Abstract
CircCRIM1 (hsa_circ_0002346) is a circular RNA derived from gene CRIM1 (the cysteine rich transmembrane BMP regulator 1 circRNAs) by back-splicing. Recent studies have suggested the diverse function of CircCRIM1 in the tumorigenesis of multiple malignancies, including osteosarcoma (OS). Here, we investigated the role and mechanism of circCRIM1 during OS progression. Differentially expressed circRNAs (including circCRIM1) in OS and human osteoblast (hFOB1.19) cell lines were selected by searching the circRNA expression microarray dataset of GSE96964. The expression levels of circCRIM1 and its sponging miRNAs and target genes were examined by RT-qPCR. The effects of circCRIM1 on the proliferation, migration, and invasion of OS cells were investigated by in vitro gain of function experiments. The in vivo function of circCRIM1 on OS was evaluated by measuring the subcutaneous and in situ tumor growth in nude mice. In addition, dual-luciferase reporter assay and in situ hybridization (FISH) were performed to explore the underlying mechanisms of circCRIM1 and its sponging miRNAs and target genes in OS. CircCRIM1 is downregulated in human OS cell lines and predominantly presents in the cytoplasm as demonstrated by RT-qPCR and FISH assays. Overexpression of circCRIM1 suppressed the migration, invasion, proliferation of OS cells in vitro and OS tumor growth in vivo. Mechanistically, we identified miR146a-5p as a sponge miRNA of circCRIM1 through bioinformatic prediction and confirmed their interaction and colocalization via reporter gene assay and FISH analysis. This interaction leads to increase expression of the downstream target gene NUMB, which will cause inhibition of the Notch signal pathway. We further demonstrated that miR146a-5p overexpression could reverse the antitumor effect induced by circCRIM1 in OS cells. Our results support that circCRIM1 acts as a tumor suppressor in OS by sponging miR146a-5p and its downstream target NUMB.