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miR-17-5p promotes human breast cancer cell migration and invasion through suppression of HBP1

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Abstract

MicroRNAs have been implicated in regulating diverse cellular pathways. Emerging evidence indicate that the miR-17-92 cluster may have a causal role in breast cancer tumorigenesis as a novel class of oncogenes, but the role of these miRNAs in breast cancer invasion and migration remains unexplored. The aims of this study were to verify the effect of miR-17-5p (an important member of the miR-17-92 cluster) on the invasive and migratory ability of breast cancer cells. The matching of miR-17-5p and HMG box-containing protein 1 (HBP1) was predicted by TargetScan and confirmed by DNA constructs and luciferase target assay. The expression levels of miR-17-5p and its candidate target-HBP1 in MCF7 and MDA-MB-231 breast cancer cells were measured by real-time PCR and western blotting. Effects of miR-17-5p in cell cycle progression, proliferation, invasion and migration were evaluated by flow cytometry assay, 3-(4,-dimethy -lthiazol-2-yl)-2,-diphenyl -tetrazoliumbromide assay, soft-agar colony formation assay, and transwell invasive and migratory assay, respectively. The results showed that miR-17-5p was highly expressed in high-invasive MDA-MB-231 breast cancer cells but not in low-invasive MCF-7 breast cancer cells. Over-expression of miR-17-5p in MCF-7 cells rendered them the invasive and migratory abilities by targeting HBP1/β-catenin pathway. On the other hand, down-regulation of endogenous miR-17-5p suppressed the migration and invasion of MDA-MB-231 cells in vitro. These findings suggest that miR-17-5p plays an important role in breast cancer cell invasion and migration by suppressing HBP1 and subsequent activation of Wnt/β-catenin.

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Abbreviations

HBP1:

HMG box-containing protein 1

miRNA:

microRNA

PTEN:

Phosphatase and tensin homolog

BIM:

Bcl-2 interacting mediator of cell death

H-DMEM:

High glucose of Dulbecco’s modified Eagle’s medium

MTT:

3-(4, -dimethy -lthiazol-2-yl)-2, -diphenyl -tetrazoliumbromide

BSA:

Bovine serum albumin

MKK4:

MAP kinase kinase 4

NMe1:

Non-metastatic cells 1

Timp1:

Tissue inhibitor of matrix metalloproteinase-1

NMe2:

Non-metastatic cells 2

KAI1:

CD82 molecule (metastasis suppressor Kangai-1 homolog)

SDF-1:

Stromal cell-derived factor 1

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Acknowledgments

This work was supported by grants from the “863 Projects” of Ministry of Science and Technology of People’s Republic of China (No. 2006AA02A109, 2006AA02A115), National Natural Science Foundation of China (No. 30830052, 30911130363, U0970181), Beijing Ministry of Science and Technology (No. D07050701350701) and major national science and technology project (No. 2008ZX09101-044, 2009ZX09503-025).

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Authors and Affiliations

  1. Institute of Basic Medical Sciences and School of Basic Medicine, Center of Excellence in Tissue Engineering, Chinese Academy of Medical Sciences and Peking Union Medical College, 5# Dongdansantiao, Beijing, People’s Republic of China

    Hongling Li, Chunjing Bian, Lianming Liao, Jing Li & Robert Chunhua Zhao

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Correspondence to Robert Chunhua Zhao.

Electronic supplementary material

Below is the link to the electronic supplementary material.

miR-17-5p mimic promoted MCF7 cells to form more colonies in soft agar (n=4) in general imaging (TIFF 17499 kb)

10549_2010_954_MOESM2_ESM.tif

miR-17-5p inhibitor suppressed the colony growth ability of 231 cells in soft agar (n=4) in general imaging (TIFF 15055 kb)

10549_2010_954_MOESM3_ESM.tif

Inhibition of endogenous miR-17-5p almost failed to suppresses activities of MCF7 cells in vitro (a) MTT assay and (b) colony formation assay showed the slight inhibition of MCF7 cell proliferation and colony formation by miR-17-5p knockdown. (c) and (d) Transwell invasion and migration assays on MCF7 cells indicated down-regulations of miR-17-5p failed to decrease cell invasive and migratory ability. Data represent the mean ±SD of at least 3 independent experiments *P<0.05, **P<0.01, ***P<0.001 (TIFF 21222 kb)

10549_2010_954_MOESM4_ESM.tif

HBP1, but not PTEN, is the direct target of miR-17-5p in breast cancer cells (a) Western blot and real-time PCR eliminated PTEN as a direct target of miR-17-5p in breast cancer cells. (b) The effect of miR-17-5p inhibitor on the luciferase activity of MCF7 cells (TIFF 1569 kb)

Si-HBP1 failed to affect colony formation in a semisolid medium (TIFF 21085 kb)

Si-HBP1 failed to reverse the colony formation of miR-17-5pI-transfected 231 cells in general imaging (TIFF 28845 kb)

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Li, H., Bian, C., Liao, L. et al. miR-17-5p promotes human breast cancer cell migration and invasion through suppression of HBP1. Breast Cancer Res Treat 126, 565–575 (2011). https://doi.org/10.1007/s10549-010-0954-4

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  • DOI: https://doi.org/10.1007/s10549-010-0954-4

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