Viral tRNA-like structure hijacks host ribosomes for poly(A)-independent translation

Positive-sense RNA viruses often use 3' tRNA-like structures (TLSs) instead of poly(A) tails to capture the host translation machinery. While TLSs resemble canonical tRNAs and engage host factors, their ability to directly recruit ribosomes has remained unresolved. Here, we present cryo-electron microscopy snapshots of the histidine-accepting TLS (TLSHis) from tobacco mosaic virus bound to the 60S subunit, the 80S ribosome, and the 80S-tRNAiMet initiation complex. Across these states, TLSHis is consistently anchored to the L1 stalk of 60S, even under cycloheximide treatment, yet remains dynamic on the 40S subunit. Structural analysis shows that TLSHis transitions from the E-site to an adjacent Z-site to accommodate initiator tRNA. Functional assays show that TLSHis preferentially associates with ribosomal subunits over polysomes and can substitute for a poly(A) tail to promote robust cap-dependent translation. Together, these findings reveal how a viral tRNA mimic hijacks host ribosomes to promote poly(A)-independent translation.