Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

12 gep oncogene deregulation of p53-responsive microRNAs promotes epithelial–mesenchymal transition of hepatocellular carcinoma

Subjects

Abstract

Hepatocellular carcinoma (HCC) has a poor prognosis owing to aggressive phenotype. Gα12 gep oncogene product couples to G-protein-coupled receptors, whose ligand levels are frequently increased in tumor microenvironments. Here, we report Gα12 overexpression in human HCC and the resultant induction of zinc-finger E-box-binding homeobox 1 (ZEB1) as mediated by microRNA deregulation. Gα12 expression was higher in HCC than surrounding non-tumorous tissue. Transfection of Huh7 cell with an activated mutant of Gα12 (Gα12QL) deregulated microRNA (miRNA or miR)-200b/a/429, -194-2/192 and -194-1/215 clusters in the miRNome. cDNA microarray analyses disclosed the targets affected by Gα12 gene knockout. An integrative network of miRNAs and mRNA changes enabled us to predict ZEB1 as a key molecule governed by Gα12. Decreases of miR-200a/b, -192 and -215 by Gα12 caused ZEB1 induction. The ability of Gα12 to decrease p53 levels, as a result of activating protein-1 (AP-1)/c-Jun-mediated mouse double minute 2 homolog induction, contributed to transcriptional deregulation of the miRNAs. Gα12QL induced ZEB1 and other epithelial–mesenchymal transition markers with fibroblastoid phenotype change. Consistently, transfection with miR-200b, -192 or -215 mimic prevented the ability of Gα12QL to increase tumor cell migration/invasion. In xenograft studies, sustained knockdown of Gα12 decreased the overall growth rate and average volume of tumors derived from SK-Hep1 cell (mesenchymal-typed). In HCC patients, miR-192, -215 and/or -200a were deregulated with microvascular invasion or growth advantage. In the HCC samples with higher Gα12 level, a correlation existed in the comparison of relative changes of Gα12 and ZEB1. In conclusion, Gα12 overexpressed in HCC causes ZEB1 induction by deregulating p53-responsive miRNAs, which may facilitate epithelial–mesenchymal transition and growth of liver tumor. These findings highlight the significance of Gα12 upregulation in liver tumor progression, implicating Gα12 as an attractive therapeutic target.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8

Accession codes

Accessions

Gene Expression Omnibus

References

  1. Mazzocca A, Dituri F, Lupo L, Quaranta M, Antonaci S, Giannelli G . Tumor-secreted lysophostatidic acid accelerates hepatocellular carcinoma progression by promoting differentiation of peritumoral fibroblasts in myofibroblasts. Hepatology 2011; 54: 920–930.

    Article  CAS  PubMed  Google Scholar 

  2. Riobo NA, Manning DR . Receptors coupled to heterotrimeric G proteins of the G12 family. Trends Pharmacol Sci 2005; 26: 146–154.

    Article  CAS  PubMed  Google Scholar 

  3. O'Hayre M, Vazquez-Prado J, Kufareva I, Stawiski EW, Handel TM, Seshagiri S et al. The emerging mutational landscape of G proteins and G-protein-coupled receptors in cancer. Nat Rev Cancer 2013; 13: 412–424.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Kelly P, Casey PJ, Meigs TE . Biologic functions of the G12 subfamily of heterotrimeric g proteins: growth, migration, and metastasis. Biochemistry 2007; 46: 6677–6687.

    Article  CAS  PubMed  Google Scholar 

  5. Kelly P, Moeller BJ, Juneja J, Booden MA, Der CJ, Daaka Y et al. The G12 family of heterotrimeric G proteins promotes breast cancer invasion and metastasis. Proc Natl Acad Sci USA 2006; 103: 8173–8178.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Kelly P, Stemmle LN, Madden JF, Fields TA, Daaka Y, Casey PJ . A role for the G12 family of heterotrimeric G proteins in prostate cancer invasion. J Biol Chem 2006; 281: 26483–26490.

