circFGD4 was down-regulated in GC and associated with the progression and poor prognosis of GC patients
(A) Schematic illustration exhibiting the overlapping of circRNAs predicted by GSE100170 and GSE131414 and our circRNA microarray (CapitalBio Technology Co., Ltd). (B) Clustered heat map displaying the tissue-specific circRNAs in five paired human GC tissues and noncancerous tissues. GC tissues: S1; Paired noncancerous tissues: S2. (C) Relative expression level of circFGD4 was detected by qRT-PCR in 51 paired GC tissues and noncancerous tissues. (D) Relative expression level of circFGD4 was examined by qRT-PCR in six GC cell lines and GES-1 cells. (E) The expression of circFGD4 and FGD4 RNA was validated by qRT-PCR after treatment with RNase R in SGC7901 cells. (F) The expression of circFGD4 and FGD4 RNA was determined by qRT-PCR after treatment with Actinomycin D at the indicated time points in SGC7901 cells. (G) qRT-PCR validated the existence of circFGD4 in BGC823 and SGC7901 cell lines. circFGD4 was amplified by divergent primers in cDNA but not gDNA. (H) Schematic representation of circFGD4 formation. The back-splice junction sequence of circFGD4 was validated by Sanger sequencing. (I) Kaplan–Meier plotter analysis of the correlation of circFGD4 expression level with the overall survival of GC patients. (J) TCGA database analysis of FGD4 mRNA levels in human GC normal tissues (n=211) and tumour tissues (n=408). (K) Violin plot of relative abundance of FGD4 mRNA levels in different stages of GC tissues from the TCGA database. The white dot represents the median. (L) Correlation analysis of circFGD4 levels with FGD4 mRNA levels in 30 GC tissues. (M) Confocal FISH was performed to observe the cellular location of circFGD4 in MGC803 and AGS cells (values are shown as the mean ± standard error of the mean based on three independent experiments; *P<0.05, **P<0.01).