Skip to main page content (VSports在线直播)
U.S. flag

An official website of the United States government

Dot gov

The . gov means it’s official. Federal government websites often end in . gov or . mil VSports app下载. 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. .

. 2020 Apr 28;40(10):e00496-19.
doi: 10.1128/MCB.00496-19. Print 2020 Apr 28.

E2F6-Mediated Downregulation of MIR22HG Facilitates the Progression of Laryngocarcinoma by Targeting the miR-5000-3p/FBXW7 Axis

Hui Chen  1 , Mudunov Ali  2 , Azizyan Ruben  3 , Dimitry Stelmakh  3 , Maksim Pak  3
Affiliations

"VSports最新版本" E2F6-Mediated Downregulation of MIR22HG Facilitates the Progression of Laryngocarcinoma by Targeting the miR-5000-3p/FBXW7 Axis

VSports - Hui Chen et al. Mol Cell Biol. .

Abstract

Recently, abundant evidence has clarified that long noncoding RNAs (lncRNAs) play an oncogenic or anticancer role in the tumorigenesis and development of diverse human cancers. Described as a crucial regulator in some cancers, MIR22HG has not yet been studied in laryngocarcinoma and therefore the underlying regulatory role of MIR22HG in laryngocarcinoma is worth detecting. In this study, MIR22HG expression in laryngocarcinoma cells was confirmed to be downregulated, and upregulated MIR22HG expression led to suppressive effects on laryngocarcinoma cell proliferation and migration. Molecular mechanism assays revealed that MIR22HG sponges miR-5000-3p in laryngocarcinoma cells. Besides, decreased expression of miR-5000-3p suppressed laryngocarcinoma cell proliferation and migration. Moreover, the FBXW7 gene was reported to be a downstream target gene of miR-5000-3p in laryngocarcinoma cells. More importantly, rescue assays verified that FBXW7 depletion or miR-5000-3p upregulation countervailed the repressive effects of MIR22HG overexpression on laryngocarcinoma progression. In addition, E2F6 was proved to be capable of inhibiting MIR22HG transcription in laryngocarcinoma cells VSports手机版. To sum up, E2F6-induced downregulation of MIR22HG promotes laryngocarcinoma progression through the miR-5000-3p/FBXW7 axis. .

Keywords: E2F6; FBXW7; MIR22HG; laryngocarcinoma; miR-5000-3p. V体育安卓版.

