Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The VSports app下载. gov means it’s official. Federal government websites often end in . gov or . mil. Before sharing sensitive information, make sure you’re on a federal government site. .

Https

The site is secure. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely V体育官网. .

. 2015;16(6):846-55.
doi: 10.1080/15384047.2015.1030545.

miR-33a functions as a tumor suppressor in melanoma by targeting HIF-1α

Jianda Zhou  1 , Dan Xu, Huiqing Xie, Jingtian Tang, Rui Liu, Jingjing Li, Shaohua Wang, Xiang Chen, Juan Su, Xiao Zhou, Kun Xia, Quanyong He, Jia Chen, Wei Xiong, Peiguo Cao, Ke Cao
Affiliations

miR-33a functions as a tumor suppressor in melanoma by targeting HIF-1α

Jianda Zhou (V体育官网) et al. Cancer Biol Ther. 2015.

Abstract

Background: Our previous findings showed that miR-33 expressed abnormally in clinical specimens of melanoma, but the exact molecular mechanism has not been elucidated. VSports手机版.

Object: To determine miR-33's roles in melanoma and confirm whether HIF-1α is a direct target gene of miR-33a V体育安卓版. .

Methods: First miR-33a/b expression levels were detected in HM, WM35, WM451, A375 and SK-MEL-1. Then lentiviral vectors were constructed to intervene miR-33a expression in melanoma cells V体育ios版. Cell proliferation, invasion and metastasis were detected. A375 cells mice model was performed to test the tumorigenesis of melanoma in vivo. Finally the dual reporter gene assay was carried out to confirm whether HIF-1α is a direct target gene of miR-33a. .

Results: MiR-33a/b exhibited a lower expression in WM35, WM451, A375 and SK-MEL-1 of the metastatic skin melanoma cell lines than that in HM. Then inhibition of miR-33a expression in WM35 and WM451 cell lines could promote cell proliferation, invasion and metastasis VSports最新版本. Conversely, increased expression of miR-33a in A375 cells could inhibit cellproliferation, invasion and metastasis. In vivo tests also confirmed that overexpression of miR-33a in A375 cells significantly inhibited melanoma tumorigenesis. Finally, we confirmed that HIF-1α is a direct target gene of miR-33a. .

Conclusion: The newly identified miR-33a/HIF-1α axis might provide a new strategy for the treatment of melanoma V体育平台登录. .

Keywords: HIF-1α; HIF-1α, hypoxia inducible factor 1, α subunit; cell proliferation; invasion; melanoma; metastasis; miR-33a; miR-33a/b, microRNA 33a/b; microRNAs VSports注册入口. .

