Fig. 2

CircLOC101928570 acted as a sponge for miR-150-5p. A ceRNAs that could competitively bind to this exonic circRNA. MiR-150 has two targets marked with red lines. B The relative levels of 10 miRNA candidates in the PBMC lysates were detected by real-time PCR. miR-150-3p/5p was pulled down by circLOC101928570 in PBMCs. C The interaction of circLOC101928570 with miR-150 was tested by RNA immunoprecipitation (RIP) of AGO2 from HEK293T cells. Relative circLOC101928570 enrichment levels were quantified by RIP-qPCR (AGO2 RIP/IgG RIP). D Schematic illustration of wild-type or mutant transcripts of the circLOC101928570 luciferase reporter. E, F Luciferase reporter and effects of miR-150-3p/5p on the expression of wild-type or mutant transcripts of the circLOC101928570 luciferase reporter. Luciferase activity was normalized to the value obtained in cells transfected with NC oligonucleotides. G The expression of miR-150-5p was analyzed by qRT–PCR in cells transfected with circLOC101928570, mock vector, shRNA-circLOC101928570 or shRNA-NC. H The expression levels of circLOC101928570 were determined with qRT–PCR in cells transfected with miR-150-3p/5p mimics or inhibitor. I The colocalization of circLOC101928570 and miR-150-5p in PBMCs was detected using a FISH assay. CircLOC101928570 was captured with a Cy3-labeled probe (red), while miR-150-5p was captured with a digoxin-labeled probe followed by further visualization using an anti-digoxin rhodamine-conjugated antibody (green). Blue: stained with DAPI. Scale bar: 10 µm. IgG: immunoglobulin G. All data are presented as the means ± SD (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001