    Article  CAS  PubMed  Google Scholar 

  7. Grzelinski M, Pinkenburg O, Buch T, Gold M, Stohr S, Kalwa H et al. Critical role of G(alpha)12 and G(alpha)13 for human small cell lung cancer cell proliferation in vitro and tumor growth in vivo. Clin Cancer Res 2010; 16: 1402–1415.

    Article  CAS  PubMed  Google Scholar 

  8. Tung-Ping Poon R, Fan ST, Wong J . Risk factors, prevention, and management of postoperative recurrence after resection of hepatocellular carcinoma. Ann Surg 2000; 232: 10–24.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Peinado H, Olmeda D, Cano A . Snail, Zeb and bHLH factors in tumour progression: an alliance against the epithelial phenotype? Nat Rev Cancer 2007; 7: 415–428.

    Article  CAS  PubMed  Google Scholar 

  10. Bartel DP . MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 2004; 116: 281–297.

    Article  CAS  PubMed  Google Scholar 

  11. Calin GA, Sevignani C, Dumitru CD, Hyslop T, Noch E, Yendamuri S et al. Human microRNA genes are frequently located at fragile sites and genomic regions involved in cancers. Proc Natl Acad Sci USA 2004; 101: 2999–3004.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Kim T, Veronese A, Pichiorri F, Lee TJ, Jeon YJ, Volinia S et al. P53 regulates epithelial–mesenchymal transition through microRNAs targeting ZEB1 and ZEB2. J Exp Med 2011; 208: 875–883.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Xu N, Bradley L, Ambdukar I, Gutkind JS . A mutant alpha subunit of G12 potentiates the eicosanoid pathway and is highly oncogenic in NIH 3T3 cells. Proc Natl Acad Sci USA 1993; 90: 6741–6745.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Yang YM, Lee S, Nam CW, Ha JH, Jayaraman M, Dhanasekaran DN et al. G(alpha)12/13 inhibition enhances the anticancer effect of bortezomib through PSMB5 downregulation. Carcinogenesis 2010; 31: 1230–1237.

    Article  CAS  PubMed  Google Scholar 

  15. Gu JL, Muller S, Mancino V, Offermanns S, Simon MI . Interaction of G alpha(12) with G alpha(13) and G alpha(q) signaling pathways. Proc Natl Acad Sci USA 2002; 99: 9352–9357.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Braun CJ, Zhang X, Savelyeva I, Wolff S, Moll UM, Schepeler T et al. P53-responsive micrornas 192 and 215 are capable of inducing cell cycle arrest. Cancer Res 2008; 68: 10094–10104.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Pichiorri F, Suh SS, Rocci A, De Luca L, Taccioli C, Santhanam R et al. Downregulation of p53-inducible microRNAs 192, 194, and 215 impairs the p53/MDM2 autoregulatory loop in multiple myeloma development. Cancer Cell 2010; 18: 367–381.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Bressac B, Galvin KM, Liang TJ, Isselbacher KJ, Wands JR, Ozturk M . Abnormal structure and expression of P53 gene in human hepatocellular-carcinoma. Proc Natl Acad Sci USA 1990; 87: 1973–1977.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Hsieh JL, Wu CL, Lee CH, Shiau AL . Hepatitis B virus X protein sensitizes hepatocellular carcinoma cells to cytolysis induced by E1B-deleted adenovirus through the disruption of p53 function. Clin Cancer Res 2003; 9: 338–345.

    CAS  PubMed  Google Scholar 

  20. Dharel N, Kato N, Muroyama R, Taniguchi H, Otsuka M, Wang Y et al. Potential contribution of tumor suppressor p53 in the host defense against hepatitis C virus. Hepatology 2008; 47: 1136–1149.

    Article  CAS  PubMed  Google Scholar 

  21. Di Como CJ, Prives C . Human tumor-derived p53 proteins exhibit binding site selectivity and temperature sensitivity for transactivation in a yeast-based assay. Oncogene 1998; 16: 2527–2539.

