Conference proceeding
Interference of MERS-CoV accessory genes with the innate immune response and their contribution to virulence
Congreso Nacional de Virología, Vol.XIV
06/11/2017
Abstract
Middle East respiratory syndrome coronavirus (MERS-CoV) is an emerging coronavirus infecting humans, associated with acute pneumoniaand a high mortality rate around 36%, which is considered a public health threat. No vaccines or antivirals are still available,making investigation for therapeutic strategies an urgent need. A collection of recombinant MERS-CoVs deficient in the genus-specific genes 3, 4a, 4b and 5 was generated from cDNA clonesusing the reverse genetics system previously developed in our laboratory.The contribution of 3, 4a, 4b and 5 proteins asantagonistsof the innate immune response (IIR) during infection was studied by analyzing in MERS-CoV deletion mutants the expression and subcellular localization of IIR mediators using qPCR, Western-blot and immunofluorescence techniques. The growth kineticsof recombinant viruses was similar to that of the wild-type (WT) virus, indicating that accessory genes werenot essential for MERS-CoV replication in cell cultures. Deletion of 4b gene significantly increased the expression of NF-kB-induced proinflammatory cytokines IL-6, IL-8 and TNF, suggesting 4b protein contribution to the inhibition of the NF-B signaling pathway in the context of infection. Mutations in 4b protein nuclear localization signal (NLS)prevented its localization to the nucleus and increased the expression of NF-kB-induced proinflammatory cytokines, suggesting that nuclear localization of protein 4b was required for its IIR antagonism. Co-immunoprecipitationand mass spectrometry analysisidentified the binding of 4b protein to karyopherins3 (KPNA3) and 4 (KPNA4), which mediate the translocation of proteins from the cytosol to the nucleus, including the transcription factor NF-kB.Binding of 4b to KPNA4was NLS-dependent.Moreover, the NF-B-KPNA4 binding was reducedin the presence of 4b, suggesting a mechanism of competitive inhibition.Mutationof 4b-NLS in recombinant MERS-CoVs caused accumulation of 4b in the cytoplasm and translocation ofNF-kBto the nucleus, leading to the expression of NF-kB-dependent proinflammatory cytokines. In contrast, in MERS-CoV WT infection, the presence of 4b protein in the nucleus was associated with the accumulation of NF-kBin the cytoplasm. Together, these results suggest that the binding of MERS-CoV 4b protein to karyopherins during infection competitively inhibits the nuclear translocation of NF-kB and, as a consequence, the expression of proinflammatory cytokines. Since 4b protein interferes with the host innate immune response, itmay contribute to MERS-CoV virulence and itsdeletion will lead to attenuated viruses that represent vaccine candidates. Inhibitors of cell signaling pathways involved in the interaction with 4b protein will inform antiviral candida
Details
- Title: Subtitle
- Interference of MERS-CoV accessory genes with the innate immune response and their contribution to virulence
- Creators
- Javier CantónAnthony R FehrFrancisco J Gutierrez-AlvarezCarlos Castaño-RodríguezJosé M HonrubiaRaúl Fernandez-DelgadoMaría T Sanchez-AparicioAdolfo García-SastreStanley Perlman - University of Iowa, Microbiology and ImmunologyLuis Enjuanes SánchezIsabel Solá Gurpegui
- Resource Type
- Conference proceeding
- Publication Details
- Congreso Nacional de Virología, Vol.XIV
- ISSN
- 2172-6523
- Language
- English
- Date published
- 06/11/2017
- Academic Unit
- Stead Family Department of Pediatrics; Microbiology and Immunology; Iowa Neuroscience Institute; Infectious Disease (Pediatrics)
- Record Identifier
- 9984072073602771
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