig. 2

MiR-107 represses the proliferation, migration and invasion of SW620 cells. a MiR-107 suppresses the proliferation of SW620 cells. The cells were treated with NC mimic or miR-107 mimic for the times shown. Cell viabilities were detected using the MTT assay. b and c MiR-107 inhibits the horizontal migration of SW620 cells. The effect of miR-107 cells on horizontal migration was evaluated with a wound-healing assay. The confluent cells were starved with serum-free medium for 6 h and then scratched with 10-μl pipette tips. After washing with PBS, the cells were treated with or without miR-107 mimic for another 8 h. The images were taken at 0 h and 8 h in the same field with an Olympus IX70 inverted microscope. Representative images (100× magnification) and quantitative data are shown in (b and c), respectively. d and e MiR-107 supresses the vertical migration and invasion abilities of SW620 cells. The effect of miR-107 on vertical migration and invasion was assessed with transwell migration assay and tranwell invasion assay, respectively. Representative images (100× magnification) are shown in D and the quantitative data are shown in E. The data were analyzed with GraphPad Prism 5.0, and are presented as means ± SEM, n = 3. *** p < 0.001 compared with the control group

Fig. 3

MiR-107 directly targeting at TFR1. a The sequence of human miR-107 and the predicted binding sites with miR-107 within the TFR1 3′-UTR are shown. b Photos of western blots showing that miR-107 inhibited the expression of miR-107 in SW620 cells. SW620 cell were treated with NC mimic or miR-107 mimic for 24 h. Then, the cells were collected and subjected to western blotting assay. c MiR-107 suppressed the transcription of TFR1 mRNA. SW620 cells were co-transfected with luciferase plasmids containing wild-type TFR1 3′-UTR or mutational TFR1 3′-UTR. The cells were also treated with NC mimic or miR-107 mimic. The cells were collected and lysed to measure the relative luciferase activity. Quantitative data are presented as the means ± SEM. *** p < 0.001 compared with the NC mimic group. d The TFR1 mRNA level in human colorectal cancer and the corresponding mucosa tissues. Quantitative data are presented as the means ± SEM, n = 50. *** p < 0.001 compared with normal tissues. e The TFR1 expressions in human colorectal cancer lines and normal human colon epithelial cell HCoEpiC were analyzed with RT-PCR. The TFR1 expression in HCoEpiC was set as 100%. Quantitative data are presented as the means ± SEM. *** p < 0.001 compared with BEA-2B cells. f The miR-107 expression in human colorectal cancer tissues is negatively correlated with that of TFR1. R² stands for goodness of fit and p stands for significance of the slope

Fig. 4

TFR1 overexpression restored the miR-107-mediated inhibitory effect on SW620 cells. a TFR1 overexpression attenuated miR-107-induced suppressive effect on SW620 cells. SW620 cells were treated with NC mimic or miR-107 mimics for 24 h, and then the cell viabilities were determined with the MTT assay. b and c TFR1 overexpression restores the miR-107-mediated effect on the horizontal migration of SW620 cells. SW620 cells were transfected with NC vector or TFR1 vector, and then used for the wound-healing assay. Representative images (100× magnification) and quantitative data are shown in (b and c), respectively. d and e TFR1 overexpression attenuates the miR-107-mediated suppressive effect on the vertical migration and invasion of SW620 cells. The SW620 cells were transfected with NC vector or TFR1 vector, and then subjected to transwell migration and transwell invasion assays. Representative images (100 × magnification) and quantitative data are shown in (d and e), representatively. Quantitative data are presented as the means ± SEM. *** p < 0.01 and *** p < 0.001 compared with NC vector group, ^#p < 0.05 compared with NC vector+miR-107 mimic group