    Article  CAS  PubMed  Google Scholar 

  22. Wang Z, Li B . Mdm2 links genotoxic stress and metabolism to p53. Protein Cell 2010; 1: 1063–1072.

    Article  CAS  PubMed  Google Scholar 

  23. Kim ES, Kim JS, Kim SG, Hwang S, Lee CH, Moon A . Sphingosine 1-phosphate regulates matrix metalloproteinase-9 expression and breast cell invasion through S1P3-Galphaq coupling. J Cell Sci 2011; 124: 2220–2230.

    Article  CAS  PubMed  Google Scholar 

  24. Sumie S, Kuromatsu R, Okuda K, Ando E, Takata A, Fukushima N et al. Microvascular invasion in patients with hepatocellular carcinoma and its predictable clinicopathological factors. Ann Surg Oncol 2008; 15: 1375–1382.

    Article  PubMed  Google Scholar 

  25. Gregory PA, Bert AG, Paterson EL, Barry SC, Tsykin A, Farshid G et al. The miR-200 family and miR-205 regulate epithelial to mesenchymal transition by targeting ZEB1 and SIP1. Nat Cell Biol 2008; 10: 593–601.

    Article  CAS  PubMed  Google Scholar 

  26. Liu SC, Jen YM, Jiang SS, Chang JL, Hsiung CA, Wang CH et al. G(alpha)12-mediated pathway promotes invasiveness of nasopharyngeal carcinoma by modulating actin cytoskeleton reorganization. Cancer Res 2009; 69: 6122–6130.

    Article  CAS  PubMed  Google Scholar 

  27. Kim MS, Lee SM, Kim WD, Ki SH, Moon A, Lee CH et al. G alpha 12/13 basally regulates p53 through Mdm4 expression. Mol Cancer Res 2007; 5: 473–484.

    Article  CAS  PubMed  Google Scholar 

  28. Fornari F, Gramantieri L, Giovannini C, Veronese A, Ferracin M, Sabbioni S et al. MiR-122/cyclin G1 interaction modulates p53 activity and affects doxorubicin sensitivity of human hepatocarcinoma cells. Cancer Res 2009; 69: 5761–5767.

    Article  CAS  PubMed  Google Scholar 

  29. Lee SJ, Yang JW, Cho IJ, Kim WD, Cho MK, Lee CH et al. The gep oncogenes, Galpha(12) and Galpha(13), upregulate the transforming growth factor-beta1 gene. Oncogene 2009; 28: 1230–1240.

    Article  CAS  PubMed  Google Scholar 

  30. Kang KW, Choi SY, Cho MK, Lee CH, Kim SG . Thrombin induces nitric-oxide synthase via Galpha12/13-coupled protein kinase C-dependent I-kappaBalpha phosphorylation and JNK-mediated I-kappaBalpha degradation. J Biol Chem 2003; 278: 17368–17378.

    Article  CAS  PubMed  Google Scholar 

  31. Lin F, Chen S, Sepich DS, Panizzi JR, Clendenon SG, Marrs JA et al. Galpha12/13 regulate epiboly by inhibiting E-cadherin activity and modulating the actin cytoskeleton. J Cell Biol 2009; 184: 909–921.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Xu J, Lamouille S, Derynck R . TGF-beta-induced epithelial to mesenchymal transition. Cell Res 2009; 19: 156–172.

    Article  CAS  PubMed  Google Scholar 

  33. Gregory PA, Bracken CP, Smith E, Bert AG, Wright JA, Roslan S et al. An autocrine TGF-beta/ZEB/miR-200 signaling network regulates establishment and maintenance of epithelial-mesenchymal transition. Mol Biol Cell 2011; 22: 1686–1698.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Wang B, Herman-Edelstein M, Koh P, Burns W, Jandeleit-Dahm K, Watson A et al. E-cadherin expression is regulated by miR-192/215 by a mechanism that is independent of the profibrotic effects of transforming growth factor-beta. Diabetes 2010; 59: 1794–1802.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Burk U, Schubert J, Wellner U, Schmalhofer O, Vincan E, Spaderna S et al. A reciprocal repression between ZEB1 and members of the miR-200 family promotes EMT and invasion in cancer cells. EMBO Rep 2008; 9: 582–589.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Liu YN, Yin JJ, Abou-Kheir W, Hynes PG, Casey OM, Fang L et al. MiR-1 and miR-200 inhibit EMT via Slug-dependent and tumorigenesis via Slug-independent mechanisms. Oncogene 2013; 32: 296–306.

