A Temperature-Dependent Translation Defect Caused by Internal Ribosome Entry Site Mutation Attenuates Foot-and-Mouth Disease Virus: Implications for Rational Vaccine Design

Foot-and-mouth disease (FMD), which is caused by FMD virus (FMDV), remains a major plague among cloven-hoofed animals worldwide, and its outbreak often has disastrous socioeconomic consequences. A live-attenuated FMDV vaccine will greatly facilitate the global control and eradication of FMD, but a safe and effec-tive attenuated FMDV vaccine has not yet been successfully developed. Here, we found that the internal ribosome entry site (IRES) element in the viral genome is a critical virulence determinant of FMDV, and a nucleotide substitution of cytosine (C) for guanine (G) at position 351 of the IRES endows FMDV with temperature-sensitive and attenuation (ts & att) phenotypes. Furthermore, we demonstrated that the C351G mutation of IRES causes a temperature-dependent translation defect by impairing its binding to cellular pyrimidine tract-binding protein (PTB), resulting in the ts & att phe-notypes of FMDV. Natural hosts inoculated with viruses carrying the IRES C351G mu-tation showed no clinical signs, viremia, virus excretion, or viral transmission but still produced a potent neutralizing antibody response that provided complete protec-tion. Importantly, the IRES C351G mutation is a universal determinant of the ts & att phenotypes of different FMDV strains, and the C351G mutant was incapable of re-version to virulence during in vitro and in vivo passages. Collectively, our findings suggested that manipulation of the IRES, especially its C351G mutation, may serve as a feasible strategy to develop live-attenuated FMDV vaccines. IMPORTANCE The World Organization for Animal Health has called for global con-trol and eradication of foot-and-mouth disease (FMD), the most economically and socially devastating disease affecting animal husbandry worldwide. Live-attenuated vaccines are considered the most effective strategy for prevention, control, and eradication of infectious diseases due to their capacity to induce potent and long-lasting protective immunity. However, efforts to develop FMD virus (FMDV) live -attenuated vaccines have achieved only limited success. Here, by structure-function study of the FMDV internal ribosome entry site (IRES), we find that the C351 mu-tation of the IRES confers FMDV with an ideal temperature-sensitive attenuation phenotype by decreasing its interaction with cellular pyrimidine tract-binding protein (PTB) to cause IRES-mediated temperature-dependent translation defects. The temperature-sensitive attenuated strains generated by manipulation of the IRES address the challenges of FMDV attenuation differences among various livestock species and immunogenicity maintenance encountered previously, and this strategy can be applied to other viruses with an IRES to rationally design and develop live -attenuated vaccines.