Fig. 5: YBX3 recruits PABPC1 to regulate ICAM-1 translation.

A Scatter plot of proteins bound to YBX3 or IgG in HK-1-EBV cells. X-axis presents the peptide number bound to IgG, while y-axis shows the peptide number bound to YBX3. PSM, the peptide-spectrum matches. YBX3 and its protein partner PABPC1 are highlighted in red. B, C Co-immunoprecipitation (Co-IP) assays showing the interaction between YBX3 and PABPC1 in 293T cells. Cells were transfected with Flag-YBX3 and/or HA-PABPC1 plasmids as indicated. The immunoprecipitation efficiency and enriched FLAG-YBX3 or HA-PABPC1 were detected by immunoblotting with antibodies against FLAG and HA, respectively. Schematic diagram showing domain structures of wild-type and mutant constructs of YBX3-FLAG (D) and HA-PABPC1 (F). Co-IP assays in 293T cells co-transfected with either wild-type or mutant combination of HA-PABPC1 and FLAG-YBX3. Immunoprecipitations were performed with anti-FLAG (E) anti-HA (G) antibodies. IP, immunoprecipitation. H Transcriptional correlation between YBX3 and PABPC1 in NPC samples (n = 95). Correlation coefficient and P value were calculated using the ‘cor. test’ function in R. I RIP-qPCR assay detecting PABPC1’s binding level to ICAM-1 in HK-1-EBV and CNE2-EBV cells with NSUN2 (siNSUN2) or YBX3 (siYBX3) knockdown, compared to control cells (siNC). IgG-IP was used as a control. Data are the mean ± SD. J Western blot analysis evaluating ICAM-1 protein expression in HK-1-EBV and CNE2-EBV cells with PABPC1 knockdown by siRNA and concurrently treated with puromycin. Representative western blot images of three biological replicates with similar results are shown in (B, C, E, G, J). Data in (I) are Mean ± SD from three independent experiments. Statistical analyses were performed using appropriate tests based on data distribution: correlation analyses for (H) were conducted using the cor. test function in R; and one-way ANOVA test was used for (I). *P < 0.05, ** P < 0.01.