    Article  CAS  PubMed  Google Scholar 

  37. Feng B, Wang R, Chen LB . Review of miR-200b and cancer chemosensitivity. Biomed Pharmacother 2012; 66: 397–402.

    Article  CAS  PubMed  Google Scholar 

  38. Sanchez-Tillo E, Fanlo L, Siles L, Montes-Moreno S, Moros A, Chiva-Blanch G et al. The EMT activator ZEB1 promotes tumor growth and determines differential response to chemotherapy in mantle cell lymphoma. Cell Death Differ 2014; 21: 247–257.

    Article  CAS  PubMed  Google Scholar 

  39. Zhou YM, Cao L, Li B, Zhang RX, Sui CJ, Yin ZF et al. Clinicopathological significance of ZEB1 protein in patients with hepatocellular carcinoma. Ann Surg Oncol 2012; 19: 1700–1706.

    Article  PubMed  Google Scholar 

  40. Chen X, Lingala S, Khoobyari S, Nolta J, Zern MA, Wu J . Epithelial mesenchymal transition and hedgehog signaling activation are associated with chemoresistance and invasion of hepatoma subpopulations. J Hepatol 2011; 55: 838–845.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Ki SH, Choi MJ, Lee CH, Kim SG . Galpha12 specifically regulates COX-2 induction by sphingosine 1-phosphate. Role for JNK-dependent ubiquitination and degradation of IkappaBalpha. J Biol Chem 2007; 282: 1938–1947.

    Article  CAS  PubMed  Google Scholar 

  42. Yang YM, Seo SY, Kim TH, Kim SG . Decrease of microRNA-122 causes hepatic insulin resistance by inducing protein tyrosine phosphatase 1B, which is reversed by licorice flavonoid. Hepatology 2012; 56: 2209–2220.

    Article  CAS  PubMed  Google Scholar 

  43. Shin KJ, Wall EA, Zavzavadjian JR, Santat LA, Liu J, Hwang JI et al. A single lentiviral vector platform for microRNA-based conditional RNA interference and coordinated transgene expression. Proc Natl Acad Sci USA 2006; 103: 13759–13764.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Georges SA, Biery MC, Kim SY, Schelter JM, Guo J, Chang AN et al. Coordinated regulation of cell cycle transcripts by p53-inducible microRNAs, miR-192 and miR-215. Cancer Res 2008; 68: 10105–10112.

    Article  CAS  PubMed  Google Scholar 

  45. Esau C, Davis S, Murray SF, Yu XX, Pandey SK, Pear M et al. MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting. Cell Metab 2006; 3: 87–98.

    Article  CAS  PubMed  Google Scholar 

  46. Lee JM, Lee WH, Kay HY, Kim ES, Moon A, Kim SG . Hemin, an iron-binding porphyrin, inhibits HIF-1alpha induction through its binding with heat shock protein 90. Int J Cancer 2012; 130: 716–727.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIP) (No. 2007-0056817) and in part by the World Class University project (R322012000100980) and National Cancer Center (No. 1110050). We thank Prof Dr Soon-Sun Hong and Hee-Seung Lee for their indispensable support.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to S G Kim.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Additional information

Supplementary Information accompanies this paper on the Oncogene website

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Yang, Y., Lee, W., Lee, C. et al.12 gep oncogene deregulation of p53-responsive microRNAs promotes epithelial–mesenchymal transition of hepatocellular carcinoma. Oncogene 34, 2910–2921 (2015). https://doi.org/10.1038/onc.2014.218

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/onc.2014.218

Search

Quick links