ISG15/USP18/STAT2 is a molecular hub regulating IFN I-mediated control of Dengue and Zika virus replication.

The establishment of a virus infection is the result of the pathogen's ability to replicate in a hostile environment generated by the host's immune system. Here, we found that ISG15 restricts Dengue and Zika viruses' replication through the stabilization of its binding partner USP18. ISG15 expression was necessary to control DV replication driven by both autocrine and paracrine type one interferon (IFN-I) signaling.

PKR-mediated stress response enhances dengue and Zika virus replication.

One of the fundamental features that make viruses intracellular parasites is the necessity to use cellular translational machinery. Hence, this is a crucial checkpoint for controlling infections. Here, we show that dengue and Zika viruses, responsible for nearly 400 million infections every year worldwide, explore such control for optimal replication.

Myeloid cell interferon secretion restricts Zika flavivirus infection of developing and malignant human neural progenitor cells.

Zika virus (ZIKV) can infect human developing brain (HDB) progenitors resulting in epidemic microcephaly, whereas analogous cellular tropism offers treatment potential for the adult brain cancer, glioblastoma (GBM). We compared productive ZIKV infection in HDB and GBM primary tissue explants that both contain SOX2+ neural progenitors. Strikingly, although the HDB proved uniformly vulnerable to ZIKV infection, GBM was more refractory, and this correlated with an innate immune expression signature.

Functional role and ribosomal position of the unique N-terminal region of DHX29, a factor required for initiation on structured mammalian mRNAs.

Translation initiation on structured mammalian mRNAs requires DHX29, a DExH protein that comprises a unique 534-aa-long N-terminal region (NTR) and a common catalytic DExH core. DHX29 binds to 40S subunits and possesses 40S-stimulated NTPase activity essential for its function. In the cryo-EM structure of DHX29-bound 43S preinitiation complexes, the main DHX29 density resides around the tip of helix 16 of 18S rRNA, from which it extends through a linker to the subunit interface forming an intersubunit domain next to the eIF1A binding site.

Mouse Ifit1b is a cap1-RNA-binding protein that inhibits mouse coronavirus translation and is regulated by complexing with Ifit1c.

Knockout mouse models have been extensively used to study the antiviral activity of IFIT (interferon-induced protein with tetratricopeptide repeats). Human IFIT1 binds to cap0 (m7GpppN) RNA, which lacks methylation on the first and second cap-proximal nucleotides (cap1, m7GpppNm, and cap2, m7GpppNmNm, respectively). These modifications are signatures of "self" in higher eukaryotes, whereas unmodified cap0-RNA is recognized as foreign and, therefore, potentially harmful to the host cell.

The flavivirus polymerase NS5 regulates translation of viral genomic RNA.

Flaviviruses, including dengue virus and Zika virus, contain a single-stranded positive sense RNA genome that encodes viral proteins essential for replication and also serves as the template for new genome synthesis. As these processes move in opposite directions along the genome, translation must be inhibited at a defined point following infection to clear the template of ribosomes to allow efficient replication. Here, we demonstrate in vitro and in cell-based assays that the viral RNA polymerase, NS5, inhibits translation of the viral genome.

Circularization of flavivirus genomic RNA inhibits de novo translation initiation.

Members of the Flaviviridae family, including dengue virus (DENV) and yellow fever virus, cause serious disease in humans, whilst maternal infection with Zika virus (ZIKV) can induce microcephaly in newborns. Following infection, flaviviral RNA genomes are translated to produce the viral replication machinery but must then serve as a template for the transcription of new genomes. However, the ribosome and viral polymerase proceed in opposite directions along the RNA, risking collisions and abortive replication.

Ifit1 regulates norovirus infection and enhances the interferon response in murine macrophage-like cells.

Norovirus, also known as the winter vomiting bug, is the predominant cause of non-bacterial gastroenteritis worldwide. Disease control is predicated on a robust innate immune response during the early stages of infection. Double-stranded RNA intermediates generated during viral genome replication are recognised by host innate immune sensors in the cytoplasm, activating the strongly antiviral interferon gene programme.

Better together: the role of IFIT protein-protein interactions in the antiviral response.

The interferon-induced proteins with tetratricopeptide repeats (IFITs) are a family of antiviral proteins conserved throughout all vertebrates. IFIT1 binds tightly to non-self RNA, particularly capped transcripts lacking methylation on the first cap-proximal nucleotide, and inhibits their translation by out-competing the cellular translation initiation apparatus. This exerts immense selection pressure on cytoplasmic RNA viruses to maintain mechanisms that protect their messenger RNA from IFIT1 recognition.

IFIT3 and IFIT2/3 promote IFIT1-mediated translation inhibition by enhancing binding to non-self RNA.

