Fig 2: CMG2 is highly expressed in GC tissues and correlated with the outcome of patients. a Representative IHC images of CMG2. Scale bar = 100 µm. aa Absence of CMG2 expression in normal gastric mucosa. ab–d CMG2 staining was observed in GC tissues and the intensity was increased with the tumor invasion depth. ae CMG2 highly expressed in both primary tumor and the corresponding metastatic lymph node. b Higher percentage of CMG2+staining in GC tissues; ***P < 0.001. c, d CMG2 expression at protein and mRNA levels in four fresh surgical gastric tumor specimens (T) and paired adjacent normal tissues (N) detected by western blotting and qRT-PCR, respectively; **P < 0.01, *P < 0.05. e Data in the GEO database GES13911 showing higher CMG2 expression in GC tumor tissues compared to adjacent normal tissues; **P < 0.01. f Kaplan–Meier estimation indicating shorter overall survival time of patients with CMG2+ GC

Fig 3: Silencing CMG2 results in reduced invasive and metastatic properties of GC cells epithelial–mesenchymal transition (EMT). a Representative images of transwell invasion test showing decreased invasion capability of GC cells with knockdown of CMG2. b Quantification of the transwell invasion assay results; Scale bar = 100 µm; **P < 0.01. c Representative images of intraperitoneal metastasis tests showing reduced number of metastatic foci formed by shCMG2 cells. d Quantification of the intraperitoneal metastasis results; **P < 0.01. e Western blot showing upregulation of E-cadherin and downregulation of CD44 and vimentin in GC cells with knockdown of CMG2

Fig 4: Silencing CMG2 reduces GCSLC population. a Decreased expression of stemness marker CD44 and stemness-related factor SOX2 in GC cells with CMG2 knockdown (shCMG2). b Representative flow cytometric histograms showing decreased percentage of CD44High cells in shCMG2 GC cells. c Markedly reduced sphere formation by CMG2 knockdown GC cells; *P < 0.05. d Decreased colony formation by shCMG2 cells. e Representative images showing decreased tumor formation in nude mice by CMG2 knockdown GC cells. f Reduced weight of xenograft tumor formed by shCMG2 GC cells; **P < 0.01

Fig 5: CMG2High cells are detected in CD44High population in GC cell lines and in GC tissues. a Representative flow histogram of the percentage of CD44High cells in SGC7901 and XN0422 cell lines. b Limiting dilution showing increased capacity of sphere formation by CD44High cells compared to CD44Low cells; *P < 0.05. c Representative immunofluorescence images (left panel) and quantitative statistic results (right panel) showing more CMG2-positive cells detected in CD44High subpopulation than CD44Low subpopulation; Scale bar = 50 µm; **P < 0.01. d The co-localization of CMG2 and CD44 in the frozen sections of fresh GC tissues; Scale bar = 100 µm