microRNA-497 inhibits cell proliferation and induces apoptosis by targeting YAP1 in human hepatocellular carcinoma - PubMed (original) (raw)

microRNA-497 inhibits cell proliferation and induces apoptosis by targeting YAP1 in human hepatocellular carcinoma

Lei Zhang et al. FEBS Open Bio. 2016.

Retraction in

Abstract

microRNAs (miRNAs) function as oncogenes or tumor suppressors in human cancers by targeting mRNAs for degradation and/or translational repression. miR-497 has been proposed as a tumor suppressive miRNA and its deregulation is observed in human cancers. However, the prognostic value of miR-497 and its underlying molecular pathways involved in the initiation and development of hepatocellular carcinoma (HCC) are poorly investigated. In the present study, we found that the mean level of miR-497 in HCC tissues was lower than that in adjacent nontumor tissues. Clinical data indicated that low expression of miR-497 was prominently associated with adverse prognostic features of HCC including high serum alpha-fetoprotein (AFP) level, large tumor size, high Edmondson-Steiner grading and advanced tumor-node-metastasis (TNM) stage. Furthermore, miR-497 was an independent prognostic factor for indicating both 5-year overall survival and disease-free survival of HCC patients. Gain- and loss-of-function studies showed that miR-497 reduced cell proliferation and induced apoptosis in HCC cells. Yes-associated protein 1 (YAP1) was identified as a direct target of miR-497 in HCC. An inverse correlation between YAP1 and miR-497 expression was observed in HCC tissues. Notably, YAP1 knockdown abrogated the effects of miR-497 deletion on HCC cells with decreased cell proliferation and increased apoptosis. In conclusion, we report that miR-497 is a potent prognostic indicator and may suppress tumor growth of HCC by targeting YAP1.

Keywords: Yes‐associated protein 1; apoptosis; hepatocellular carcinoma; microRNA‐497; proliferation.

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Figures

Figure 1

Figure 1

The expression and prognostic significance of miR‐497 in hepatocellular carcinoma (

HCC

). (A) Comparing differences in the expression levels of miR‐497 between

HCC

(T) and matched adjacent nontumor tissues (

NT

). *P < 0.05 by _t_‐test. (B, C) According to the level of miR‐497 expression, Kaplan–Meier 5‐year overall and disease‐free survival curves of

HCC

patients showed that low expression of miR‐497 was correlated with poor prognosis. The median expression value obtained for miR‐497 of the 86

HCC

samples detected by qRT‐

PCR

was chosen as the cutoff value.

Figure 2

Figure 2

miR‐497 reduces cell proliferation and induces apoptosis in hepatocellular carcinoma (

HCC

) cells. (A) HepG2 cells that were transfected with miR‐control (control) and miR‐497, respectively, were subjected to qRT‐

PCR

for miR‐497 expression. n = three independent experiments, *P < 0.05 by _t_‐test. (B) Cell proliferation as measured by BrdU incorporation assays was inhibited by up‐regulation of miR‐497 in HepG2 cells as compared with control cells. n = three repeats with similar results, *P < 0.05 by _t_‐test. (C) miR‐497‐overexpressing HepG2 cells conferred a large subgroup of apoptotic cells as compared with control cells. n = three repeats with similar results, *P < 0.05 by _t_‐test. (D) Huh7 cells that were transfected with negative control (

NC

) and miR‐497 inhibitor (anti‐miR‐497), respectively, were subjected to qRT‐

PCR

for miR‐497 expression. n = three independent experiments, *P < 0.05 by _t_‐test. (E) and (F) Down‐regulation of miR‐497 promoted cell proliferation and inhibited apoptosis in Huh7 cells. n = three repeats with similar results, *P < 0.05 by _t_‐test.

Figure 3

Figure 3

YAP

1 is identified as a functional target of miR‐497 in hepatocellular carcinoma (

HCC

). (A) qRT‐

PCR

and (B) western blot analysis of

YAP

1 expression in HepG2 cells with miR‐497 or control vectors transfection. n = three independent experiments; *P < 0.05 by _t_‐test. (C) miR‐497 and its putative binding sequence in the 3′‐

UTR

of

YAP

1. The mutant miR‐497 binding site was generated in the complementary site for the seed region of miR‐497 (wt, wild‐type; mt, mutant type). (D) miR‐497 significantly suppressed the luciferase activity that carried wt but not mt 3′‐

UTR

of

YAP

1. Anti‐miR‐497 led to a noticeable increase in luciferase activity of wt 3′‐

UTR

of

YAP

1. n = three repeats with similar results; *P < 0.05 by _t_‐test.

Figure 4

Figure 4

miR‐497 is inversely correlated with

YAP

1 in hepatocellular carcinoma (

HCC

). (A) Comparing differences in the expression levels of

YAP

1 between

HCC

(T) and matched adjacent nontumor tissues (

NT

). *P < 0.05 by _t_‐test. (B) Representative immunostaining showed negative expression of

YAP

1 in miR‐497 high‐expressing

HCC

tissue and positive expression of

YAP

1 in miR‐497 low‐expressing tumor. A significant inverse correlation between miR‐497 and

YAP

1 expression was observed in

HCC

tissues. Scale bar: 50 μm; *P < 0.05 by chi‐square test.

Figure 5

Figure 5

YAP

1 knockdown reduces cell proliferation and induces apoptosis in miR‐497 down‐regulating Huh7 cells. (A) Huh7 cells with anti‐miR‐497 vector transfection (Huh7‐anti‐miR‐497) that was transfected with scrambled si

RNA

and

YAP

1 si

RNA

, respectively, were subjected to western blot for

YAP

1 expression. n = three independent experiments, *P < 0.05 by _t_‐test. (B) Cell proliferation as measured by BrdU incorporation assays was inhibited by

YAP

1 knockdown in Huh7‐anti‐miR‐497 cells as compared with control cells. n = three repeats with similar results, *P < 0.05 by _t_‐test. (C)

YAP

1 down‐regulating Huh7‐anti‐miR‐497 cells conferred a large subgroup of apoptotic cells as compared with control cells. n = three repeats with similar results, *P < 0.05 by _t_‐test.

References

    1. Dhanasekaran R, Limaye A and Cabrera R (2012) Hepatocellular carcinoma: current trends in worldwide epidemiology, risk factors, diagnosis, and therapeutics. Hepat Med 4, 19–37. - PMC - PubMed
    1. Yang LY, Fang F, Ou DP, Wu W, Zeng ZJ and Wu F (2009) Solitary large hepatocellular carcinoma: a specific subtype of hepatocellular carcinoma with good outcome after hepatic resection. Ann Surg 249, 118–123. - PubMed
    1. Aravalli RN, Steer CJ and Cressman EN (2008) Molecular mechanisms of hepatocellular carcinoma. Hepatology 48, 2047–2063. - PubMed
    1. Llovet JM and Bruix J (2008) Molecular targeted therapies in hepatocellular carcinoma. Hepatology 48, 1312–1327. - PMC - PubMed
    1. He L and Hannon GJ (2004) MicroRNAs: small RNAs with a big role in gene regulation. Nat Rev Genet 5, 522–531. - PubMed

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