Viperin is an iron-sulfur protein that inhibits genome synthesis of tick-borne encephalitis virus via radical SAM domain activity

Upadhyay, Arunkumar S.; Vonderstein, Kirstin; Pichlmair, Andreas; Stehling, Oliver; Bennett, Keiryn L.; Dobler, Gerhard; Guo, Ju‐Tao; Superti‐Furga, Giulio; Lill, Roland; Överby, Anna K.; Weber, Friedemann

doi:10.1111/cmi.12241

Cited by 92 publications

(148 citation statements)

References 51 publications

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“…In contrast to our results with FPPS, it was shown that, whereas localization of viperin to the ER was essential to reduce the titer of TBEV, this was not dependent on identity of the ER-localizing sequence (24). Significantly, mutation of the conserved cysteine residues or the C-terminal tryptophan of viperin, which is important for installation of the [Fe 4 S 4 ] cluster by CIAO1, abolished viperin inhibition of TBEV replication, indicating that an intact [Fe 4 S 4 ] cluster was essential for activity.…”

Section: Discussioncontrasting

confidence: 99%

“…Unlike viruses such as influenza A that bud from cholesterol-rich lipid rafts and are thus sensitive to FPPS inhibition, flaviviruses such as TBEV do not require lipid rafts for budding. Indeed it has been shown that inhibition of FPPS by ibandronate has no effect TBEV replication (24).…”

Section: Discussionmentioning

confidence: 99%

“…It is one of eight putative radical SAM enzymes identified in humans; four of these appear to be involved in tRNA modifications, two seem to be involved in co-factor biosynthesis, and the remaining two, including viperin, have unknown enzymatic activities (23). The human enzyme is a 361-residue protein that comprises three distinct domains: an N-terminal amphipathic helix thought to play a role in localizing viperin to the endoplasmic reticulum and lipid droplets (19); a large, central domain that contains four motifs characteristic of radical SAM enzymes; and a conserved C-terminal domain that may be involved in [Fe 4 S 4 ] cluster assembly, the proper folding of viperin, and/or substrate recognition (20,24). Although no enzymatic function has yet been assigned to viperin, the radical SAM domain has been shown to be important in limiting HIV and Bunyamwera viruses (20).…”

Section: Radical S-adenosyl-l-methionine-dependent (Sam)mentioning

confidence: 99%

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Does Viperin Function as a Radical S-Adenosyl-l-methionine-dependent Enzyme in Regulating Farnesylpyrophosphate Synthase Expression and Activity?

Makins

Ghosh²,

Román‐Meléndez³

et al. 2016

Journal of Biological Chemistry

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Edited by Charles SamuelViperin is an endoplasmic reticulum-associated antiviral responsive protein that is highly up-regulated in eukaryotic cells upon viral infection through both interferon-dependent and independent pathways. Viperin is predicted to be a radical S-adenosyl-L-methionine (SAM) enzyme, but it is unknown whether viperin actually exploits radical SAM chemistry to exert its antiviral activity. We have investigated the interaction of viperin with its most firmly established cellular target, farnesyl pyrophosphate synthase (FPPS). Numerous enveloped viruses utilize cholesterol-rich lipid rafts to bud from the host cell membrane, and it is thought that by inhibiting FPPS activity (and therefore cholesterol synthesis), viperin retards viral budding from infected cells. We demonstrate that, consistent with this hypothesis, overexpression of viperin in human embryonic kidney cells reduces the intracellular rate of accumulation of FPPS but does not inhibit or inactivate FPPS. The endoplasmic reticulum-localizing, N-terminal amphipathic helix of viperin is specifically required for viperin to reduce cellular FPPS levels. However, although viperin reductively cleaves SAM to form 5-deoxyadenosine in a slow, uncoupled reaction characteristic of radical SAM enzymes, this cleavage reaction is independent of FPPS. Furthermore, mutation of key cysteinyl residues ligating the catalytic [Fe 4 S 4 ] cluster in the radical SAM domain, surprisingly, does not abolish the inhibitory activity of viperin against FPPS; indeed, some mutations potentiate viperin activity. These observations imply that viperin does not act as a radical SAM enzyme in regulating FPPS. Radical S-adenosyl-L-methionine-dependent (SAM)3 enzymes constitute a superfamily of enzymes that use SAM to generate free radicals (1-4). The superfamily is typified by a common CXXXCXXC motif, the cysteinyl residues of which coordinate a [Fe 4 S 4 ] cluster that is essential for radical generation. These enzymes catalyze a remarkably wide range of reactions involving an equally diverse set of substrates (2). For example, radical SAM-dependent enzymes participate in the biosynthesis of herbicides, antibiotics, vitamins, co-factors such as biotin and thiamin, and various other natural products (2, 5-8). They also function in the modification of ribosomal and transfer RNAs (9, 10) and DNA repair (11) and in the post-translational modification of peptides and proteins (12)(13)(14). Sequence analyses indicate that there are potentially thousands of members of the radical SAM superfamily, but to date, relatively few of these enzymes have been isolated and characterized (15, 16). Radical SAM enzymes were until recently thought to be confined to the microbial realm but intriguingly have now been identified in higher aerobic organisms, including plants and animals (3).Central to the mechanism of all radical SAM enzymes is the generation of a highly reactive 5Ј-deoxyadenosyl radical (Ado ⅐ ) (1,4,17). This is accomplished through one-electron reduction of SAM by the [Fe 4...