PubMed Disclaimer

Figures

FIG 1
FIG 1
MIR22HG was expressed at low levels, and its upregulation inhibits cell proliferation and migration in laryngocarcinoma. (A) RT-qPCR analysis of MIR22HG expression in laryngocarcinoma cell lines (UM-SCC-10B, SNU899, and SNU46) and a normal nasopharyngeal epithelial cell line (NP69). (B) Measurement of the efficacy of MIR22HG overexpression in SNU46 and SNU899 cells by RT-qPCR. (C to E) Evaluation of cell proliferation ability in SNU46 and SNU899 cells transfected with pcDNA3.1/MIR22HG or pcDNA3.1 using CCK-8, colony formation, and EdU. (F and G) Evaluation of cell migratory ability in transfected cells by conducting wound healing and transwell assays. **, P < 0.01.
FIG 2
FIG 2
MIR22HG directly interacted with miR-5000-3p in laryngocarcinoma. (A) Detection the subcellular localization of MIR22HG in laryngocarcinoma cells with the application of subcellular fractionation and FISH assays. (B) miRNAs predicted to bind with MIR22HG based on the starBase database. (C) The binding capacity of MIR22HG and predicted miRNAs was analyzed via an RNA pulldown assay in SNU46 and SNU899 cells. (D) Binding site between MIR22HG and miR-5000-3p. (E) The interaction between MIR22HG and miR-5000-3p was confirmed by a luciferase reporter assay. (F) The effects of miR-5000-3p overexpression on MIR22HG were analyzed using qRT-PCR. (G) RT-qPCR analysis of miR-5000-3p expression in laryngocarcinoma cells and NP69 cells. (H) Analysis of miR-5000-3p expression in transfected cells via RT-qPCR. (I) RT-qPCR analysis of the efficacy of miR-5000-3p inhibition in SNU46 and SNU899 cells. (J and K) Evaluation of cell proliferative ability in SNU46 and SNU899 cells transfected with different plasmids utilizing CCK-8, colony formation, and EdU. (L) Analysis of cell migratory capability in different groups by applying wound healing and transwell assays. **, P < 0.01.
FIG 3
FIG 3
FBXW7 was the downstream target of miR-5000-3p in laryngocarcinoma. (A) Analysis overlaps illustrated by a Venn diagram. (B) RT-qPCR analysis of FBXW7 expression in SNU46 and SNU899 cells transfected with miR-5000-3p inhibitor or NC inhibitor. (C) RT-qPCR and Western blot analyses of FBXW7 expression in laryngocarcinoma cells and NP69 cells. (D) Measurement of FBXW7 expression in transfected cells through RT-qPCR and Western blotting. (E) FBXW7 expression and protein levels were appraised by RT-qPCR and Western blotting in cells with miR-5000-3p inhibitor or mimics. (F) The existence of RNAs (MIR22HG, miR-5000-3p and FBXW7) in RISC confirmed by RIPA. (G) Binding site between miR-5000-3p and FBXW7 predicted using starBase. (H) Interaction between miR-5000-3p and FBXW7 validated using a luciferase reporter assay. (I) Measurement of FBXW7 expression in different groups via RT-qPCR and Western blotting. **, P < 0.01.
FIG 4
FIG 4
miR-5000-3p inhibition recovered the impacts of MIR22HG silence on cell proliferation and migration. (A) The downregulation of MIR22HG in SNU46 cells was confirmed by qRT-PCR. (B to D) Cell proliferative abilities were examined by using CCK-8, colony formation, and EdU assays. (E and F) The migratory abilities were evaluated in wound healing and transwell assays. **, P < 0.01.
FIG 5
FIG 5
The suppressed function of MIR22HG-overexpressed SNU46 cells was normalized by enhanced miR-5000-3p. (A to C) Evaluation of cell proliferation ability in SNU46 cells transfected with different plasmids by conducting cell proliferation assays. (D and E) Analysis of cell migration capability in different groups by carrying out cell migration assays. **, P < 0.01.
FIG 6
FIG 6
MIR22HG affected laryngocarcinoma cell proliferation and migration by targeting FBXW7. (A) Measurement of the efficacy of FBXW7 knockdown in SNU46 cells via RT-qPCR. (B to D) Cell proliferation was assessed in rescue assays. (E and F) Cell migration was examined in rescue assays. (G) RT-qPCR detected the expression of miR-5000-3p in cells transfected with mutant MIR22HG (MIR22HG-M). (H) The effects of MIR22HG-M on proliferation and migration were analyzed by using colony formation and transwell assays. (I) RT-qPCR and Western blot analyses of mRNA and protein expression of mutant FBXW7 (FBXW7-M). (J) Colony formation and transwell assays revealed that FBXW7-M had no impact on miR-5000-3p-regulated proliferation and migration.**, P < 0.01; n.s., not significant.
FIG 7
FIG 7
E2F6 inhibited MIR22HG transcription in laryngocarcinoma. (A) RT-qPCR analysis of the efficacy of E2F6 overexpression in SNU46 and SNU899 cells. (B) RT-qPCR analysis of the expression of MIR22HG in transfected cells. (C) DNA motif of E2F6 obtained from JASPAR. (D) Two binding sites between E2F6 and MIR22HG promoter in SNU46 and SNU899 cells. (E and F) Binding sites between E2F6 and MIR22HG promoter analyzed by ChIP and luciferase reporter assays. **, P < 0.01.
See this image and copyright information in PMC

References

    1. McGuire S. 2016. World cancer report 2014—Geneva, Switzerland: World Health Organization, International Agency for Research on Cancer, WHO Press, 2015. Adv Nutr 7:418–419. - V体育安卓版 - PMC - PubMed
    1. Miller KD, Goding Sauer A, Ortiz AP, Fedewa SA, Pinheiro PS, Tortolero-Luna G, Martinez-Tyson D, Jemal A, Siegel RL. 2018. Cancer statistics for Hispanics/Latinos, 2018. CA Cancer J Clin 68:425–445. doi:10.3322/caac.21494. - DOI - PubMed
    1. Lampri ES, Chondrogiannis G, Ioachim E, Varouktsi A, Mitselou A, Galani A, Briassoulis E, Kanavaros P, Galani V. 2015. Biomarkers of head and neck cancer, tools or a gordian knot? Int J Clin Exp Med 8:10340–10357. - PMC - PubMed
    1. Li P, Liu H, Wang Z, He F, Wang H, Shi Z, Yang A, Ye J. 2016. MicroRNAs in laryngeal cancer: implications for diagnosis, prognosis and therapy. Am J Transl Res 8:1935–1944. - PMC - PubMed
    1. Hoffman HT, Porter K, Karnell LH, Cooper JS, Weber RS, Langer CJ, Ang KK, Gay G, Stewart A, Robinson RA. 2006. Laryngeal cancer in the United States: changes in demographics, patterns of care, and survival. Laryngoscope 116:1–13. doi:10.1097/01.mlg.0000236095.97947.26. - DOI - PubMed

MeSH terms

LinkOut - more resources