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Viral non-coding RNA inhibits HNF4α expression in HCV associated hepatocellular carcinoma.

Wang Z, Ceniccola K, Florea L, Wang BD, Lee NH, Kumar A - Infect. Agents Cancer (2015)

Bottom Line: Results show inhibition of HNF4α expression by targeting of HNF4α 3'-UTR by HCV-derived small non-coding RNA, vmr11.Vmr11 enhances the invasive properties of HCV-infected cells.These results suggest a direct viral role in the development of hepatocellular carcinoma.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, The George Washington University, Washington, DC USA.

ABSTRACT

Background: Hepatitis C virus (HCV) infection is an established cause of chronic hepatitis, cirrhosis and hepatocellular carcinoma (HCC); however, it is unclear if the virus plays a direct role in the development of HCC. Hepatocyte nuclear factor 4α (HNF4α) is critical determinant of epithelial architecture and hepatic development; depletion of HNF4α is correlated with oncogenic transformation. We explored the viral role in the inhibition of HNF4α expression, and consequent induction of tumor-promoting genes in HCV infection-associated HCC.

Methods: Western blot analysis was used to monitor the changes in expression levels of oncogenic proteins in liver tissues from HCV-infected humanized mice. The mechanism of HNF4α depletion was studied in HCV-infected human hepatocyte cultures in vitro. Targeting of HNF4α expression by viral non-coding RNA was examined by inhibition of Luciferase HNF4α 3'-UTR reporter. Modulation of invasive properties of HCV-infected cells was examined by Matrigel cell migration assay.

Results: Results show inhibition of HNF4α expression by targeting of HNF4α 3'-UTR by HCV-derived small non-coding RNA, vmr11. Vmr11 enhances the invasive properties of HCV-infected cells. Loss of HNF4α in HCV-infected liver tumors of humanized mice correlates with the induction of epithelial to mesenchymal transition (EMT) genes.

Conclusions: We show depletion of HNF4α in liver tumors of HCV-infected humanized mice by HCV derived small non-coding RNA (vmr11) and resultant induction of EMT genes, which are critical determinants of tumor progression. These results suggest a direct viral role in the development of hepatocellular carcinoma.

No MeSH data available.


Related in: MedlinePlus

Luc-HNF4a 3’-UTR assay: Indicated concentrations of 22 nucleotide vmr11 “mimic” oligonucleotides or 2’-Fluoro modified vmr11 mimic oligonucleotides were co-transfected with Luciferase HNF4α 3’UTR reporter plasmid [13] and luciferase activity was quantitated three days post transfection as described
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Fig6: Luc-HNF4a 3’-UTR assay: Indicated concentrations of 22 nucleotide vmr11 “mimic” oligonucleotides or 2’-Fluoro modified vmr11 mimic oligonucleotides were co-transfected with Luciferase HNF4α 3’UTR reporter plasmid [13] and luciferase activity was quantitated three days post transfection as described

Mentions: We further investigated whether the predicted vmr11 target site within HNF4α mRNA 3’-UTR is recognized by vmr11 RNA to block HNF4α expression. To do this, we determined the extent of inhibition of Luciferase-HNF4α-3’UTR reporter gene expression with increasing amounts of vmr11 oligonucleotides. To ascertain that vmr11-directed inhibition of HNF4α is not due to the instability of vmr11 oligonucleotides introduced into the cells, we used equimolar amounts of standard vmr11 oligonucleotides and 2’-Fluoro-stabilized vmr11 RNA. We co-transfected Luciferase HNF4α 3’-UTR reporter plasmid [13] with increasing amounts of either the wild-type vmr11 (“mimic”) oligunucleotides, or 2’-Fluoro-modified vmr11 oligonucleotides (TriLink BioTechnolgies). Luciferase reporter assays suggest that introduction of either normal vmr11 oligonucleotides, or 2’-Fluoro-stabilized vmr11 RNA was equally efficient in blocking HNF4α expression; about 75 % down regulation of HNF4α protein was observed, compared to cells transfected with irrelevant, scrambled oligonucleotides (Fig. 6).Fig. 6


Viral non-coding RNA inhibits HNF4α expression in HCV associated hepatocellular carcinoma.

