Fig. 1

CircZNF800 expression is up-regulated in in CRC-derived spheroids and CRC tumor tissues. A Schematic illustration of circZNF800 biogenesis via backsplicing of exons 4 and 5 of the ZNF800 pre-mRNA (NC_000007.14). Two ZNF800 alternative transcripts carry either exon 6a (NM_176814.5) or 6b (XM_011515859.4) are shown. F and R indicate the forward and reverse primers used in sequencing of the backsplicing junction. B, C Stability analysis of circZNF800 in WiDr cells on RNase R (B) and actinomycin D (C) treatments. The relative RNA levels were determined by qRT-PCR. *P < 0.05 and **P < 0.01 relative to those of the untreated sample (B) or the ZNF800 transcript (C). D Morphology of the CRC-derived spheroids. E Up-regulated circZNF800 expression in spheroids derived from CRC cell lines. F Passage (P)-dependent up-regulated circZNF800 expression in CRC cell-derived spheroids and on serum-induced differentiation of the spheroids at passage 5 (Diff. P5). G, H Co-localization of circZNF800 and intestinal stem cell markers LGR5 and SOX9 in the adjacent normal tissue (G) or Ki-67 high proliferative marker (H) in a CRC tumor tissue in dual RNA FISH-immunofluorescent assays. Bars represent 100 µm. I Significant circZNF800 over-expression in paired tumor and normal tissues of CRC patients, and in late-stage CRC tumors. Paired tissue samples (n = 30) were subjected to qRT-PCR analysis to determine the circZNF800 and ZNF800 expression levels (left two panels); the circZNF800 expression data from the same patient cohort were further grouped according to pathological stages 1 & 2 (n = 17) and 3 & 4 (n = 13) (right two panels) for circZNF800 expression analysis. *p < 0.05, **p < 0.01 and ***P < 0.001 relative to the control samples as indicated or the adjacent normal tissues