Interferon-induced proteins with tetratricopeptide repeats (IFITs) are highly expressed during the cell-intrinsic immune response to viral infection. IFIT1 inhibits translation by binding directly to the 5' end of foreign RNAs, particularly those with non-self cap structures, precluding the recruitment of the cap-binding eukaryotic translation initiation factor 4F and ribosome recruitment. The presence of IFIT1 imposes a requirement on viruses that replicate in the cytoplasm to maintain mechanisms to avoid its restrictive effects.

Neurodevelopmental protein Musashi-1 interacts with the Zika genome and promotes viral replication.

A recent outbreak of Zika virus in Brazil has led to a simultaneous increase in reports of neonatal microcephaly. Zika targets cerebral neural precursors, a cell population essential for cortical development, but the cause of this neurotropism remains obscure. Here we report that the neural RNA-binding protein Musashi-1 (MSI1) interacts with the Zika genome and enables viral replication.

The RNA Helicase eIF4A Is Required for Sapovirus Translation.

The eukaryotic initiation factor 4A (eIF4A) is a DEAD box helicase that unwinds RNA structure in the 5' untranslated region (UTR) of mRNAs. Here, we investigated the role of eIF4A in porcine sapovirus VPg-dependent translation. Using inhibitors and dominant-negative mutants, we found that eIF4A is required for viral translation and infectivity, suggesting that despite the presence of a very short 5' UTR, eIF4A is required to unwind RNA structure in the sapovirus genome to facilitate virus translation.

A Cell-based Fluorescence Resonance Energy Transfer (FRET) Sensor Reveals Inter- and Intragenogroup Variations in Norovirus Protease Activity and Polyprotein Cleavage.

The viral protease represents a key drug target for the development of antiviral therapeutics. Because many protease inhibitors mimic protease substrates, differences in substrate recognition between proteases may affect their sensitivity to a given inhibitor. Here we use a cell-based FRET sensor to investigate the activity of different norovirus proteases upon cleavage of various norovirus cleavage sites inserted into a linker region separating cyan fluorescent protein and yellow fluorescent protein.

Inhibition of Translation Initiation by Protein 169: A Vaccinia Virus Strategy to Suppress Innate and Adaptive Immunity and Alter Virus Virulence.

Vaccinia virus (VACV) is the prototypic orthopoxvirus and the vaccine used to eradicate smallpox. Here we show that VACV strain Western Reserve protein 169 is a cytoplasmic polypeptide expressed early during infection that is excluded from virus factories and inhibits the initiation of cap-dependent and cap-independent translation. Ectopic expression of protein 169 causes the accumulation of 80S ribosomes, a reduction of polysomes, and inhibition of protein expression deriving from activation of multiple innate immune signaling pathways.

Inhibition of translation by IFIT family members is determined by their ability to interact selectively with the 5'-terminal regions of cap0-, cap1- and 5'ppp- mRNAs.

Ribosomal recruitment of cellular mRNAs depends on binding of eIF4F to the mRNA's 5'-terminal 'cap'. The minimal 'cap0' consists of N7-methylguanosine linked to the first nucleotide via a 5'-5' triphosphate (ppp) bridge. Cap0 is further modified by 2'-O-methylation of the next two riboses, yielding 'cap1' (m7GpppNmN) and 'cap2' (m7GpppNmNm).

The mechanism of translation initiation on Type 1 picornavirus IRESs.

Picornavirus Type 1 IRESs comprise five principal domains (dII-dVI). Whereas dV binds eIF4G, a conserved AUG in dVI was suggested to stimulate attachment of 43S ribosomal preinitiation complexes, which then scan to the initiation codon. Initiation on Type 1 IRESs also requires IRES trans-acting factors (ITAFs), and several candidates have been proposed.

Roles of individual domains in the function of DHX29, an essential factor required for translation of structured mammalian mRNAs.

On most eukaryotic mRNAs, initiation codon selection involves base-by-base inspection of 5' UTRs by scanning ribosomal complexes. Although the eukaryotic initiation factors 4A/4B/4G can mediate scanning through medium-stability hairpins, scanning through more stable structures additionally requires DHX29, a member of the superfamily 2 DEAH/RNA helicase A (RHA) helicase family that binds to 40S subunits and possesses 40S-stimulated nucleoside triphosphatase (NTPase) activity. Here, sequence alignment and structural modeling indicated that DHX29 comprises a unique 534-aa-long N-terminal region (NTR), central catalytic RecA1/RecA2 domains containing a large insert in the RecA2 domain, and the C-terminal part, which includes winged-helix, ratchet, and oligonucleotide/oligosaccharide-binding (OB) domains that are characteristic of DEAH/RHA helicases.

