Abstract
Poliovirus IRES-mediated translation requires the functions of certain canonical as well as non-canonical factors for the recruitment of ribosomes to the viral RNA. The interaction of cellular proteins PCBP2 and SRp20 in extracts from poliovirus-infected cells has been previously described, and these two proteins were shown to function synergistically in viral translation. To further define the mechanism of ribosome recruitment for the initiation of poliovirus IRES-dependent translation, we focused on the role of the interaction between cellular proteins PCBP2 and SRp20. Work described here demonstrates that SRp20 dramatically re-localizes from the nucleus to the cytoplasm of poliovirus-infected neuroblastoma cells during the course of infection. Importantly, SRp20 partially co-localizes with PCBP2 in the cytoplasm of infected cells, corroborating our previous in vitro interaction data. In addition, the data presented implicate the presence of these two proteins in viral translation initiation complexes. We show that in extracts from poliovirus-infected cells, SRp20 is associated with PCBP2 bound to poliovirus RNA, indicating that this interaction occurs on the viral RNA. Finally, we generated a mutated version of SRp20 lacking the RNA recognition motif (SRp20ΔRRM) and found that this protein is localized similar to the full length SRp20, and also partially co-localizes with PCBP2 during poliovirus infection. Expression of this mutated version of SRp20 results in a ∼100 fold decrease in virus yield for poliovirus when compared to expression of wild type SRp20, possibly via a dominant negative effect. Taken together, these results are consistent with a model in which SRp20 interacts with PCBP2 bound to the viral RNA, and this interaction functions to recruit ribosomes to the viral RNA in a direct or indirect manner, with the participation of additional protein-protein or protein-RNA interactions.
Highlights
Translation of eukaryotic mRNAs most often occurs via a capdependent mechanism of initiation
Since SRp20 is important for poliovirus infection, which occurs in the cytoplasm of the cell, we predicted that we would observe a greater amount of SRp20 in the cytoplasm of poliovirus-infected cells compared to mock-infected cells
We conclude that expression of SRp20DRRM significantly reduced the levels of poliovirus produced, and that this protein may function as a dominant-negative in the cell. Overall these results provide important new insights into the localization and functions of IRES trans-activating factors (ITAFs) during poliovirus infection
Summary
Translation of eukaryotic mRNAs most often occurs via a capdependent mechanism of initiation (see Figure 1A). Cellular mRNAs contain a 7-methyl guanosine cap at their 59 ends, and this cap structure is recognized by the eukaryotic initiation factor 4F (eIF4F) cap binding complex. The eIF4F complex consists of the initiation factors 4A, 4G, and 4E and recruits the ribosome to the mRNA for translation initiation. The 40S ribosomal subunit binds a protein complex that consists of eIF1, eIF2-GTP-MettRNA (i.e., the ternary complex), eIF3, and eIF5. The assembled 43S pre-initiation complex binds the mRNA at the cap structure via interaction of a central domain of eIF4G with eIF3. The bound pre-initiation complex scans along the RNA until an AUG start codon is recognized in a favorable context [1], at which point GTP is hydrolyzed to GDP in the presence of eIF5. Changes to the cellular environment, which can occur during viral infection or under various conditions of stress, can result in a down-regulation of capdependent translation often by interfering with initiation factors that play important roles in cap-dependent translation initiation
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
