Fig. 5. The recruitment of RQT complex into the colliding ribosome in the ubiquitin decoding-dependent manner.

a Schematic of the experiments. b Binding assay between the colliding ribosome and the RQT complex. The ubiquitinated or nonubiquitinated colliding ribosomes were prepared by HEL2-containing or hel2-knockout (hel2∆) IVT reaction and mixed with the indicated RQT complex in the presence of AMPPMP as described in (a). After the binding reaction, free and bound RQT complexes were separated using a sucrose cushion. The RQT factors and uS10 in each fraction were detected by immunoblotting using anti-Flag and anti-HA antibodies, respectively. c Model of the recognition of colliding ribosomes by the RQT complex. All experiments were performed at least twice with highly reproducible results.