title: The ubiquitin-proteasome pathway is important for dengue virus infection in primary human endothelial cells.
abstract: Dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) are the most severe forms of dengue virus infection with hemorrhage and plasma leakage. However, pathogenic mechanisms of DHF and DSS remain poorly understood. We therefore investigated host responses as determined by changes in the cellular proteome of primary human endothelial cells upon infection with dengue virus serotype 2 (DEN-2) at a multiplicity of infection (MOI) of 10 for 24 h. Two-dimensional PAGE and quantitative intensity analysis revealed 38 significantly altered protein spots (16 upregulated and 22 downregulated) in DEN-2-infected cells compared to mock controls. These altered proteins were successfully identified by mass spectrometry, including those involved in oxidative stress response, transcription and translation, cytoskeleton assembly, protein degradation, cell growth regulation, apoptosis, cellular metabolism, and antiviral response. The proteomic data were validated by Western blot analyses [upregulated ubiquitin-activating enzyme E1 (UBE1) and downregulated annexin A2] and an immunofluorescence study (upregulated MxA). Interestingly, we found that MxA was colocalized with DEN-2 viral capsid protein, strengthening its role as an antiviral protein. Moreover, we also identified upregulation of a proteasome subunit. Our functional study revealed the significant role of ubiquitination in dengue infection and UBE1 inhibition by its specific inhibitor (UBEI-41) caused a significant reduction in the level of viral protein synthesis and its infectivity. Our findings suggest that various biological processes were triggered in response to dengue infection, particularly antiviral IFN and ubiquitin-proteasome pathways.
authors: Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
authors: Rattiyaporn
authors: R
authors: Kanlaya
authors: Sa-nga
authors: SN
authors: Pattanakitsakul
authors: Supachok
authors: S
authors: Sinchaikul
authors: Shui-Tein
authors: ST
authors: Chen
authors: Visith
authors: V
authors: Thongboonkerd
mesh_terms: D017306:Annexin A2 / Q000378:metabolism; D015153:Blotting, Western; D036022:Capsid Proteins / Q000378:metabolism; D002478:Cells, Cultured; D003716:Dengue Virus / Q000378:metabolism / Q000502:physiology; D015180:Electrophoresis, Gel, Two-Dimensional; D042783:Endothelial Cells / Q000378:metabolism* / Q000821:virology; D019204:GTP-Binding Proteins / Q000378:metabolism; D054884:Host-Pathogen Interactions; D006801:Humans; D064547:Myxovirus Resistance Proteins; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D015398:Signal Transduction; D025801:Ubiquitin / Q000378:metabolism*; D044764:Ubiquitin-Activating Enzymes / Q000378:metabolism
chemical_list: D017306:Annexin A2; D036022:Capsid Proteins; C579071:MX1 protein, human; D064547:Myxovirus Resistance Proteins; C471538:UBA1 protein, human; D025801:Ubiquitin; D046988:Proteasome Endopeptidase Complex; D019204:GTP-Binding Proteins; D044764:Ubiquitin-Activating Enzymes