Acute EIF4G1 depletion reveals transcript-wide determinants of initiation factor dependence and activates initiation-specific ribosome quality control

Canonical cap-dependent translation initiation requires the eIF4F complex, in which the scaffold protein EIF4G1 bridges cap-bound EIF4E, the RNA helicase EIF4A, and ribosome-associated EIF3. Using an inducible degron to acutely deplete EIF4G1 in human cells, we combined polysome profiling with mRNA-seq to quantify transcript-specific relative polysome association (RPA). EIF4G1 depletion causes heavy polysome collapse with reciprocal 80S monosome accumulation and activates initiation-specific ribosome quality control (iRQC), evidenced by RPS3 mono-ubiquitination enriched on 80S monosomes. Using RPA analysis we find the ribosome association of most transcripts is unaffected. However, we identify 1,250 mRNAs that are sensitive to EIF4G1 loss and 1,663 mRNAs that are translationally enhanced under the same conditions. As a class, the enhanced transcripts tend to be longer and have a higher GC content in the coding sequence relative to sensitive or unaffected transcripts. A synthetic reporter confirms that elevated CDS GC content can promote translation upon EIF4G1 depletion. These results suggest that the architecture of the entire transcript, beyond the 5'UTR, affects EIF4G1 usage and that removing EIF4G1 triggers iRQC consistent with an altered initiation architecture.