Hepatitis B virus X protein promotes vimentin expression via LIM and SH3 domain protein 1 to facilitate epithelial-mesenchymal transition and hepatocarcinogenesis

Abstract

Background Hepatitis B virus (HBV) X protein (HBX) has been reported to be responsible for the epithelial-mesenchymal transition (EMT) in HBV-related hepatocellular carcinoma (HCC). Vimentin is an EMT-related molecular marker. However, the importance of vimentin in the pathogenesis of HCC mediated by HBX has not been well determined. Methods The expression of vimentin induced by HBX, and the role of LIM and SH3 domain protein 1 (LASP1) in HBX-induced vimentin expression in hepatoma cells were examined by western blot and immunohistochemistry analysis. Both the signal pathways involved in the expression of vimentin, the interaction of HBX with vimentin and LASP1, and the stability of vimentin mediated by LASP1 in HBX-positive cells were assessed by western blot, Co-immunoprecipitation, and GST-pull down assay. The role of vimentin in EMT, proliferation, and migration of HCC cells mediated by HBX and LASP1 were explored with western blot, CCK-8 assay, plate clone formation assay, transwell assay, and wound healing assay. Results Vimentin expression was increased in both HBX-positive hepatoma cells and HBV-related HCC tissues, and the expression of vimentin was correlated with HBX in HBV-related HCC tissues. Functionally, vimentin was contributed to the EMT, proliferation, and migration of hepatoma cells mediated by HBX. The mechanistic analysis suggested that HBX was able to enhance the expression of vimentin through LASP1. On the one hand, PI3-K, ERK, and STAT3 signal pathways were involved in the upregulation of vimentin mediated by LASP1 in HBX-positive hepatoma cells. On the other hand, HBX could directly interact with vimentin and LASP1, and dependent on LASP1, HBX was capable of promoting the stability of vimentin via protecting it from ubiquitination mediated protein degradation. Besides these, vimentin was involved in the growth and migration of hepatoma cells mediated by LASP1 in HBX-positive hepatoma cells. Conclusion Taken together, these findings demonstrate that, dependent on LASP1, vimentin is crucial for HBX-mediated EMT and hepatocarcinogenesis, and may serve as a potential target for HBV-related HCC treatment.

Impact of N-Terminal Tags on De Novo Vimentin Intermediate Filament Assembly

“…These fusion proteins have been used in combination with untagged vimentin or transfected into vimentin expressing cells in order to study in vivo filament dynamics in live cells [ 40 ], fluorescence recovery after photobleaching [ 43 ] and fluorescence energy transfer to study intermolecular interactions [ 48 ]. Apart from these fluorescent tags, there are non-fluorescent tags such as N-Myc (EQKLISEEDL) [ 49 , 50 ], FLAG (DYKDDDDK) and 3 × FLAG [ 51 , 52 ], HA Tag (YPYDVPDYA) [ 53 ], Spot Tag (PDRVRAVSHWSS) [ 54 , 55 ], V5 Tag (GKPIPNPLLGLDST) [ 56 ] and His Tag (HHHHHH) [ 57 , 58 ]. These are small non-fluorescent peptides, which are fused in frame with the protein of interest so the fusion proteins can be tracked using tag-specific antibodies and therefore they are not suitable for live cell imaging.…”

Regulation of Pattern-Recognition Receptor Signaling by HBX During Hepatitis B Virus Infection

“…Depending on STAT3, HBX promotes the expression of HMGB1 to enhance EMT in liver cancer cells ( 121 ). Additionally, our results have shown that HBX can activate STAT3 through LASP1 to facilitate vimentin expression and enhance EMT ( 128 ). Similar to STAT3, STAT5b participates in the induction of EMT mediated by HBX ( 129 ).…”

HBV HBx-Downregulated lncRNA LINC01010 Attenuates Cell Proliferation by Interacting with Vimentin

“…According to previous reports, the expression of vimentin protein is associated with HBx in HBV-related tumor tissues. The researcher showed that HBx could enhance vimentin expression to facilitate EMT in hepatoma cells [40]. Recently, it has been reported that HBx also regulates the transcription of lncRNAs.…”