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UL26 Attenuates IKK?-Mediated Induction of Interferon-Stimulated Gene (ISG) Expression and Enhanced Protein ISGylation during Human Cytomegalovirus Infection.

ABSTRACT: Viruses must negotiate cellular antiviral responses in order to replicate. Human cytomegalovirus (HCMV) is a prevalent betaherpesvirus that encodes a number of viral gene products that modulate cellular antiviral signaling. The HCMV UL26 gene has previously been found to attenuate cytokine-activated NF-?B signaling, yet the role that UL26 plays in modulating the host cell's global transcriptional response to infection is not clear. Here, we find that infection with a UL26 deletion virus (?UL26) induces a proinflammatory transcriptional environment that includes substantial increases in the expression of cytokine signaling genes relative to wild-type HCMV. These increases include NF-?B-regulated genes as well as interferon-stimulated genes (ISGs), such as ISG15 and bone marrow stromal cell antigen 2 (BST2). The ?UL26 mutant-mediated induction of ISG15 expression was found to drive increases in global protein ISGylation during ?UL26 mutant infection. However, short hairpin RNA (shRNA) and CRISPR-mediated targeting of ISG15 indicated that its induction does not restrict HCMV infection. In contrast, shRNA-mediated targeting of BST2 demonstrated that BST2 restricts HCMV cell-to-cell spread. In addition, the increased expression of both of these ISGs and the global enhancement in protein ISGylation were found to be dependent on the activity of the canonical inhibitor of NF-?B kinase beta (IKK?). Both CRISPR-based and pharmacologically mediated inhibition of IKK? blocked the induction of ISG15 and BST2. These results suggest significant cross-talk between the NF-?B and interferon signaling pathways and highlight the importance of IKK signaling and the HCMV UL26 protein in shaping the antiviral response to HCMV.IMPORTANCE Modulation of cellular antiviral signaling is a key determinant of viral pathogenesis. Human cytomegalovirus (HCMV) is a significant source of morbidity in neonates and the immunosuppressed that contains many genes that modulate antiviral signaling, yet how these genes contribute to shaping the host cell's transcriptional response to infection is largely unclear. Our results indicate that the HCMV UL26 protein is critical in preventing the establishment of a broad cellular proinflammatory transcriptional environment. Further, we find that the host gene IKK? is an essential determinant governing the host cell's antiviral transcriptional response. Given their importance to viral pathogenesis, continuing to elucidate the functional interactions between viruses and the cellular innate immune response could enable the development of therapeutic strategies to limit viral infection.

SUBMITTER: Goodwin CM

PROVIDER: S-EPMC6854504 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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UL26 Attenuates IKKβ-Mediated Induction of Interferon-Stimulated Gene (ISG) Expression and Enhanced Protein ISGylation during Human Cytomegalovirus Infection.

Goodwin Christopher M CM Schafer Xenia X Munger Joshua J

Journal of virology 20191113 23

Viruses must negotiate cellular antiviral responses in order to replicate. Human cytomegalovirus (HCMV) is a prevalent betaherpesvirus that encodes a number of viral gene products that modulate cellular antiviral signaling. The HCMV UL26 gene has previously been found to attenuate cytokine-activated NF-κB signaling, yet the role that UL26 plays in modulating the host cell's global transcriptional response to infection is not clear. Here, we find that infection with a U ...[more]

PMID: 31534044

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Project description:Bunyaviruses pose a significant threat to human health, prosperity, and food security. In response to viral infections, interferons (IFNs) upregulate the expression of hundreds of interferon-stimulated genes (ISGs), whose cumulative action can potently inhibit the replication of bunyaviruses. We used a flow cytometry-based method to screen the ability of ?500 unique ISGs from humans and rhesus macaques to inhibit the replication of Bunyamwera orthobunyavirus (BUNV), the prototype of both the Peribunyaviridae family and the Bunyavirales order. Candidates possessing antibunyaviral activity were further examined using a panel of divergent bunyaviruses. Interestingly, one candidate, ISG20, exhibited potent antibunyaviral activity against most viruses examined from the Peribunyaviridae, Hantaviridae, and Nairoviridae families, whereas phleboviruses (Phenuiviridae) largely escaped inhibition. Similar to the case against other viruses known to be targeted by ISG20, the antibunyaviral activity of ISG20 is dependent upon its functional RNase activity. Through use of an infectious virus-like particle (VLP) assay (based on the BUNV minigenome system), we confirmed that gene expression from all 3 viral segments is strongly inhibited by ISG20. Using in vitro evolution, we generated a substantially ISG20-resistant BUNV and mapped the determinants of ISG20 sensitivity/resistance. Taking all the data together, we report that ISG20 is a broad and potent antibunyaviral factor but that some bunyaviruses are remarkably ISG20 resistant. Thus, ISG20 sensitivity/resistance may influence the pathogenesis of bunyaviruses, many of which are emerging viruses of clinical or veterinary significance.IMPORTANCE There are hundreds of bunyaviruses, many of which cause life-threatening acute diseases in humans and livestock. The interferon (IFN) system is a key component of innate immunity, and type I IFNs limit bunyaviral propagation both in vitro and in vivo Type I IFN signaling results in the upregulation of hundreds of IFN-stimulated genes (ISGs), whose concerted action generates an "antiviral state." Although IFNs are critical in limiting bunyaviral replication and pathogenesis, much is still unknown about which ISGs inhibit bunyaviruses. Using ISG-expression screening, we examined the ability of ?500 unique ISGs to inhibit Bunyamwera orthobunyavirus (BUNV), the prototypical bunyavirus. Using this approach, we identified ISG20, an interferon-stimulated exonuclease, as a potent inhibitor of BUNV. Interestingly, ISG20 possesses highly selective antibunyaviral activity, with multiple bunyaviruses being potently inhibited while some largely escape inhibition. We speculate that the ability of some bunyaviruses to escape ISG20 may influence their pathogenesis.

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Dataset Information

UL26 Attenuates IKK?-Mediated Induction of Interferon-Stimulated Gene (ISG) Expression and Enhanced Protein ISGylation during Human Cytomegalovirus Infection.

Publications

U<sub>L</sub>26 Attenuates IKKβ-Mediated Induction of Interferon-Stimulated Gene (ISG) Expression and Enhanced Protein ISGylation during Human Cytomegalovirus Infection.

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