The Double-Stranded RNA Binding Protein 76:NF45 Heterodimer Inhibits Translation Initiation at the Rhinovirus Type 2 Internal Ribosome Entry Site

Abstract

Eukaryotic mRNA translation regulation is most often achieved by interference with initiation events (10). Unimpeded initiation occurs upon assembly of eukaryotic initiation factor 4F at the cap, recruitment of the 43S preinitiation complex, scanning, formation of the 48S initiation complex at the initiation codon, and 60S ribosomal subunit joining. Translation initiation repressors binding to 5Ј and 3Ј untranslated regions (UTR) have been reported to interfere with distinct steps of the initiation cascade. These include obstructed eukaryotic initiation factor 4F cap assembly (26), inhibited stable association of the 40S ribosomal subunit (9, 12), and blocked 60S ribosomal subunit joining (31).Some mRNAs, however, can initiate translation by noncanonical mechanisms. Picornaviruses feature uncapped (29) plus-strand RNA genomes whose translation depends on internal ribosome entry sites (IRES) within their highly structured 5Ј UTR for initiation in a 5Ј-end, cap-independent manner (21, 34). Poliovirus (PV), a member of the family Picornaviridae, is responsible for paralytic poliomyelitis, resulting from specific targeting of spinal cord motor neurons in the human central nervous system. Neurological disease is the result of complex interactions determined by the host, e.g., aggravating susceptibility factors (44), viral receptors in the central nervous system (17), and the innate immune response (19), and the virus itself, e.g., capsid structure (43) and the viral RNA-dependent RNA polymerase (33; reviewed in reference 16).An important viral determinant of neuropathogenesis is the IRES. Single point mutations in IRES stem loop domain V of the PV (Sabin) oral vaccine strains have been implicated in neurovirulence attenuation (8,22,37), and deliberate manipulation of the PV IRES reduces its neurovirulent potential (1). Replacement with a heterologous IRES element of human rhinovirus type 2 (HRV2) eliminates PV propagation in cells of neuronal derivation (5, 13, 14) and abolishes neuropathogenicity in mice transgenic for the human PV receptor (13) and in nonhuman primates (14). Despite severely repressed viral propagation in neurons, the chimeric PV-RIPO replicates with wild-type kinetics in malignant glioma (15) and breast cancer (30) cells. These observations suggest that the conditions for HRV2 IRES-mediated translation in neurons differ from those in glioma and other cancer cells. HRV2 IRES-driven translation could require enabling factors in glioma cells that are absent in neuronal cells, or, alternatively, negative regulators in neurons could block PV-RIPO propagation.Neuroblast:glioma heterokaryon analyses revealed that neuron-specific repression is trans dominant over robust PV-RIPO propagation in glioma cells (27). We identified association of double-stranded RNA binding protein 76 (DRBP76), nuclear factor of activated T cells, 45 kDa (NF45), and RNA helicase A (RHA) with the HRV2 IRES by affinity chromatography with lysates from neuron-derived cells (27). These proteins have been reported to associate with e...

NF90 Exerts Antiviral Activity through Regulation of PKR Phosphorylation and Stress Granules in Infected Cells

“…NF90 was shown to inhibit replication of a broad range of viruses and to form complexes with NF45 and shuttle between the nucleus and cytoplasm (13)(14)(15)(16)(17)(18)(19). However, a detailed mechanism describing how NF90 negatively regulates virus replication remains unclear.…”

“…It suppresses the function of Ebola virus polymerases through interaction with VP35 (14), inhibits HIV replication through interaction with HIV-1 TAR RNA (15,16), represses internal ribosome entry sites in rhinoviruses (17,18) and negatively regulates influenza virus replication through interaction with viral nucleoprotein (NP) (19). However, other studies (20)(21)(22)(23) found that NF90 is required for the replication of some positivestranded RNA viruses and is important for expression of E6 protein in human papillomavirus-infected cells.…”

Picornavirus IRES elements: RNA structure and host protein interactions

“…For instance, the nuclear protein FBP2 is a KH protein that shuttles to the cytoplasm in infected cells negatively regulating EV71 IRES activity (Lin et al, 2009a). The double stranded RNA-binding protein DRBP76:NF45 is also a nuclear heterodimeric protein that interacts with HRV IRES repressing its activity in neuron-derived cells (Merrill and Gromeier, 2006). Similarly, the cellular mRNA decay protein AU-binding factor (AUF1) behaves as a negative regulator of EV71 and HRV infections .…”

Characterization of proteins associating with 5’ terminus of PGHS-1 mRNA

“…Nevertheless, NF45 and NF90 were shown to bind to dsRNA as well as to proteins [26,39]. In neuronal cells, the NF45/DRBP76 heterodimer complex can inhibit translational initiation by binding to the 5'UTR of human rhinovirus type 2 that encoding the internal ribosome entry site (IRES) [40]. NF90 has been shown to increase the half-life of IL-2 mRNA following T-cell activation [41].…”