Repression of bone morphogenetic protein 4 by let-7i attenuates mesenchymal migration of head and neck cancer cells
Highlights
► let-7i represses BMP4 expression through binding to 3′-UTR. ► Knockdown of BMP4 attenuates mesenchymal migration of head and neck cancer cells. ► let-7i expression inversely correlates with BMP4 in head and neck cancer samples.
Introduction
Head and neck cancer, including cancers arising from the oral cavity, oropharynx, hypopharynx and larynx, is one of the leading causes of cancer-related deaths worldwide [1]. More than 90% of head and neck cancers are squamous cell carcinoma (HNSCC), which is associated with smoking, drinking and betel nut chewing [2]. The clinical characteristics of HNSCC are distinct from cancers originating from other tissues/organs: advanced HNSCC is often associated with severe destruction of surrounding tissues, and locoregional recurrence is the major pattern of treatment failure. When disease recurs, the response rate to treatment reduces significantly. Intriguingly, compared with other types of cancers, the frequency of distant metastasis in HNSCC is relatively low [3]. However, understanding of the unique mechanism responsible for HNSCC local invasion is limited.
The three-dimensional (3D) cultivation system represents the living microenvironment and is superior to traditional culture systems for studying the migration of cancer cells [4], [5]. In 3D environments, epithelial cancer cells migrate using multi-cellular collective movement [6] or individual cell movement, the latter of which includes mesenchymal- or amoeboid-mode movement [7]. For mesenchymal-mode movement, the cells possess an elongated shape with protrusion, whereas cells moving in an amoeboid mode often reveal a round shape with membrane blebbing [8], [9]. Recent studies have suggested that mesenchymal-mode movement is important for local invasion; by contrast, the amoeboid-mode movement directs distant metastasis [10]. However, knowledge concerning controlling the switch between mesenchymal and amoeboid movement is limited.
MicroRNAs (miRNAs) are small, non-protein-coding RNA molecules that repress gene expression at the posttranscriptional level by partially base-pairing to the 3′-untranslated regions (3′-UTRs) of target mRNAs [11], [12]. The role of miRNAs in cancer metastasis has been highlighted recently [13], [14]. We recently discovered that, in HNSCC, the epithelial–mesenchymal transition (EMT) inducer Twist1 promotes mesenchymal-mode movement through repression of let-7i, a member of let-7 family microRNAs that are expressed during stem cell differentiation and that act as a tumor suppressor [15], [16], [17]. Repression of let-7i elicits mesenchymal-mode movement through activation of Ras-related C3 botulinum toxin substrate 1 (RAC1) [18]. In the present study, we further identified a novel let-7i target involved in regulating HNSCC migration in 3D environments. We found that bone morphogenetic protein 4 (BMP4), a TGF-β superfamily protein that plays a critical role in embryogenesis [19], is a target repressed by let-7i. Knockdown of BMP4 reduces mesenchymal-mode movement in HNSCC. The present study uncovers both a novel target of let-7i and a new function of BMP4 in cancer cells migration.
Section snippets
Cell lines and plasmids
The human HNSCC cell line FaDu and the human embryonic kidney cell line HEK-293T were obtained from the Bioresource Collection and Research Center of Taiwan. The HNSCC cell lines OECM-1, CAL-27, Ca-9 and HSC-3 were provided by Dr. Cheng-Chi Chang (National Taiwan University). OECM-1 and FaDu were cultivated in Roswell Park Memorial Institute (RPMI)-1640 medium with 10% heat-inactivated fetal bovine serum (FBS), and HEK-293T, Ca-9, HSC-3 and CAL-27 were cultured in Dulbecco’s Modified Eagle’s
The expression of let-7i inversely correlates with BMP4
We initially aimed to identify the novel let-7i target gene(s) that regulate(s) cellular migration because we previously found that let-7i represses mesenchymal-mode movement in HNSCC [18]. The cancer miRNA Regulatory Network (http://cmrn.systemsbiology.net/) [23] was applied to predict candidate targets of let-7i. We found that BMP4, a morphogenic protein that is critically involved in development and cancer progression [19], [24], [25], [26], [27], [28], [29], [30], [31], is a putative target
Discussion
BMP4 is a morphogenic protein that plays an important role in embryogenesis and stem cell development [19], [24]. The significance of BMP4 in cancer cells has been highlighted recently: increased expression of BMP4 has been shown in different types of cancers, including melanoma, ovarian carcinoma, gastric carcinoma, hepatocellular carcinoma, renal cell carcinoma and HNSCC [25], [26], [27], [28], [29], [30], [31], and increased BMP4 expression is associated with a worse outcome of HNSCC [29].
Acknowledgments
This work was supported by National Health Research Institutes (NHRI-EX100-10037BI to M.H.Y.), National Science Council (101-2321-B-010-007 to M.H.Y.), Taipei Veterans General Hospital (V102-E8-002; V102C-036 to M.H.Y.), a grant from Ministry of Education, Aim for the Top University Plan (to M.H.Y.), and a grant from Department of Health, Center of Excellence for Cancer Research (DOH101-TD-C-111-007 to M.H.Y).
References (33)
- et al.
Habitual risk factors for head and neck cancer
Otolaryngol. Head Neck Surg.
(2004) - et al.
Rac activation and inactivation control plasticity of tumor cell movement
Cell
(2008) - et al.
The plasticity of cytoskeletal dynamics underlying neoplastic cell migration
Curr. Opin. Cell Biol.
(2010) MicroRNAs: genomics, biogenesis, mechanism, and function
Cell
(2004)- et al.
Temporal regulation of metamorphic processes in Drosophila by the let-7 and miR-125 heterochronic microRNAs
Curr. Biol.
(2008) - et al.
Expression of bone morphogenetic proteins, the subfamily of the transforming growth factor-beta superfamily, in renal cell carcinoma
J. Urol.
(2007) - et al.
Bone morphogenetic protein-4-induced epithelial–mesenchymal transition and invasiveness through Smad1-mediated signal pathway in squamous cell carcinoma of the head and neck
Arch. Med. Res.
(2011) - et al.
Overexpression of bone morphogenetic protein 4 enhances the invasiveness of Smad4-deficient human colorectal cancer cells
Cancer Lett.
(2009) - et al.
Cancer statistics
CA Cancer J. Clin.
(2010) - et al.
Risk factors for distant metastases in head and neck squamous cell carcinoma
Arch. Otolaryngol. Head Neck Surg.
(2006)