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MiR-143 and MiR-145 regulate IGF1R to suppress cell proliferation in colorectal cancer.

Su J, Liang H, Yao W, Wang N, Zhang S, Yan X, Feng H, Pang W, Wang Y, Wang X, Fu Z, Liu Y, Zhao C, Zhang J, Zhang CY, Zen K, Chen X, Wang Y - PLoS ONE (2014)

Bottom Line: We identified specific target sites for miR-143 and miR-145 (miR-143/145) in the 3'-untranslated region (3'-UTR) of the IGF1R gene.These miRNAs are members of a cluster of miRNAs that have been reported to exhibit tumor suppressor activity.We demonstrated that the repression of IGF1R by miR-143/145 suppressed the proliferation of Caco2 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, The First Affiliated Hospital of Anhui Medical University, Hefei, Anhui 230022, China.

ABSTRACT
Insulin-like growth factor 1 receptor (IGF1R) is a transmembrane receptor that is activated by insulin-like growth factor 1 (IGF-1) and by a related hormone called IGF-2. It belongs to the large class of tyrosine kinase receptors and plays an important role in colorectal cancer etiology and progression. In this study, we used bioinformatic analyses to search for miRNAs that potentially target IGF1R. We identified specific target sites for miR-143 and miR-145 (miR-143/145) in the 3'-untranslated region (3'-UTR) of the IGF1R gene. These miRNAs are members of a cluster of miRNAs that have been reported to exhibit tumor suppressor activity. Consistent with the bioinformatic analyses, we identified an inverse correlation between miR-143/145 levels and IGF1R protein levels in colorectal cancer tissues. By overexpressing miR-143/145 in Caco2, HT29 and SW480 colorectal cancer cells, we experimentally validated that miR-143/145 directly recognizes the 3'-UTR of the IGF1R transcript and regulates IGF1R expression. Furthermore, the biological consequences of the targeting of IGF1R by miR-143/145 were examined by cell proliferation assays in vitro. We demonstrated that the repression of IGF1R by miR-143/145 suppressed the proliferation of Caco2 cells. Taken together, our findings provide evidence for a role of the miR-143/145 cluster as a tumor suppressor in colorectal cancer through the inhibition of IGF1R translation.

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The effect of miR-143/145 on the proliferation of colorectal cancer cells.(A, E and I) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (A), HT29 (E) and SW480 (I) cells with a scrambled control, pre-miR-143, pre-miR-145 or both pre-miR-143 and pre-miR-145. (B, F and J) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (B), HT29 (F) and SW480 (J) cells with either a scrambled control siRNA or an IGF1R siRNA. (C, G and K) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (C), HT29 (G) and SW480 (K) cells with either a control vector or an IGF1R overexpression vector. (D, H and L) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (D), HT29 (H) and SW480 (L) cells with either a scrambled control, pre-miR-143/145 or both pre-miR-143/145 and the IGF1R overexpression vector. *P<0.05; **P<0.01.
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pone-0114420-g003: The effect of miR-143/145 on the proliferation of colorectal cancer cells.(A, E and I) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (A), HT29 (E) and SW480 (I) cells with a scrambled control, pre-miR-143, pre-miR-145 or both pre-miR-143 and pre-miR-145. (B, F and J) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (B), HT29 (F) and SW480 (J) cells with either a scrambled control siRNA or an IGF1R siRNA. (C, G and K) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (C), HT29 (G) and SW480 (K) cells with either a control vector or an IGF1R overexpression vector. (D, H and L) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (D), HT29 (H) and SW480 (L) cells with either a scrambled control, pre-miR-143/145 or both pre-miR-143/145 and the IGF1R overexpression vector. *P<0.05; **P<0.01.

Mentions: We next focused on studying the roles of miR-143/145 in IGF1R regulation. Because IGF1R is essential for the regulation of proliferation and cell cycle progression, we evaluated the effects of miR-143/145 on colorectal cancer cell proliferation using a CCK-8 assay. In support of the notion that miR-143/145 function as key cancer suppressed miRNAs [26], Caco2, HT29 and SW480 cells transfected with pre-miR-143/145 showed suppressed proliferation (Figure 3A, 3E and 3I). Furthermore, we assessed the role of IGF1R on cell proliferation. To knock down IGF1R, an siRNA targeting IGF1R was designed and transfected into Caco2, HT29 and SW480 cells. To overexpress IGF1R, a plasmid expressing the IGF1R ORF was transfected into Caco2, HT29 and SW480 cells. The efficient overexpression or knockdown of IGF1R is shown in Figure S1 in File S1. Consistent with previous studies showing that IGF1R promotes cell proliferation [27], Caco2, HT29 and SW480 cells transfected with the IGF1R overexpression plasmid proliferated at a significantly higher rate (Figure 3C, 3G and 3K), whereas IGF1R knockdown with the siRNA significantly suppressed proliferation (Figure 3B, 3F and 3J). Thus, the anti-proliferative effect of IGF1R knockdown was similar to miR-143/145 overexpression. Furthermore, we constructed a IGF1R overexpression plasmid resistant to miR-143/145 (by elimination of its 3′-UTR). Compared to cells transfected with pre-miR-143/145, the cells transfected with pre-miR-143/145 and the IGF1R overexpression plasmid exhibited significantly higher proliferation rates (Figure 3D, 3H and 3L), suggesting that miR-143/145–resistant IGF1R rescued the suppression of IGF1R by miR-143/145 and attenuated the anti-proliferative effect of miR-143/145.


MiR-143 and MiR-145 regulate IGF1R to suppress cell proliferation in colorectal cancer.

