Fig. 4

The effects of circZFR and HNRNPLL on alternative splicing of MYO1B. a GSEA results of the differential genes affected by HNRNPLL knockdown. b, c Distribution of alternative splicing events affected by circZFR (b) and HNRNPLL (c). SE skipped exon, RI retained intron, A5SS alternative 5’ splice site, A3SS alternative 3’ splice site, MXE mutually exclusive exons. d Venn diagram illustrating the overlap of altered exon skipping events upon circZFR overexpression or HNRNPLL depleting. P value was calculated by χ2 test. e HNRNPLL binding site analyses on alternatively spliced exons. f Pie chart showing the screening of potential functional target of both circZFR and HNRNPLL.g Kaplan-Meier analysis of the DFI of the LUAD patients. h Top 6 hallmark pathways from GSEA enriched in patients with high MYO1B PSI scores from TCGA project. i Left: Diagram of the splicing variants of MYO1B mRNA and the primers for RT-PCR detection of exon 23 (primer 1) and exons 23 and 24 (primer 2) inclusion/exclusion. Right: RT-PCR validation of MYO1B exons upon knockdown or overexpression of circZFR in A549 cells. in%, percentage of inclusion. Red, exon 23, blue, exon 24, purple, exon 23/24. j The fluorescence images of circZFR (green) and MYO1B exon23 (red). Nuclei was stained with DAPI (blue). Scale bars, 50 μm. k Splicing pattern of MYO1B in LUAD cell lines (upper panel) and tissues (lower panel) as detected by RT-PCR. T, tumor tissue, N, nontumorous tissue. l Correlation analysis on circZFR and MYO1B PSI in LUAD tissues. m The effects of HNRNPLL on MYO1B exons in A549 cells. n Interaction of MYO1B exons and HNRNPLL examined by RT-PCR with RIP assays in A549 cells. Data are shown as mean ± SD (n = 3). Similar results were obtained in three independent experiments or typical photographs of one representative experiment. *p < 0.05, **p < 0.01, ***p < 0.001, two-tailed Student’s t test