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Fig. 1 | Genome Biology

Fig. 1

From: A novel FLI1 exonic circular RNA promotes metastasis in breast cancer by coordinately regulating TET1 and DNMT1

Fig. 1

Identification of FLI1 circular RNA by Cas9IP. a Overexpression of FLI1 in breast cancer tissues. FLI1 expression was quantitated by immunohistochemical staining and was evaluated as the expression score. **p < 0.01 in breast carcinoma tissues as compared with their adjacent tissues. b High expression of FLI1 in carcinoma as compared with adjacent tissues. Red arrow: dark brown immunohistochemical staining of FLI1 oncoprotein. c CRISPR Cas9-guided chromatin immunoprecipitation (CasIP). Cas9, CRISPR Cas9; FLI1 gRNA, Cas9 guiding RNAs that target the FLI1 promoter. Cas9 binds to the FLI1 promoter through a mechanism of base pairing between the gRNA and target DNA. After fixation, the Cas9-FLI1 promoter chromatin complex was immunoprecipitated by an anti-Cas9 antibody. The CasIP-captured RNAs were sequenced to identify the RNA components that regulate the gene activity in breast cancers. d Identification of FLI1 circular RNA by CasIP. In the FLI1 Cas9-gRNA cassette vector, two Cas9 gRNAs are transcribed by U6 and H1 promoters, respectively, and guide Cas9 to the FLI1 promoter. The CasIP sequencing identifies a novel FLI1 exonic circular RNA that interacts with the gene promoter. e Enrichment of FECR1 in the FLI1 promoter. After CasIP, the captured RNAs were reverse transcribed to quantitate the abundance of FECR1 in the Cas9-captured promoter complex. M, 100 bp marker; IgG, ChIP with antibody control; Cas9, ChIP with anti-Cas9 antibody; nAb, the ChIP negative control, in which the anti dCas9-FLAG antibody was replaced by the equal amount of albumin protein

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