Dataset Information

Hepatitis B virus X protein promotes human hepatoma cell growth via upregulation of transcription factor AP2? and sphingosine kinase 1.

ABSTRACT:

Aim

Sphingosine kinase 1 (SPHK1) is involved in various cellular functions, including cell growth, migration, apoptosis, cytoskeleton architecture and calcium homoeostasis, etc. As an oncogenic kinase, SPHK1 is associated with the development and progression of cancers. The aim of this study was to investigate whether SPHK1 was involved in hepatocarcinogenesis induced by the hepatitis B virus X protein (HBx).

Methods

The expression of SPHK1 in hepatocellular carcinoma (HCC) tissue and hepatoma cells were measured using qRT-PCR and Western blot analysis. HBx expression levels in hepatoma cells were modulated by transiently transfected with HBx or psi-HBx plasmids. The SPHK1 promoter activity was measured using luciferase reporter gene assay, and the interaction of the transcription factor AP2? with the SPHK1 promoter was studied with chromatin immunoprecipitation assay. The growth of hepatoma cells was evaluated in vitro using MTT and colony formation assays, and in a tumor xenograft model.

Results

A positive correlation was found between the mRNA levels of SPHK1 and HBx in 38 clinical HCC samples (r=+0.727, P<0.01). Moreover, the expression of SPHK1 was markedly increased in the liver cancer tissue of HBx-transgenic mice. Overexpressing HBx in normal liver cells LO2 and hepatoma cells HepG2 dose-dependently increased the expression of SPHK1, whereas silencing HBx in HBx-expressing hepatoma cells HepG2-X and HepG2.2.15 suppressed SPHK1 expression. Furthermore, overexpressing HBx in HepG2 cells dose-dependently increased the SPHK1 promoter activity, whereas silencing HBx in HepG2-X cells suppressed this activity. In HepG2-X cells, AP2? was found to directly interact with the SPHK1 promoter, and silencing AP2? suppressed the SPHK1 promoter activity and SPHK1 expression. Silencing HBx in HepG2-X cells abolished the HBx-enhanced proliferation and colony formation in vitro, and tumor growth in vivo.

Conclusion

HBx upregulates SPHK1 through the transcription factor AP2?, which promotes the growth of human hepatoma cells.

SUBMITTER: Lu ZP

PROVIDER: S-EPMC4814204 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Hepatitis B virus X protein promotes human hepatoma cell growth via upregulation of transcription factor AP2α and sphingosine kinase 1.

Lu Zhan-ping ZP Xiao Ze-lin ZL Yang Zhe Z Li Jiong J Feng Guo-xing GX Chen Fu-quan FQ Li Ying-Hui YH Feng Jin-yan JY Gao Yu-en YE Ye Li-hong LH Zhang Xiao-dong XD

Acta pharmacologica Sinica 20150615 10

<h4>Aim</h4>Sphingosine kinase 1 (SPHK1) is involved in various cellular functions, including cell growth, migration, apoptosis, cytoskeleton architecture and calcium homoeostasis, etc. As an oncogenic kinase, SPHK1 is associated with the development and progression of cancers. The aim of this study was to investigate whether SPHK1 was involved in hepatocarcinogenesis induced by the hepatitis B virus X protein (HBx).<h4>Methods</h4>The expression of SPHK1 in hepatocellular carcinoma (HCC) tissue ...[more]

PMID: 26073327

Similar Datasets

Project description:Sphingosine-1-phosphate (SPP) is a bioactive lipid that has recently been identified as the ligand for the EDG family of G protein-coupled cell surface receptors. However, the mitogenic and survival effects of exogenous SPP may not correlate with binding to cell-surface receptors (Van Brocklyn, J.R., M.J. Lee, R. Menzeleev, A. Olivera, L. Edsall, O. Cuvillier, D.M. Thomas, P.J.P. Coopman, S. Thangada, T. Hla, and S. Spiegel. 1998. J. Cell Biol. 142:229-240). The recent cloning of sphingosine kinase, a unique lipid kinase responsible for the formation of SPP, has provided a new tool to investigate the role of intracellular SPP. Expression of sphingosine kinase markedly increased SPP levels in NIH 3T3 fibroblasts and HEK293 cells, but no detectable secretion of SPP into the medium was observed. The increased sphingosine kinase activity in NIH 3T3 fibroblasts was sufficient to promote growth in low- serum media, expedite the G(1)/S transition, and increase DNA synthesis and the proportion of cells in the S phase of the cell cycle with a concomitant increase in cell numbers. Transient or stable overexpression of sphingosine kinase in NIH 3T3 fibroblasts or HEK293 cells protected against apoptosis induced by serum deprivation or ceramide elevation. N,N-Dimethylsphingosine, a competitive inhibitor of sphingosine kinase, blocked the effects of sphingosine kinase overexpression on cell proliferation and suppression of apoptosis. In contrast, pertussis toxin did not abrogate these biological responses. In Jurkat T cells, overexpression of sphingosine kinase also suppressed serum deprivation- and ceramide-induced apoptosis and, to a lesser extent, Fas-induced apoptosis, which correlated with inhibition of DEVDase activity, as well as inhibition of the executionary caspase-3. Taken together with ample evidence showing that growth and survival factors activate sphingosine kinase, our results indicate that SPP functions as a second messenger important for growth and survival of cells. Hence, SPP belongs to a novel class of lipid mediators that can function inside and outside cells.

Project description:Both alcohol and hepatitis C virus (HCV) infection could induce cellular autophagy in liver cells, which is considered to be essential for productive HCV replication. However, whether alcohol-induced autophagy is involved in the pathogenesis of HCV infection is still poorly understood. Alcohol treatment could induce autophagy in Huh7 cells (a hepatoma cell line that supports HCV JFH-1 replication), evidenced by the increase of LC3B-II levels, the conversion of LC3B-I to LC3B-II, and the formation of GFP-LC3 puncta as well as the decrease of p62 level in alcohol-treated cells compared with control cells. Alcohol treatment also significantly increased PIASy (a member of the PIAS family) expression, which can act as a SUMO (small ubiquitin-like modifier protein) E3 ligase to regulate a broader range of cellular processes including autophagy. Overexpression or the silencing expression of PIASy in alcohol-treated Huh7 cells could increase or decrease autophagic activation caused by alcohol treatment, respectively, and thus affect HCV replication correspondingly. In the absence of alcohol, overexpression or silencing expression of PIASy increase or decrease the level of cellular autophagy, judged by the changes of LC3B-II and p62 levels in the presence or absence of chloroquine (CQ), a lysosome inhibitor. More importantly, in the presence of 3-methyladenine (3-MA), an inhibitor in the early stage of autophagy, the effects of overexpression or silencing expression of PIASy on HCV replication were largely blocked. Furthermore, PIASy could selectively drive the accumulation of SUMO1-conjugated proteins, along with upregulation of the expression of several important autophagy factors, including ATG7 and ATG5-ATG12. In conclusion, alcohol promotes HCV replication through activation of autophagy in Huh7 cells, which partly attributes to its induction of PIASy expression. PIASy-enhanced accumulation of SUMO1-conjugated proteins may contribute to its inducing effect of autophagy. Our findings provide a novel mechanism for the action of alcohol-promoting HCV replication in the context of cellular autophagy.