Wang Z, Ceniccola K, Florea L, Wang BD, Lee NH, Kumar A - Infect. Agents Cancer (2015)

Luc-HNF4a 3’-UTR assay: Indicated concentrations of 22 nucleotide vmr11 “mimic” oligonucleotides or 2’-Fluoro modified vmr11 mimic oligonucleotides were co-transfected with Luciferase HNF4α 3’UTR reporter plasmid [13] and luciferase activity was quantitated three days post transfection as described
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4495692&req=5

Fig6: Luc-HNF4a 3’-UTR assay: Indicated concentrations of 22 nucleotide vmr11 “mimic” oligonucleotides or 2’-Fluoro modified vmr11 mimic oligonucleotides were co-transfected with Luciferase HNF4α 3’UTR reporter plasmid [13] and luciferase activity was quantitated three days post transfection as described
Mentions: We further investigated whether the predicted vmr11 target site within HNF4α mRNA 3’-UTR is recognized by vmr11 RNA to block HNF4α expression. To do this, we determined the extent of inhibition of Luciferase-HNF4α-3’UTR reporter gene expression with increasing amounts of vmr11 oligonucleotides. To ascertain that vmr11-directed inhibition of HNF4α is not due to the instability of vmr11 oligonucleotides introduced into the cells, we used equimolar amounts of standard vmr11 oligonucleotides and 2’-Fluoro-stabilized vmr11 RNA. We co-transfected Luciferase HNF4α 3’-UTR reporter plasmid [13] with increasing amounts of either the wild-type vmr11 (“mimic”) oligunucleotides, or 2’-Fluoro-modified vmr11 oligonucleotides (TriLink BioTechnolgies). Luciferase reporter assays suggest that introduction of either normal vmr11 oligonucleotides, or 2’-Fluoro-stabilized vmr11 RNA was equally efficient in blocking HNF4α expression; about 75 % down regulation of HNF4α protein was observed, compared to cells transfected with irrelevant, scrambled oligonucleotides (Fig. 6).Fig. 6

Bottom Line: Results show inhibition of HNF4α expression by targeting of HNF4α 3'-UTR by HCV-derived small non-coding RNA, vmr11.Vmr11 enhances the invasive properties of HCV-infected cells.These results suggest a direct viral role in the development of hepatocellular carcinoma.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, The George Washington University, Washington, DC USA.

ABSTRACT

Background: Hepatitis C virus (HCV) infection is an established cause of chronic hepatitis, cirrhosis and hepatocellular carcinoma (HCC); however, it is unclear if the virus plays a direct role in the development of HCC. Hepatocyte nuclear factor 4α (HNF4α) is critical determinant of epithelial architecture and hepatic development; depletion of HNF4α is correlated with oncogenic transformation. We explored the viral role in the inhibition of HNF4α expression, and consequent induction of tumor-promoting genes in HCV infection-associated HCC.

Methods: Western blot analysis was used to monitor the changes in expression levels of oncogenic proteins in liver tissues from HCV-infected humanized mice. The mechanism of HNF4α depletion was studied in HCV-infected human hepatocyte cultures in vitro. Targeting of HNF4α expression by viral non-coding RNA was examined by inhibition of Luciferase HNF4α 3'-UTR reporter. Modulation of invasive properties of HCV-infected cells was examined by Matrigel cell migration assay.

Results: Results show inhibition of HNF4α expression by targeting of HNF4α 3'-UTR by HCV-derived small non-coding RNA, vmr11. Vmr11 enhances the invasive properties of HCV-infected cells. Loss of HNF4α in HCV-infected liver tumors of humanized mice correlates with the induction of epithelial to mesenchymal transition (EMT) genes.

Conclusions: We show depletion of HNF4α in liver tumors of HCV-infected humanized mice by HCV derived small non-coding RNA (vmr11) and resultant induction of EMT genes, which are critical determinants of tumor progression. These results suggest a direct viral role in the development of hepatocellular carcinoma.

No MeSH data available.


Related in: MedlinePlus