Poliovirus Switches to an eIF2-Independent Mode of Translation during Infection

Abstract

Inhibition of translation is an integral component of the innate antiviral response and is largely accomplished via interferon-activated phosphorylation of the ␣ subunit of eukaryotic initiation factor 2 (eIF2␣). To successfully infect a host, a virus must overcome this blockage by either controlling eIF2␣ phosphorylation or by utilizing a noncanonical mode of translation initiation. Here we show that enterovirus RNA is sensitive to translation inhibition resulting from eIF2␣ phosphorylation, but it becomes resistant as infection progresses. Further, we show that the cleavage of initiation factor eIF5B during enteroviral infection, along with the viral internal ribosome entry site, plays a role in mediating viral translation under conditions that are nonpermissive for host cell translation. Together, these results provide a mechanism by which enteroviruses evade the antiviral response and provide insight into a noncanonical mechanism of translation initiation.Initiation of cap-dependent translation occurs in three initial steps. The canonical first step is recruitment of an eIF2-GTP-Met-tRNA iMet ternary complex to the 40S subunit in a binding step stabilized by initiation factors eIF3 (eukaryotic initiation factor 3), eIF1, and eIF1A to form a 43S complex. The 43S complex is competent to bind mRNA complexed with eIF4F and eIF4B bound to the 5Ј cap structure to form 48S initiation complexes that are capable of scanning for initiator codons, aided by eIF4A/eIF4B RNA helicase functions (20). Stable 48S complexes arise with formation of codon-anticodon base pairing in the P site of the 40S ribosome subunit. Proper placement of the tRNA anticodon is mediated by eIF2, which is aided by eIF1 and perhaps eIF5B in rejection of poor-context triplets (20, 28). Upon proper codon-anticodon base pairing, eIF2 hydrolyzes its bound GTP, followed by eIF5B-mediated ejection of factors from the 40S subunit and 60S subunit joining to form the complete 80S ribosome (20,42).Before 60S subunit joining, hydrolyzed eIF2-GDP is released from the 48S complex and must be recycled by interaction with the guanine nucleotide exchange factor eIF2B to exchange the GDP for GTP for use in the next round of translation initiation. Exposure to several types of stress stimuli restricts translation through the activation of eukaryotic initiation factor 2 (eIF2␣) kinases PKR (protein kinase R), PERK (double-stranded RNA-activated ER kinase), HRI (heme-regulated inhibitor), and GCN2 (general control nonderepressing 2). Phosphorylation of eIF2␣ turns eIF2 into a competitive inhibitor of eIF2B function through an increased affinity of phospho-eIF2 for eIF2B, causing the accumulation of eIF2-GDP and a corresponding global inhibition of translation initiation (25, 51).Infection of mammalian cells with viruses often induces an antiviral response that can produce interferon and activate the eIF2␣ kinase PKR, causing a suppression of translation by eIF2␣ phosphorylation (34). However, poliovirus (PV), a member of the Picornaviridae family and Enterovirus gen...

Upregulation of eIF5B controls cell-cycle arrest and specific developmental stages

“…In serum-starved cells, the increased levels of eIF5B lead to increased formation of eIF5B complexes with tRNAMet i , consistent with its previously described role with viral and stress mRNAs (12,(17)(18)(19). Additionally, increased phosphorylation of eIF2α is observed in these three conditions.…”

“…Translation is regulated and active in X. laevis oocytes after maturation (3,4); however, a subset of mRNAs is translated in immature oocytes (3,5), indicating that uncharacterized factors support translation in these conditions. eIF5B is known to recruit 60S ribosome subunits (2,11,13) and to support the translation of viral, stress-related, and other specific mRNAs in embryos and in stress conditions in which translation is regulated (12,(17)(18)(19)(20). Therefore, we compared eIF5B levels in folliculated immature, defolliculated, and mature oocytes (5).…”

“…in the case of specific viral and stress-related mRNA translation (2,(11)(12)(13)(17)(18)(19). We, therefore, tested whether increased eIF5B expression leads to correspondingly increased complexes with tRNAMet i in THP1 cells that had been serum-starved for 1 d. eIF5B immunoprecipitation (Fig.…”

“…eIF5B functions in 60S ribosome subunit joining during canonical translation (2,13,14) and is involved in pre-40S ribosome subunit proofing (15,16). eIF5B is also required for the translation of a few viral and specialized mRNAs (12,(17)(18)(19)(20) and can contribute to supporting or stabilizing tRNA-Met i association, including in specific conditions where phosphorylation of the eIF2 subunit, eIF2α, is also observed (12,(17)(18)(19). In some cases, phosphorylation of eIF2α is sufficiently increased relative to the levels of its guanine nucleotide exchange factor (eIF2B), which leads to decreased release of phosphorylated eIF2α from eIF2B.…”

“…Since it has been observed that eIF5B is important for translation of viral and stress-related mRNAs in conditions that also exhibit increased eIF2α phosphorylation (12,(17)(18)(19), we examined eIF2α (2, 21) in folliculated immature, defolliculated, and mature oocytes (5). We found that eIF2α phosphorylation is increased in late immature oocytes stages V and VI that precede maturation (Western blot, Fig.…”