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Section: Discussioncontrasting

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Radical S-adenosyl-l-methionine-dependent (Sam)mentioning

confidence: 99%

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Does Viperin Function as a Radical S-Adenosyl-l-methionine-dependent Enzyme in Regulating Farnesylpyrophosphate Synthase Expression and Activity?

Makins

Ghosh²,

Román‐Meléndez³

et al. 2016

Journal of Biological Chemistry

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“…Early studies showed that viperin is induced in various cell types by IFN-α and IFN-β, associates with the cytosolic face of the endoplasmic reticulum (ER), and inhibits human cytomegalovirus replication when preexpressed in human fibroblasts (8). Since then, viperin has been shown to be induced by several factors, including lipopolysaccharide (10)(11)(12), and to inhibit a broad range of viruses, including HIV-1 (13), West Nile virus (14), hepatitis C virus (15,16), dengue virus type 2 (17), influenza A virus (18), and tick-borne encephalitis virus (19). Gene-profiling microarray studies have shown that the viperin gene is one of the most highly inducible ISGs upon infection with a wide range of RNA viruses (20).…”

mentioning

confidence: 99%

Structural studies of viperin, an antiviral radical SAM enzyme

Fenwick

Cresswell

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

112

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Viperin is an IFN-inducible radical S-adenosylmethionine (SAM) enzyme that inhibits viral replication. We determined crystal structures of an anaerobically prepared fragment of mouse viperin (residues 45-362) complexed with S-adenosylhomocysteine (SAH) or 5′-deoxyadenosine (5′-dAdo) and L-methionine (L-Met). Viperin contains a partial (βα) 6 -barrel fold with a disordered N-terminal extension (residues 45-74) and a partially ordered C-terminal extension (residues 285-362) that bridges the partial barrel to form an overall closed barrel structure. Cys84, Cys88, and Cys91 located after the first β-strand bind a [4Fe-4S] cluster. The active site architecture of viperin with bound SAH (a SAM analog) or 5′-dAdo and L-Met (SAM cleavage products) is consistent with the canonical mechanism of 5′-deoxyadenosyl radical generation. The viperin structure, together with sequence alignments, suggests that vertebrate viperins are highly conserved and that fungi contain a viperin-like ortholog. Many bacteria and archaebacteria also express viperin-like enzymes with conserved active site residues. Structural alignments show that viperin is similar to several other radical SAM enzymes, including the molybdenum cofactor biosynthetic enzyme MoaA and the RNA methyltransferase RlmN, which methylates specific nucleotides in rRNA and tRNA. The viperin putative active site contains several conserved positively charged residues, and a portion of the active site shows structural similarity to the GTPbinding site of MoaA, suggesting that the viperin substrate may be a nucleoside triphosphate of some type.radical SAM | IFN-stimulated gene | antiviral cellular factor | free radical | S-adenosyl methionine V iruses exploit the metabolic machinery of host cells to replicate and spread to other cells. Although cytotoxic T cells and antibody-producing B cells can ultimately be produced in an adaptive response to the virus, innate immune mechanisms are used to respond to infection rapidly. Upon infection, cells can sense the presence of virus via pattern recognition receptors (1, 2) and produce IFNs that limit the spread of infection to other cells (3). IFNs induce the expression of hundreds of IFN-stimulated genes (ISGs), many of which are involved in various antiviral processes, including antigen presentation, apoptosis, and inhibition of viral replication (4-7).Viperin, the product of rsad-2, was first identified as a protein induced by exposure of human macrophages to IFN-γ and by infection of primary human fibroblasts with human cytomegalovirus (8,9). Early studies showed that viperin is induced in various cell types by IFN-α and IFN-β, associates with the cytosolic face of the endoplasmic reticulum (ER), and inhibits human cytomegalovirus replication when preexpressed in human fibroblasts (8). Since then, viperin has been shown to be induced by several factors, including lipopolysaccharide (10-12), and to inhibit a broad range of viruses, including HIV-1 (13), West Nile virus (14), hepatitis C virus (15, 16), dengue virus type 2 (17), influ...

