Identification and characterization of cellular proteins interacting with Hepatitis E virus untranslated regions

• RNA affinity chromatography was employed to identify HEV 5′ and 3′ UTR interacting proteins.
• The UTR–host protein interaction was further validated using viral RNA immunoprecipitation.
• DHX9, PTK-7, DIS3 and TCR E chain of three cell lines were found interacting with HEV 3′UTR.
• RAD50 and TLE-4 of three cell lines were found interacting with HEV 5′UTR.
• Exosomes might have important implication in HEV life cycle.

Lack of robust cell culture systems for Hepatitis E virus (HEV) infection has hampered understanding of HEV biology. We attempted to identify the host cellular factors that interact with HEV 5′ and 3′ untranslated regions (UTRs) by RNA affinity chromatography followed by mass spectrometry analysis. Hepatitis E virus genotype-1 (HEV-1) and Hepatitis E virus genotype-4 (HEV-4) and three cell lines (HepG2/C3A, A549 and Caco2) were employed to understand the UTR–host protein interaction. RNA pull-down and matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI TOF/TOF) analysis revealed that DHX9, PTK-7, DIS3 and TCR E chain (CD3ɛ) of all the three cell lines interacted with HEV 3′UTR while RAD50 and TLE-4 interacted with HEV 5′UTR. RNA immuno-precipitation studies further confirmed the interaction of DHX9, DIS3 and TCR E chain. The expression changes in genes associated with the identified proteins were quantitated in the peripheral blood mononuclear cells (PBMCs) of Hepatitis E patients during acute and recovery phases. The data revealed that HEV infection influences the exosomes, T cell receptor signalling and Wnt signalling pathways. Interactions of DIS3 with HEV UTRs suggest that exosomes might have important implication in HEV life cycle.