Fig. 3

Regulation of miR-106a-5p packaging into exosomes by hnRNPA1. (A) The potential RBPs that may bind to miR-106a-5p. (B) The specific interaction between the miR-106a-5p sequence and hnRNPA1 motifs. C-D. After transfecting HCT 116 and SW620 cells with sh-hnRNPA1, the expression of hnRNPA1 was detected by qRT-PCR and western blot (C), and the expression of miR-106a-5p was detected by qRT-PCR (D). E-G. Mφ cells were co-cultured with SW620/HCT 116 pre-transfected with sh-hnRNPA1 and Cy3-miR-106a-5p (red). Immunofluorescence was performed to detect the red fluorescent signals in macrophages. Scale bar = 10 μm. H-I. Mφ cells were incubated with exosomes derived from SW620/HCT 116 transfected with sh-hnRNPA1 and Cy3-miR-106a-5p (red). Immunofluorescence was performed to detect the red fluorescent signals in macrophages. Scale bar = 10 μm. J. Western blot was employed to assess hnRNPA1 expression in samples obtained from miRNA pulldowns, utilizing nuclear, cytoplasmic, or exosomal lysates from HCT116 cells. K. RIP assay was executed using an anti-hnRNPA1 antibody (or IgG as a control) on lysates derived from HCT 116 cells or exosomes. qRT-PCR was used to quantify miR-106a-5p levels in the immunoprecipitated samples, expressed as percentages relative to the input (% input). The data presented herein represent the outcomes of a minimum of three independent experiments and are depicted as the mean ± standard deviation (SD). ns = No significant difference, ** P < 0.01, *** P < 0.001