Fig. 4

Interaction and co-localization of circHAS2 with USP10. A The enrichment of circHAS2-MS2 complex formation was detected with MS2-CP-Flag in SW620 cells. B The Silver staining of circHAS2-associated proteins by the biotin-labeled probe. C The RNA pull-down confirmed the interaction between circHAS2 and USP10. The relative expression of circHAS2 was determined using the RIP assay with AGO2 and IgG antibodies in SW620 cells. D The RIP assays were conducted under the specified treatment conditions using anti-USP10 and anti-IgG antibodies in SW620 cells. E Representative images of the FISH assay in SW620 cells revealed the colocalization of circHAS2 and USP10 in the cytoplasm with the target probe labeled with Cy3 and nuclei stained with DAPI (scale bar: 20 μm). F The colocalization of circHAS2 and USP10 in CRC tissues and paired normal tissues of microarrays were verified by FISH (scale bar: 400 μm; scale bar: 100 μm). G The interaction profile between circHAS2 fragments and USP10 was predicted by the catRAPID database. H The interaction between circHAS2 full-length and truncations with USP10 was validated using RIP assays in HEK-293T cells. I The heat map illustrated the interaction between circHAS2 and the N-terminal region (1-100 amino acids) of USP10 by the catRAPID database. Statistical significance in two-group experiments was assessed using a two-sided Student’s t-test, and multiple-group comparisons were conducted through one-way analysis of variance. Data are represented as mean ± SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, no significance