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
miR-33a/b was downregulated in metastatic melanoma cell lines: real-time PCR assay was used to detect miR-33a / b expression in melanoma cell lines with different metastatic potentials. (A) miR-33a in WM35, WM451, A375 and SK-MEL-1 was significantly reduced, normalized with that in HM, respectively (*P < 0.05). (B) miR-33b in WM35, WM451, A375 and SK-MEL-1 was significantly reduced, normalized with that in HM, respectively (*P < 0.05).
Figure 2.
Figure 2.
miR-33a was down-regulated or overexpressed in melanoma cells. (A) Real-time PCR detection of the relative expression level of miR-33a in pYr-LVX-miR-33a-sponge-transfected WM35 and WM451 as compared with that after transfection with the blank vector (*P < 0.05). Real-time PCR detect the miR-33a relative expressionin pYr-LVX-pri-miR-33a-transfected as compared with that in empty vector-transfected A375 (*P < 0.05). (B) When miR-33a was downregulated, HIF-1α protein level in WM35 and WM451 cells was significantly increased; when miR-33a overexpressed, HIF-1α protein level in A375 cells was significantly reduced. (C) Relative expression levels of HIF-1α protein in corresponding cells, normalized with that in WM35 cells (*P <0 .05).
Figure 3.
Figure 3.
miR-33a suppresses melanoma cell proliferation. MTT assay and colony formation assay were performed for miR-33a effects on cell proliferation and clone formation. In colony formation test, there were 1000 cells per well. (A) WM35 cells transfected with pYr-LVX-miR-33a-sponge had a stronger proliferation ability than those transfected with blank vector, and the colony formation efficiency also increased after transfection with pYr-LVX-miR-33a-sponge (*P < 0.05). (B) The proliferative ability and colony formation efficiency were significantly higher in pYr-LVX-miR-33a-sponge-transfected WM451 cells than the blank vector group (*P < 0.05). (C) The proliferative ability and colony formation efficiency of pYr-LVX-pri-miR-33a-transfected A375 cells was significantly lower than those transfected with the blank vector (*P < 0.05).
Figure 4.
Figure 4.
miR-33a inhibits melanoma cell migration. Cell scratch test and Transwell assay were used to detect miR-33a effect on melanoma cell migration. (A) WM35 cells transfected with pYr-LVX-miR-33a-sponge had a higher cell migration ability and more cells passing through the Transwell chamber as compared with those transfected with blank vector (*P < 0.05). (B) WM451 cells transfected with pYr-LVX-miR-33a-sponge had a higher cell migration ability and more cells passing through the Transwell chamber as compared with those transfected with blank vector (*P < 0.05). (C) The cell migration ability of pYr-LVX-pri-miR-33a-transfected A375 cells was significantly decreased as compared with the blank vector group, as well as the number of cells passing through the Transwell chamber significantly reduced (*P < 0.05).
Figure 5.
Figure 5.
miR-33a reduces in vivo growth and metastasis of A375 xenografts. miR-33a effects on in vivo growth of A375 xenografts in nude mice were detected through subcutaneous tumor implantation. (A) Left figure shows tumor formation in nude mice of the blank vector group and pYr-LVX-pir-miR-33a group. Right figure shows the excision tumor of A375 xenografts in the blank vector and pYr-LVX-pir-miR-33a groups (*P < 0.05). (B) This figure show the tumor volume of excision tumor by the formula V(mm3) = 0.5 × a × b2(a: maximum length to diameter, b: maximum transverse diameter) (*P < 0.05). (C) This figure show the tumor weight of excision tumor (*P < 0.05). (D) Mice were injected i.v. with melanoma cells to determine their metastatic activity. The lung tissue of mice in pYr-LVX-pir-miR-33a group was normal, whereas the lung tissue of mice in the blank vector group remarkably manifested melanoma colonies (*P < 0.05).
Figure 6.
Figure 6.
miR-33a directly targets HIF-1α. (A) pYr-Mir-Target-HIF-1α-3U luciferase report plasmid profile. The left is HIF-1α 3'UTR (NM_001530) sequence inserted into pYr-Mir-Target-HIF-1-3U dual luciferase reporter plasmid. Red area is the predicted loci which hsa-miR-33a binds to. The right is pYr-Mir-Target-HIF-1-3U dual luciferase reporter plasmid sketch. (B) Real-time PCR detection of miR-33a relative expression levels in WM35 and 293 cells. When miR-33a overexpressed in 293 cells, the dual luciferase activity decreased; while when endogenous miR-33a was down-regulated in 293 cells, the dual luciferase activity increased (*P < 0.05).
See this image and copyright information in PMC

References

    1. Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell 2011; 144:646-74; PMID:21376230; http://dx.doi.org/10.1016/j.cell.2011.02.013 - DOI - PubMed
    1. Segura MF, Greenwald HS, Hanniford D, Osman I, Hernando E. MicroRNA and cutaneous melanoma: from discovery to prognosis and therapy. Carcinogenesis 2012; 33:1823-32; PMID:22693259; http://dx.doi.org/10.1093/carcin/bgs205 - DOI - PMC - PubMed
    1. Dong F, Lou D. MicroRNA-34b/c suppresses uveal melanoma cell proliferation and migration through multiple targets. Mol Vis 2012; 18:537-46; PMID:22419847 - "V体育官网入口" PMC - PubMed
    1. Luo C, Tetteh PW, Merz PR, Dickes E, Abukiwan A, Hotz-Wagenblatt A, Holland-Cunz S, Sinnberg T, Schittek B, Schadendorf D, et al.. miR-137 inhibits the invasion of melanoma cells through downregulation of multiple oncogenic target genes. J Invest Dermatol 2013; 133:768-75; PMID:23151846; http://dx.doi.org/10.1038/jid.2012.357 - DOI - PubMed
    1. Dar AA, Majid S, Rittsteuer C, de Semir D, Bezrookove V, Tong S, Nosrati M, Sagebiel R, Miller JR 3rd, Kashani-Sabet M. The role of miR-18b in MDM2-p53 pathway signaling and melanoma progression. J Natl Cancer Inst 2013; 105:433-42; PMID:23365201; "V体育2025版" http://dx.doi.org/10.1093/jnci/djt003 - DOI - PMC - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources