The p48 isoform of the PA2G4/EBP1/ITAF45 oncoprotein is required for the encephalomyocarditis virus IRES-driven translation initiation

Internal ribosome entry sites (IRESs) enable cap-independent initiation of picornaviral RNA translation and, together with canonical translation initiation factors, typically require specific cellular proteins known as IRES trans-acting factors (ITAFs). While the type II IRES of foot-and-mouth disease virus (FMDV, an aphthovirus) has been shown to depend on the oncoprotein ITAF45, also known as Proliferation-Associated 2G4 (PA2G4) or ErbB-3 receptor Binding Protein (EBP1), for in vitro assembly of the 48S pre-initiation complex, some related type II IRESs, such as that of encephalomyocarditis virus (EMCV, a cardiovirus), can form the initiation complex independently of ITAF45. In this study, we performed a genome-wide CRISPR screen and identified knockouts of PA2G4/EBP1/ITAF45 in cells that survive EMCV infection, suggesting an important role for this factor. We show that the p48 isoform of ITAF45, but not the p42 isoform, is crucial for efficient EMCV/Mengovirus replication and for propagation of replicons in human cell culture. Loss of ITAF45 markedly diminishes EMCV and FMDV IRES activities, which can be rescued by re-expression of ITAF45-p48. Interestingly, cell-free translation assays reveal that EMCV IRES activity is less ITAF45-dependent in vitro, in contrast to FMDV, raising questions about the versatile functions of ITAFs in IRES-driven translation. These findings reveal an isoform-specific function of ITAF45 in supporting cardiovirus infection and provide new insights into the complex regulation of IRES-driven translation, with implications for developing targeted antiviral strategies.