Su J, Liang H, Yao W, Wang N, Zhang S, Yan X, Feng H, Pang W, Wang Y, Wang X, Fu Z, Liu Y, Zhao C, Zhang J, Zhang CY, Zen K, Chen X, Wang Y - PLoS ONE (2014)

The effect of miR-143/145 on the proliferation of colorectal cancer cells.(A, E and I) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (A), HT29 (E) and SW480 (I) cells with a scrambled control, pre-miR-143, pre-miR-145 or both pre-miR-143 and pre-miR-145. (B, F and J) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (B), HT29 (F) and SW480 (J) cells with either a scrambled control siRNA or an IGF1R siRNA. (C, G and K) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (C), HT29 (G) and SW480 (K) cells with either a control vector or an IGF1R overexpression vector. (D, H and L) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (D), HT29 (H) and SW480 (L) cells with either a scrambled control, pre-miR-143/145 or both pre-miR-143/145 and the IGF1R overexpression vector. *P<0.05; **P<0.01.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4256231&req=5

pone-0114420-g003: The effect of miR-143/145 on the proliferation of colorectal cancer cells.(A, E and I) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (A), HT29 (E) and SW480 (I) cells with a scrambled control, pre-miR-143, pre-miR-145 or both pre-miR-143 and pre-miR-145. (B, F and J) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (B), HT29 (F) and SW480 (J) cells with either a scrambled control siRNA or an IGF1R siRNA. (C, G and K) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (C), HT29 (G) and SW480 (K) cells with either a control vector or an IGF1R overexpression vector. (D, H and L) CCK-8 viability assays were performed 12, 24, 48 and 72 h after the transfection of Caco2 (D), HT29 (H) and SW480 (L) cells with either a scrambled control, pre-miR-143/145 or both pre-miR-143/145 and the IGF1R overexpression vector. *P<0.05; **P<0.01.
Mentions: We next focused on studying the roles of miR-143/145 in IGF1R regulation. Because IGF1R is essential for the regulation of proliferation and cell cycle progression, we evaluated the effects of miR-143/145 on colorectal cancer cell proliferation using a CCK-8 assay. In support of the notion that miR-143/145 function as key cancer suppressed miRNAs [26], Caco2, HT29 and SW480 cells transfected with pre-miR-143/145 showed suppressed proliferation (Figure 3A, 3E and 3I). Furthermore, we assessed the role of IGF1R on cell proliferation. To knock down IGF1R, an siRNA targeting IGF1R was designed and transfected into Caco2, HT29 and SW480 cells. To overexpress IGF1R, a plasmid expressing the IGF1R ORF was transfected into Caco2, HT29 and SW480 cells. The efficient overexpression or knockdown of IGF1R is shown in Figure S1 in File S1. Consistent with previous studies showing that IGF1R promotes cell proliferation [27], Caco2, HT29 and SW480 cells transfected with the IGF1R overexpression plasmid proliferated at a significantly higher rate (Figure 3C, 3G and 3K), whereas IGF1R knockdown with the siRNA significantly suppressed proliferation (Figure 3B, 3F and 3J). Thus, the anti-proliferative effect of IGF1R knockdown was similar to miR-143/145 overexpression. Furthermore, we constructed a IGF1R overexpression plasmid resistant to miR-143/145 (by elimination of its 3′-UTR). Compared to cells transfected with pre-miR-143/145, the cells transfected with pre-miR-143/145 and the IGF1R overexpression plasmid exhibited significantly higher proliferation rates (Figure 3D, 3H and 3L), suggesting that miR-143/145–resistant IGF1R rescued the suppression of IGF1R by miR-143/145 and attenuated the anti-proliferative effect of miR-143/145.

Bottom Line: We identified specific target sites for miR-143 and miR-145 (miR-143/145) in the 3'-untranslated region (3'-UTR) of the IGF1R gene.These miRNAs are members of a cluster of miRNAs that have been reported to exhibit tumor suppressor activity.We demonstrated that the repression of IGF1R by miR-143/145 suppressed the proliferation of Caco2 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, The First Affiliated Hospital of Anhui Medical University, Hefei, Anhui 230022, China.

ABSTRACT
Insulin-like growth factor 1 receptor (IGF1R) is a transmembrane receptor that is activated by insulin-like growth factor 1 (IGF-1) and by a related hormone called IGF-2. It belongs to the large class of tyrosine kinase receptors and plays an important role in colorectal cancer etiology and progression. In this study, we used bioinformatic analyses to search for miRNAs that potentially target IGF1R. We identified specific target sites for miR-143 and miR-145 (miR-143/145) in the 3'-untranslated region (3'-UTR) of the IGF1R gene. These miRNAs are members of a cluster of miRNAs that have been reported to exhibit tumor suppressor activity. Consistent with the bioinformatic analyses, we identified an inverse correlation between miR-143/145 levels and IGF1R protein levels in colorectal cancer tissues. By overexpressing miR-143/145 in Caco2, HT29 and SW480 colorectal cancer cells, we experimentally validated that miR-143/145 directly recognizes the 3'-UTR of the IGF1R transcript and regulates IGF1R expression. Furthermore, the biological consequences of the targeting of IGF1R by miR-143/145 were examined by cell proliferation assays in vitro. We demonstrated that the repression of IGF1R by miR-143/145 suppressed the proliferation of Caco2 cells. Taken together, our findings provide evidence for a role of the miR-143/145 cluster as a tumor suppressor in colorectal cancer through the inhibition of IGF1R translation.

Show MeSH
Related in: MedlinePlus