The fact that reinitiation, albeit at reduced efficiency, can occur when the first AUG is moved 63 nt downstream raises the question of whether the reinitiating 40S subunits access this site by linear scanning from the TURBS. We examined this by exploiting the dependence of ribosomal scanning on eIF4G [34] . The BM2wt and BM2start +63 mRNAs were modified such that translation of the upstream M1 ORF was driven by the cricket paralysis virus (CrPV) IRES, which does not require eIF4G for function. These mRNAs (CrPV-BM2wt, CrPV-BM2start+63) were then translated alongside capped brome mosaic virus RNAs (BMV, Promega) and BM2wt RNAs (as a control for the efficacy of eIF4G depletion) in mock-and eIF4G-depleted rabbit reticulocyte lysates ( Figure 6C ). Translations were also performed in the absence or presence of dominant-negative eIF4A (eIF4A R362Q [35] ) to inhibit the activity of any residual eIF4G that had escaped depletion. As expected, the translation of the capped wildtype RNA (BM2wt) and BMV RNAs were inhibited in the eIF4Gdepleted RRL, but the CrPV IRES-containing mRNAs were translated efficiently in both control and depleted RRL ( Figure 6C ). Importantly, depletion of eIF4G had no effect on the reinitiation efficiency in either the CrPV-BM2wt or CrPV-BM2start +63 mRNAs, suggesting that the reinitiation site can be located in a scanning-independent manner.