Fig. 5: hnRNP A2B1 interacts with SON, but not with RBFOX2, and facilitates target RNA binding of SON while inhibiting the RBFOX2-RNA interaction.

a Immunoprecipitation (IP) and Western blot analysis confirming the interaction of SON or RBFOX2 with indicated hnRNP proteins. U87MG cell lysates were immunoprecipitated with control IgG, SON antibody, or RBFOX2 antibody and analyzed by Western blotting with the indicated antibodies. Data are representative of n = 3 independent experiments. b The hnRNP components within the SON complex and the RBFOX2 complex identified from (a). c RT-qPCR analyses showing the effect of knockdown of three different hnRNPs (hnRNP A2B1, hnRNP K, and hnRNP M) on the expression of PTBP1 and PTBP2 in U87MG cells (n = 3). The expression levels of indicated hnRNPs were also measured to confirm the knockdown efficiency. d RT-qPCR analysis of SON, RBFOX2, hnRNP A2B1, and PTBP2 in U87MG cells expressing control or SON shRNA and infected with or without hnRNP A2B1 shRNA (n = 3). e The effects of SON knockdown and/or RBFOX2 knockdown on PTBP2 exon 10 inclusion were examined by RT-PCR. Data are representative of n = 3 independent experiments. f Cellular localization of SON (red) and hnRNPA2B1 (green) in U87MG cells were shown by immunostaining. Blue, DAPI. Scale bar: 20 μm. g, h The effects of hnRNP A2B1 knockdown on SON or RBFOX2 binding to the PTBP2 pre-mRNAs were measured by CLIP-qPCR. The locations of the qPCR primer sets indicated as 1–8 are shown in Fig. 4e. The qPCR experiments were performed three times. Error bars in all graphs represent the standard deviation (SD) of tests. NS not significant, *p < 0.05, **p < 0.01. Statistical significance was determined by an unpaired two-tailed t-test. Source data are provided in the Source data file.