Fig. 5: RPL24C serves as the substrate for the OPNR complex.

A, B: AlphaFold predicted structures and the interaction between the Arabidopsis OPNR-OAP1-OAP2 trimer and RPL24C (A), and the human C1ORF109-CINP dimer and RSL24D1 (B). The contact sites and conserved sequences are also shown. C Recombinant protein pull-down assay results for GST-RPL24C and the His-OAP1-OPNR-OAP2 trimer. Free GST proteins served as the control. 5% input used for the His-OAP1-OPNR-OAP2 trimer and 100% input used for the GST-RPL24C and GST. D CSLM images show the localization of RPL24C-YFP before (upper panel) and after (lower panel) inducible CRISPR/Cas9-facilitated mutation of OPNR (icr-opnr). The same settings of YFP and mScarlet channels were used for upper and lower panels. The OPNR-mScarlet signals almost disappeared after 3 days of inducing in the lower panel. E, G Bar charts show measurements of the RPL24C-YFP (E) and YFP-NOG1-1 (G) signals in the cytosol before and after CRISPR/Cas9 induced mutation of OPNR, CDC48D and CIP111. Data represent mean of five measured cells ± SD. The P values were generated from a two-tailed Student’s t-tests. F, H Localization of YFP-NOG1-1 (F) and BUD20-YFP (H) before (upper panel) and after (lower panel) CRISPR/Cas9-induced mutation of OPNR (icr-opnr), respectively. I, J Proteomic analyses of the 60S fractions from mutant and wild-type plants. Volcano plot results for icr-oap1 vs CK (I) and icr-cdc48D vs CK (J) with -log10 P-value on the y-axis and log2 intensity differences on the x-axis. P-values were calculated using two-tailed Student’s t-test, moderated by Benjamini–Hochberg’s method. FDR = 0.05, s0 = 0.5. The intensity of each protein was normalized to the total signal of each sample. Each dot represents a protein with the green dots representing proteins that are part of large ribosome subunits and blue dots representing small ribosome subunits. RPL24C, NOG1-1 and BUD20 are shown in red. The full lists are shown in Supplementary Data 6. K Sucrose density gradient centrifugation obtained ribosome profiles for wild-type, icr-oap2, icr-cip111 and icr-cdc48D calli after CRISPR/Cas9-induced mutation. Scar bars: 5 µm.