A distinct class of internal ribosomal entry site in members of the Kobuvirus and proposed Salivirus and Paraturdivirus genera of the Picornaviridae.

The 5'-untranslated regions (5' UTRs) of picornavirus genomes contain an internal ribosomal entry site (IRES) that promotes the end-independent initiation of translation. Picornavirus IRESs are classified into four structurally distinct groups, each with different initiation factor requirements. Here, we identify a fifth IRES class in members of Kobuvirus, Salivirus, and Paraturdivirus genera of Picornaviridae: Aichi virus (AV), bovine kobuvirus (BKV), canine kobuvirus (CKoV), mouse kobuvirus (MKoV), sheep kobuvirus (SKV), salivirus A (SV-A), turdivirus 2 (TV2), and TV3.

The mechanism of translation initiation on Aichivirus RNA mediated by a novel type of picornavirus IRES.

Picornavirus mRNAs contain IRESs that sustain their translation during infection, when host protein synthesis is shut off. The major classes of picornavirus IRESs (Types 1 and 2) have distinct structures and sequences, but initiation on both is determined by their specific interaction with eIF4G. We report here that Aichivirus (AV), a member of the Kobuvirus genus of Picornaviridae, contains an IRES that differs structurally from Type 1 and Type 2 IRESs.

Foot-and-mouth disease virus 2C is a hexameric AAA+ protein with a coordinated ATP hydrolysis mechanism.

Foot-and-mouth disease virus (FMDV), a positive sense, single-stranded RNA virus, causes a highly contagious disease in cloven-hoofed livestock. Like other picornaviruses, FMDV has a conserved 2C protein assigned to the superfamily 3 helicases a group of AAA+ ATPases that has a predicted N-terminal membrane-binding amphipathic helix attached to the main ATPase domain. In infected cells, 2C is involved in the formation of membrane vesicles, where it co-localizes with viral RNA replication complexes, but its precise role in virus replication has not been elucidated.

Insights into cleavage specificity from the crystal structure of foot-and-mouth disease virus 3C protease complexed with a peptide substrate.

Picornavirus replication is critically dependent on the correct processing of a polyprotein precursor by 3C protease(s) (3C(pro)) at multiple specific sites with related but non-identical sequences. To investigate the structural basis of its cleavage specificity, we performed the first crystallographic structural analysis of non-covalent complexes of a picornavirus 3C(pro) with peptide substrates. The X-ray crystal structure of the foot-and-mouth disease virus 3C(pro), mutated to replace the catalytic Cys by Ala and bound to a peptide (APAKQ|LLNFD) corresponding to the P5-P5' region of the VP1-2A cleavage junction in the viral polyprotein, was determined up to 2.

Control of choline oxidation in rat kidney mitochondria.

Choline is a quaternary amino cationic organic alcohol that is oxidized to betaine in liver and kidney mitochondria. Betaine acts as an intracellular organic osmolyte in the medulla of the kidney. Evidence is provided that kidney mitochondria have a choline transporter in their inner membrane.

Structural insights into the transcriptional and translational roles of Ebp1.

The ErbB3-binding protein 1 (Ebp1) is an important regulator of transcription, affecting eukaryotic cell growth, proliferation, differentiation and survival. Ebp1 can also affect translation and cooperates with the polypyrimidine tract-binding protein (PTB) to stimulate the activity of the internal ribosome entry site (IRES) of foot-and-mouth disease virus (FMDV). We report here the crystal structure of murine Ebp1 (p48 isoform), providing the first glimpse of the architecture of this versatile regulator.

Tumor necrosis factor and interleukin 1 decrease RXRalpha, PPARalpha, PPARgamma, LXRalpha, and the coactivators SRC-1, PGC-1alpha, and PGC-1beta in liver cells.

During the acute phase response, cytokines induce marked alterations in lipid metabolism including an increase in serum triglyceride levels and a decrease in hepatic fatty acid oxidation, in bile acid synthesis, and in high-density lipoprotein levels. Here we demonstrate that tumor necrosis factor (TNF) and interleukin 1 (IL-1), but not IL-6, decrease the expression of retinoid X receptor alpha (RXRalpha), peroxisome proliferator-activated receptor alpha (PPARalpha), PPARgamma, liver X receptor alpha (LXRalpha), and coactivators PPARgamma coactivator 1alpha (PGC-1alpha), PGC-1beta, and steroid receptor coactivator 1 (SRC-1) in Hep3B human hepatoma cells. In addition, treatment of mice with TNF and IL-1 also decreased RXRalpha, PPARalpha, PPARgamma, LXRalpha, and PGC-1alpha messenger RNA (mRNA) levels in the liver.