ISG15 overexpression compensates the defect of Crimean-Congo hemorrhagic fever virus polymerase bearing a protease-inactive ovarian tumor domain

• Published in: PLoS neglected tropical diseases 2020-09, Vol.14 (9), p.e0008610-e0008610
• Main Authors: Devignot, Stephanie, Kromer, Thilo, Mirazimi, Ali, Weber, Friedemann
• Format: Article
• Language: English
• Subjects: Amino acids; Antiviral agents; Attenuation; Binding sites; Biology and Life Sciences; Conjugation; Context; Crimean hemorrhagic fever; Cytokines; Development and progression; Disease control; Domains; Fever; Genetic aspects; Genomes; Haemorrhage; Hemorrhage; Infections; Interferon; L protein; Length; Medicine and Health Sciences; Molecules; Mutation; Neoplasms; Ovarian cancer; Ovaries; Pathogens; Plasmids; Protease; Proteases; Protein-protein interactions; Proteinase; Proteins; Recombinants; Research and Analysis Methods; RNA polymerase; RNA polymerases; Signaling; Structure; Transcription; Tropical diseases; Tumors; Ubiquitin; Vaccines; Veterinary medicine; Viral proteins; Viral research; Virology; Virus-like particles; Viruses
• ISSN: 1935-2735; 1935-2727; 1935-2735
• DOI: 10.1371/journal.pntd.0008610

Crimean-Congo Hemorrhagic Fever virus (CCHFV; family Nairoviridae) is an extremely pathogenic member of the Bunyavirales order. Previous studies have shown that the N-terminal domain of the CCHFV polymerase (L) contains an ovarian tumor-type protease (OTU) domain with the capability to remove both ubiquitin and ISG15 molecules from proteins. The approximately 200 amino acids-long OTU domain, if ectopically expressed, can interfere with both the induction of antiviral type I interferons (IFN) as well as the IFN-stimulated signaling. A OTU protease mutant (C40A), by contrast, was inactive in that respect. However, the effect of the OTU protease activity in the context of the full-length L protein (approximately 4000 amino acids) is only poorly characterized, and recombinant CCHFV with the C40A mutation could not be rescued. Here, we employed transcriptionally active virus-like particles (tc-VLPs) to investigate the interaction between the L-embedded OTU protease and the IFN system. Our data show a cis requirement of the OTU protease for optimal CCHFV polymerase activity in human HuH-7 cells. The L-embedded OTU did not influence IFN signaling, the sensitivity to IFN, or IFN induction. Moreover, the attenuation of OTU C40A-mutated L could not be relieved by inactivating the IFN response, but after overexpression of conjugation-competent ISG15 the polymerase activity recovered to wild-type levels. Consequently, ISG15 was used to produce OTU-deficient tc-VLPs, a potential vaccine candidate. Our data thus indicate that in the context of full-length L the OTU domain is important for the regulation of CCHFV polymerase by ISG15.