Fig 2: SLC34A2 promotes the stemness of neuroblastoma cells. (A) Representative image of spheroids formed by SH‐SY5Y cells. Scale bar, 100 μm. (B,C) SLC34A2 expression was detected in SH‐SY5Y cells and spheroids. (D,E) Expression of stemness markers (ALDH1 and Nanog) was examined in SH‐SY5Y cells and spheroids. (F,G) Expression of stemness markers was determined in SH‐SY5Y cells with or without SLC34A2 overexpression. (H) The capacity of spheroid formation was evaluated in cells depicted in (F). Scale bar, 100 μm. (I,J) Expression of stemness markers was measured in SH‐SY5Y cell spheroids with or without SLC34A2 knockdown. (K) The capacity of spheroid formation was detected in spheroids described in (I). (L,M) The overexpression and knockdown efficiency of SLC34A2 was confirmed in SH‐SY5Y cells and spheroids, respectively. The difference was assayed using one‐way ANOVA with the Tukey–Kramer post hoc test. Data are presented as mean ± SD; n ≥ 3, **P < 0.01 vs control.

Fig 3: SLC34A2 attenuates the stemness of neuroblastoma cells through miR‐25–Gsk3β axis. (A) MiR‐25 and Gsk3β expression was detected in SH‐SY5Y cells with SLC34A2 overexpression plus miR‐25 knockdown or Gsk3β overexpression. (B,C) Expressions of stemness markers were measured in cells depicted in (A). (D,E) The capacity of spheroid formation was determined in cells described in (A) by detecting the spheroid size (D) and number (E). Scale bar, 100 μm. (F,G) Expression of Wnt3a and β‐catenin was examined in cells depicted in (A). The difference was assayed using one‐way ANOVA with the Tukey–Kramer post hoc test. Data are presented as mean ± SD; n ≥ 3, *P < 0.05, **P < 0.01 vs control.

Fig 4: SLC34A2 expression is negatively correlated with the overall survival and event‐free survival of neuroblastoma patients. (A–F) KM‐Plotter analysis showed the correlation between SLC34A2 expression and overall survival (A–C) and relapse‐free survival (D–F) of neuroblastoma patients.

Fig 5: Solute carrier family 34 member 2 (SLC34A2) promotes esophageal squamous cell carcinoma (ESCC) cell proliferation by inducing autophagy. (a) The proliferation‐associated signaling pathways among the enriched pathways were shown by gene set enrichment analysis (GSEA) based on RNA‐sequencing data of SLC34A2‐overexpressed cells. (b, c) Gene sets of autophagosome and autophagosome_membrane were positively correlated with high SLC34A2 expression in ESCC by GSEA. (d) Protein of LC3B and P62 were examined by western blot (WB) when SLC34A2‐upregulated or SLC34A2‐downregulated cells were cultured in Earle's balanced salt solution (EBSS) for 2 h. (e) SLC34A2‐overexpressed/−downregulated cells were transfected with mRFP‐GFP‐LC3B lentiviruses and cultured for 48 h. After 2 h of EBSS treatment, green and red fluorescence were detected under laser confocal microscope. Scale bar, 5 μm. The number of yellow and red dots per cell were counted in 20 cells of each group. (f) Electron microscopy was conducted to visualize the ultrastructure of the ESCC cells after SLC34A2 overexpression or knockdown. Red arrows represent autophagic vacuoles (AVs). Magnification scale bar, 2 μm (upper) and 500 nm (lower). (g–j) Cell counting‐kit 8 (CCK‐8) and 5‐ethynyl‐2′‐deoxyuridine (EdU) assays were carried out to detect the proliferative ability of ESCC cells by treatment with BafA1 or RAPA. The grayscale value of the target protein in the WB was quantified using ImageJ software and normalized to glyceraldehyde 3‐phosphate dehydrogenase (GAPDH). Scale bar, 20 μm. *p < 0.05; **p < 0.01; ***p < 0.001.