Fig. 2: Quercetin upregulates the translation of PANX1 mRNA in RMS cells.

Rh18 (eRMS) and Rh30 (aRMS) cells were treated with increasing dosages of quercetin or its vehicle control (DMSO) for 24 h. DMSO concentration corresponded to the highest dosage of quercetin. Representative Western blots and quantification for PANX1 in Rh18 (n = 3) (A) and Rh30 (n = 3) (B) cells are shown. Statistical analysis was performed using a one-way ANOVA with Tukey’s post-hoc test. *P < 0.05 and **P < 0.01 compared to DMSO. GAPDH was used as loading control. Rh18 and Rh30 cells were treated with 50 μM and 10 μM, respectively, for 24 h and subjected to RT-qPCR or polysome fractionation. C RT-qPCR analysis of PANX1 transcript levels in Rh18 (n = 3) and Rh30 (n = 3) using primers specific to its exon 1 region. Results are relative to GAPDH transcript levels. ns: not significant. qPCR program: 95 °C for 3 min followed by 40 cycles of 95 °C for 15 s, 60 °C for 30 s. Statistical analysis was performed using a paired Student’s t test. ns not significant. D Representative polysome traces of Rh30 cells treated with quercetin or DMSO. RT-qPCR analysis (n = 4) showing (E) distribution of PANX1 mRNA levels in polysome fractions and (F) their proportions in subpolysomes (fractions 1–4) and polysomes (fractions 5–8) fractions from DMSO- or quercetin-treated Rh30 cells. Results are expressed as the percentage of total PANX1 mRNA. Statistical analysis was performed using a paired Student’s t test. *P < 0.05. RT-qPCR analysis (n = 4) showing (G) distribution of GAPDH mRNA levels in polysome fractions and (H) their proportions in combined subpolysomes (fractions 1–4) (translationally inactive) and polysomes (fractions 5–8) (translationally active) fractions from the DMSO- or quercetin-treated Rh30 cells in (E) and (F). Results are expressed as the percentage of total GAPDH mRNA. Statistical analysis was performed using a paired Student’s t test. ns not significant.