Background:Tumor metastasis is a major factor for poor prognosis of hepatocellular carcinoma(HCC),but the relationship between ubiquitination and metastasis need to be studiedmore systematically.We analyzed the ubiqui...Background:Tumor metastasis is a major factor for poor prognosis of hepatocellular carcinoma(HCC),but the relationship between ubiquitination and metastasis need to be studiedmore systematically.We analyzed the ubiquitinome of HCC in this study to have a more comprehensive insight into human HCC metastasis.Methods:The protein ubiquitination levels in 15 HCC specimens with vascular invasion and 15 without vascular invasion were detected by ubiquitinome.Proteins with significantly different ubiquitination levels between HCCs with and without vascular invasion were used to predict E3 ubiquitin ligases associated with tumor metastasis.The topological network of protein substrates and corresponding E3 ubiquitin ligaseswas constructed to identify the key E3 ubiquitin ligase.Besides,the growth,migration and invasion ability of LM3 and HUH7 hepatoma cell lines with andwithout SYVN1 expression interferencewere measured by cell proliferation assay,subcutaneous tumor assay,umphal vein endothelium tube formation assay,transwell migration and invasion assays.Finally,the interacting proteins of SYVN1 were screened and verified by protein interaction omics,immunofluorescence,and immunoprecipitation.Ubiquitin levels of related protein substrates in LM3 and HUH7 cells were compared in negative control,SYVN1 knockdown,and SYVN1 overexpression groups.Results:In this study,our whole-cell proteomic dataset and ubiquitinomic dataset contained approximately 5600 proteins and 12,000 ubiquitinated sites.We discovered increased ubiquitinated sites with shorter ubiquitin chains during the progression ofHCC metastasis.In addition,proteomic and ubiquitinomic analyses revealed that high expression of E3 ubiquitin-protein ligase SYVN1 is related with tumor metastasis.Furthermore,we found that SYVN1 interacted with heat shock protein 90(HSP90)and impacted the ubiquitination of eukaryotic elongation factor 2 kinase(EEF2K).Conclusions:The ubiquitination profiles of HCC with and without vascular invasion were significantly different.SYVN1 was the most important E3 ubiquitin-protein ligase responsible for this phenomenon,and itwas related with tumormetastasis and growth.Therefore,SYVN1might be a potential therapeutic target for HCC.展开更多
基金National Key Research and Development Program,Grant/Award Number:2017YFC1200100National Natural Science Foundation of China,Grant/Award Number:81400589Chinese National Science and Technology Major Project of the 13th Five-year plan,Grant/Award Number:2017ZX10202202-001-008。
文摘Background:Tumor metastasis is a major factor for poor prognosis of hepatocellular carcinoma(HCC),but the relationship between ubiquitination and metastasis need to be studiedmore systematically.We analyzed the ubiquitinome of HCC in this study to have a more comprehensive insight into human HCC metastasis.Methods:The protein ubiquitination levels in 15 HCC specimens with vascular invasion and 15 without vascular invasion were detected by ubiquitinome.Proteins with significantly different ubiquitination levels between HCCs with and without vascular invasion were used to predict E3 ubiquitin ligases associated with tumor metastasis.The topological network of protein substrates and corresponding E3 ubiquitin ligaseswas constructed to identify the key E3 ubiquitin ligase.Besides,the growth,migration and invasion ability of LM3 and HUH7 hepatoma cell lines with andwithout SYVN1 expression interferencewere measured by cell proliferation assay,subcutaneous tumor assay,umphal vein endothelium tube formation assay,transwell migration and invasion assays.Finally,the interacting proteins of SYVN1 were screened and verified by protein interaction omics,immunofluorescence,and immunoprecipitation.Ubiquitin levels of related protein substrates in LM3 and HUH7 cells were compared in negative control,SYVN1 knockdown,and SYVN1 overexpression groups.Results:In this study,our whole-cell proteomic dataset and ubiquitinomic dataset contained approximately 5600 proteins and 12,000 ubiquitinated sites.We discovered increased ubiquitinated sites with shorter ubiquitin chains during the progression ofHCC metastasis.In addition,proteomic and ubiquitinomic analyses revealed that high expression of E3 ubiquitin-protein ligase SYVN1 is related with tumor metastasis.Furthermore,we found that SYVN1 interacted with heat shock protein 90(HSP90)and impacted the ubiquitination of eukaryotic elongation factor 2 kinase(EEF2K).Conclusions:The ubiquitination profiles of HCC with and without vascular invasion were significantly different.SYVN1 was the most important E3 ubiquitin-protein ligase responsible for this phenomenon,and itwas related with tumormetastasis and growth.Therefore,SYVN1might be a potential therapeutic target for HCC.
