The CCCH-Type Zinc Finger Antiviral Protein Relieves Immunosuppression of T Cells Induced by Avian Leukosis Virus Subgroup J via the NLP-PKC-δ-NFAT Pathway.

The CCCH-type zinc finger antiviral protein (ZAP) can recognize and induce the degradation of mRNAs and proteins of certain viruses, as well as exerting its antiviral activity by activating T cells. However, the mechanism of ZAP that mediates T cell activation during virus infection remains unclear. Here, we found a potential function of ZAP that relieves immunosuppression of T cell induced by avian leukosis virus subgroup J (ALV-J) via a novel signaling pathway that involves norbin-like protein (NLP), protein kinase C delta (PKC-δ), and nuclear factor of activated T cell (NFAT). Specifically, ZAP expression activated T cells by promoting the dephosphorylation and nuclear translocation of NFAT. Furthermore, knockdown of ZAP weakened the reactivity and antiviral response of T cells. Mechanistically, ZAP reduced PKC-δ activity by upregulating and reactivating NLP by competitively binding with viral protein. Knockdown of NLP decreased the dephosphorylation of PKC-δ by ZAP expression. Moreover, we show that knockdown of PKC-δ reduced the phosphorylation levels of NFAT and enhanced its nuclear translocation. Taken together, these data revealed that ZAP relieves immunosuppression caused by ALV-J and mediates T cell activation through the NLP-PKC-δ-NFAT pathway. IMPORTANCE The evolution of the host defense system is driven synchronously in the process of resisting virus invasion. Accordingly, host innate defense factors effectively work to suppress virus replication. However, it remains unclear whether the host innate defense factors are involved in antiviral immune responses against the invasion of immunosuppressive viruses. Here, we found that CCCH-type zinc finger antiviral protein (ZAP) effectively worked in resistance to immunosuppression caused by avian leukosis virus subgroup J (ALV-J), a classic immunosuppressive virus. Evidence showed that ZAP released the phosphatase activity of NLP inhibited by ALV-J and further activated NFAT by inactivating PKC-δ. This novel molecular mechanism, i.e., ZAP regulation of the antiviral immune response by mediating the NLP-PKC-δ-NFAT pathway, has greatly enriched the understanding of the functions of host innate defense factors and provided important scientific ideas and a theoretical basis for research on immunosuppressive viruses and antiviral immunity.

Serum amyloid A regulates TLR2/4-mediated IFN-ß signaling pathway against Marek's disease virus.

Serum amyloid A (SAA), an acute response phase protein (APP), is crucial for the innate immune response during pathogenic microorganisms' invasion. Marek's disease virus (MDV) is a highly oncogenic alphaherpesvirus that activates multiple innate immune molecules, including SAA, in the host during infection. However, the pathway through which SAA participates in MDV-induced host innate immunity remains unknown. The present study aimed to elucidate the pathway through which SAA exerts its anti-MDV function. We observed that MDV infection in vivo and in vitro significantly elevated SAA expression. Furthermore, through SAA overexpression and knockdown experiments, we demonstrated that SAA could inhibit MDV replication. Subsequently, we found that SAA activated Toll-Like Receptor 2/4 (TLR2/4) -mediated Interferon Beta (IFN-ß) promoter activity and IFN regulatory factor 7 (IRF7) promoter activity. During MDV infection, SAA enhanced TLR2/4-mediated IFN-ß signal transduction and messenger RNAs (mRNAs) expression of type I IFN (IFN-I) and interferon-stimulated genes (ISGs). Finally, TLR2/4 inhibitor OxPAPC inhibits the anti-MDV activity of SAA. These results demonstrated that SAA inhibits MDV replication and enhancing TLR2/4-mediated IFN-ß signal transduction to promote IFNs and ISGs expression. This finding is the first to demonstrate the signaling pathway by which SAA exerts its anti-MDV function. It also provides new insights into the control of oncogenic herpesviruses from the perspective of acute response phase proteins.

Assembly encapsulation of BSA and CCCH-ZAP in the sodium alginate/atractylodis macrocephalae system.

Zinc finger antiviral proteins (ZAP) can significantly inhibit the replication of avian leukosis virus subgroup J (ALV-J), but the traditional method of ZAP administration is by injection, which can easily cause stress effects in chickens. In this work, we established a sodium alginate/atractylodis macrocephalae system for the encapsulation of CCCH-type zinc finger antiviral protein (CCCH-ZAP). Because of the high cost of ZAP, we first chose bovine serum albumin (BSA) as a model protein to investigate the encapsulation performance. The SEM images clearly confirmed that BSA and the sodium alginate/atractylodis macrocephalae system can assemble easily to form relatively stable nanostructures, and the encapsulation amount of BSA can reach 68%. Subsequently, the encapsulation of ZAP was studied. The SEM and the encapsulation experiments confirmed that ZAP can also be assembly encapsulated in the sodium alginate/atractylodis macrocephalae system with the encapsulation amount of 80%. Release studies showed that the SA/AM-ZAP nanocomposite was able to achieve a release rate of 32% of ZAP. This work successfully confirms the assembly encapsulation of ZAP, which will be beneficial for the usage of ZAP-based animal drugs.

Interaction of p10/p27 with macrophage migration inhibitory factor promotes avian leukosis virus subgroup J infection.

Avian leukosis virus subgroup J (ALV-J), an oncogenic retrovirus, induces myelocytoma and various other tumors in broilers and layers. Many recent studies have shown that ALV-J can hijack host molecules to facilitate infection. However, the molecular mechanisms of this process are not clear. Here, we aimed to elucidate the molecular mechanisms contributing to ALV-J infection. ALV-J hijacked MIF via p10 and p27 to facilitate ALV-J infection. ALV-J persistently activated MIF expression in DF-1 cells, and MIF significantly facilitated ALV-J internalization and replication, which demonstrated by MIF overexpression and knockdown experiments and treatment with the MIF antagonist ISO-1. Furthermore, we found that the two subunit proteins of Gag, p10 and p27, interacted with MIF in the cytoplasm, respectively. These results suggested that the p10 and p27 subunit in Gag protein recruited MIF to promote ALV-J infection, providing insights into the roles of the p10/p27 and the host factor MIF in ALV-J infection. The finding may facilitate the development of new strategies for controlling ALV-J or retrovirus infections.