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“…The chemical versatility of these clusters is utilized in fundamental life processes such as energy production, metabolic conversions, DNA maintenance, gene expression regulation, protein translation, and the antiviral response ( Fig. 1) (2)(3)(4). In eukaryotes, Fe/S proteins are found in or associated with the mitochondrion, endoplasmic reticulum, cytosol, and the nucleus.…”

Section: Ubiquitous Iron-sulfur Clusters and Their Synthesis: An Overmentioning

confidence: 99%

Iron–sulfur cluster biogenesis and trafficking in mitochondria

Braymer

Lill

2017

Journal of Biological Chemistry

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314

293

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The biogenesis of iron-sulfur (Fe/S) proteins in eukaryotes is a multistage, multicompartment process that is essential for a broad range of cellular functions, including genome maintenance, protein translation, energy conversion, and the antiviral response. Genetic and cell biological studies over almost 2 decades have revealed some 30 proteins involved in the synthesis of cellular [2Fe-2S] and [4Fe-4S] clusters and their incorporation into numerous apoproteins. Mechanistic aspects of Fe/S protein biogenesis continue to be elucidated by biochemical and ultrastructural investigations. Here, we review recent developments in the pursuit of constructing a comprehensive model of Fe/S protein assembly in the mitochondrion. Ubiquitous iron-sulfur clusters and their synthesis: an overviewIron-sulfur (Fe/S) 2 clusters are inorganic cofactors that are essential for the proper functioning of virtually all biological cells (1). The chemical versatility of these clusters is utilized in fundamental life processes such as energy production, metabolic conversions, DNA maintenance, gene expression regulation, protein translation, and the antiviral response ( Fig. 1) (2-4). In eukaryotes, Fe/S proteins are found in or associated with the mitochondrion, endoplasmic reticulum, cytosol, and the nucleus. These cofactors participate in electron transfer reactions, Lewis acid catalysis, transfer of sulfur atoms, or facilitating structural roles (5, 6). Although the activities of some Fe/S proteins are dispensable for cell survival under certain conditions, for example fungal Fe/S enzymes in the metabolism of amino acids, others such as Fe/S proteins involved in DNA maintenance or protein translation are essential for cell viability (Fig. 1). The growing number of diseases that implicate Fe/S proteins or their assembly factors illustrates the essentiality of the various functions of these protein cofactors (7-9). The phenotypes associated with these "Fe/S diseases" and the in vivo work in model systems such as Saccharomyces cerevisiae and human cell culture have led to a mechanistic model of eukaryotic Fe/S protein biogenesis, in which the sequence of events required for proper synthesis and trafficking of Fe/S clusters has been elucidated (2). This model has been invaluable to diagnosing new mitochondrial disorders, and its continued advancement will enhance the ability for early diagnosis (10 -13). The focus of this review will be on the latest developments of the functional and mechanistic aspects that have advanced the model of mitochondrial Fe/S protein biogenesis.Cells typically maintain a strict balance of iron and sulfide ion concentrations due to their damaging redox reactions when present in excess (14, 15). The cell also uses a complex biosynthetic system to ensure that the sometimes redox-sensitive and labile Fe/S clusters are assembled correctly, trafficked to specific target apoproteins, and remain protected during these processes. This cellular control is exemplified by the 18 known "Fe/S cluster assembly" (ISC) proteins...

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Viperin is an iron-sulfur protein that inhibits genome synthesis of tick-borne encephalitis virus via radical SAM domain activity

Cited by 92 publications

References 51 publications

Does Viperin Function as a Radical S-Adenosyl-l-methionine-dependent Enzyme in Regulating Farnesylpyrophosphate Synthase Expression and Activity?

Does Viperin Function as a Radical S-Adenosyl-l-methionine-dependent Enzyme in Regulating Farnesylpyrophosphate Synthase Expression and Activity?

Structural studies of viperin, an antiviral radical SAM enzyme

Iron–sulfur cluster biogenesis and trafficking in mitochondria

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