Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, Jiangsu, People's Republic of China.
Clinical Laboratory, Xuzhou TCM Hospital Affiliated to Nanjing University of Chinese Medicine, Xuzhou, Jiangsu, People's Republic of China.
Cell Commun Signal. 2021 Mar 15;19(1):33. doi: 10.1186/s12964-021-00714-1.
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.
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.
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.
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. Video abstract.
乙型肝炎病毒（HBV）X 蛋白（HBX）已被报道负责 HBV 相关肝细胞癌（HCC）中的上皮-间充质转化（EMT）。波形蛋白是 EMT 相关的分子标志物。然而，HBX 介导的 HCC 发病机制中波形蛋白的重要性尚未得到很好的确定。
通过 Western blot 和免疫组织化学分析，检测 HBX 诱导的波形蛋白表达，以及 LIM 和 SH3 结构域蛋白 1（LASP1）在肝癌细胞中 HBX 诱导的波形蛋白表达中的作用。通过 Western blot、免疫共沉淀和 GST 下拉实验评估参与波形蛋白表达的信号通路、HBX 与波形蛋白和 LASP1 的相互作用，以及 LASP1 介导的 HBX 阳性细胞中波形蛋白的稳定性。通过 Western blot、CCK-8 检测、平板克隆形成检测、Transwell 检测和划痕愈合检测，探讨 HBX 和 LASP1 介导的波形蛋白在 HCC 细胞 EMT、增殖和迁移中的作用。
HBX 阳性肝癌细胞和 HBV 相关 HCC 组织中波形蛋白表达增加，且 HBX 相关 HCC 组织中波形蛋白的表达与 HBX 相关。功能上，波形蛋白有助于 HBX 介导的肝癌细胞 EMT、增殖和迁移。机制分析表明，HBX 能够通过 LASP1 增强波形蛋白的表达。一方面，PI3-K、ERK 和 STAT3 信号通路参与 LASP1 在 HBX 阳性肝癌细胞中上调波形蛋白。另一方面，HBX 可以直接与波形蛋白和 LASP1 相互作用，并且依赖于 LASP1，HBX 通过保护其免受泛素化介导的蛋白降解来促进波形蛋白的稳定性。除此之外，波形蛋白参与 HBX 阳性肝癌细胞中 LASP1 介导的肝癌细胞的生长和迁移。
总之，这些发现表明，依赖于 LASP1，波形蛋白对于 HBX 介导的 EMT 和肝癌发生至关重要，并且可能成为治疗 HBV 相关 HCC 的潜在靶点。
