The expression profile of the ubiquitin-like modifier FAT10 in immune cells suggests cell type-specific functions.

The TNF and IFN-γ-inducible ubiquitin-like modifier HLA-F adjacent transcript 10 (FAT10) is most prominently expressed in immunological tissues but information regarding basal expression and inducibility of FAT10 in the different types of immune cells is still lacking. Hence, we investigated FAT10 mRNA expression in the major human and murine immune cell subsets, and FAT10 protein expression in human leukocytes. We isolated the different human leukocytes from peripheral blood and the murine immune cell subsets from spleen. The purified leukocytes were left untreated or stimulated with TNF and INF-γ or LPS to induce FAT10 followed by quantitative real-time PCR or western blot analysis. Basal expression of FAT10 mRNA and protein was generally low but strongly up-regulated by IFN-γ and TNF in all immune cell subsets. LPS treatment induced FAT10 expression marginally in human CD8+ T cells and murine granulocytes, but it increased Fat10 expression significantly in murine regulatory T cells. Yet, in human CD8+ T cells, natural killer cells, natural killer T cells, and dendritic cells, the FAT10 mRNA was expressed without induction. Similarly, murine macrophages, monocytes, and regulatory T cells expressed Fat10 in the absence of stimulation. In summary, our findings suggest particular functions of FAT10 in these cell types. Furthermore, we observed not only a cell type-specific but also a species-specific basal FAT10 expression profile. Our data will serve as a guideline for future investigations to further elucidate FAT10's role in the immune system.

FAT10 is phosphorylated by IKKß to inhibit the antiviral type-I interferon response.

IFN-I secretion provides a rapid host defense against infection with RNA viruses. Within the host cell, viral RNA triggers the activation of the RIG-I signaling pathway, leading to the production of IFN-I. Because an exaggerated IFN-I response causes severe tissue damage, RIG-I signaling is tightly regulated. One of the factors that control the IFN-I response is the ubiquitin-like modifier FAT10, which is induced by TNF and IFNγ and targets covalently FAT10-linked proteins for proteasomal degradation. However, the mechanism of how FAT10 modulates IFN-I secretion remains to be fully elucidated. Here, we provide strong evidence that FAT10 is phosphorylated by IκB kinase ß (IKKß) upon TNF stimulation and during influenza A virus infection on several serine and threonine residues. FAT10 phosphorylation increases the binding of FAT10 to the TRAF3-deubiquitylase OTUB1 and its FAT10-mediated activation. Consequently, FAT10 phosphorylation results in a low ubiquitylation state of TRAF3, which is unable to maintain interferon regulatory factor 3 phosphorylation and downstream induction of IFN-I. Taken together, we reveal a mechanism of how phosphorylation of FAT10 limits the production of tissue-destructive IFN-I in inflammation.

Regulation of Interferon Induction by the Ubiquitin-Like Modifier FAT10.

The revelation that the human major histocompatibility complex (MHC) class I locus encodes a ubiquitin-like protein designated HLA-F adjacent transcript 10 (FAT10) or ubiquitin D (UBD) has attracted increasing attention to the function of this protein. Interestingly, the pro-inflammatory cytokines interferon (IFN)-γ and tumor necrosis factor (TNF) α synergize to strongly induce FAT10 expression, thereby suggesting a role of FAT10 in the immune response. Recent reports that FAT10 downregulates type I interferon production while it upregulates IFN-γ pose mechanistic questions on how FAT10 differentially regulates interferon induction. Several covalent and non-covalent binding partners of FAT10 involved in signal transduction pathways leading to IFN synthesis have been identified. After introducing FAT10, we review here recent insights into how FAT10 affects proteins in the interferon pathways, like the virus-responsive pattern recognition receptor RIG-I, the ubiquitin ligase ZNF598, and the deubiquitylating enzyme OTUB1. Moreover, we outline the consequences of FAT10 deficiency on interferon synthesis and viral expansion in mice and human cells. We discuss the need for covalent isopeptide linkage of FAT10 to the involved target proteins and the concomitant targeting for proteasomal degradation. After years of investigating the elusive biological functions of this fascinating ubiquitin-like modifier, we review the emerging evidence for a novel role of FAT10 in interferon regulation.

The ubiquitin-like modifier FAT10 is required for normal IFN-γ production by activated CD8+ T cells.

FAT10 is the only ubiquitin-like modifier which directly targets its substrate proteins for rapid degradation by the proteasome. While the conjugation and proteasomal targeting of FAT10 are fairly well understood, the biological functions of FAT10 have remained largely elusive. Here we have investigated the role of FAT10 in cytokine responses in mice upon viral infection. We used lymphocytic choriomeningitis virus (LCMV) infection of mice to induce the IFN-γ and TNF-α-dependent expression of FAT10. We found that TCR-stimulated splenocytes derived from LCMV-infected FAT10-/- mice secreted less IFN-γ and expressed less mRNA for IL-12 p40 but secreted more IFN-α and IFN-ß compared to FAT10+/- mice. The reduction in IFN-γ secretion could be assigned to CD8+ T cells. Nevertheless, LCMV viral clearance was similar in FAT10-/- as compared to FAT10+/- mice. Since FAT10 has previously been reported to promote influenza A virus (IAV) replication in vitro we have studied the effect of FAT10 deficiency during IAV infection in mice. Unexpectedly, IAV titers and disease symptoms were not changed in FAT10-/- mice even though the Fat10 mRNA was rapidly induced in the lung upon IAV infection. In conclusion, we find that FAT10 fine-tunes the balance of interferons during viral infection by lowering the production of type I and enhancing type II interferons.