Fig. 6

SPON2 was a target of SOX13; SPON2 knockdown suppressed the GEC angiogenesis in U87 glioma. (a) A schematic depiction of the predicted binding sites for SOX13 on the promoter fragment of SPON2 and the site mutagenesis design for the reporter assay. Promoter activities were measured to determine responsive SOX13-binding sites in the promoter of SPON2 using luciferase reporter assays. Data are presented as the means ± SD (n = 3, each group). ^**P < 0.01 vs. SPON2-Mut4 group. (b) A schematic representation of the SPON2 promoter region 3000 bp upstream or 200 bp downstream of the TSS, designated as + 1. ChIP PCR products for the putative SOX13 binding sites and an upstream region not expected to associate with SOX13 are depicted with bold lines. (c-d) The qRT-PCR and western blot analysis of the SOX13 regulation of SPON2 mRNA and protein expression levels. Values represent the means ± SD (n = 3, each group). ^**P < 0.01 vs. SOX13(+)NC group; ^##P < 0.01 vs. SOX13(−)NC group. (e-f) The mRNA and protein expression levels of SPON2 in ECs and GECs were evaluated by qRT-PCR and western blot. Data are presented as the means ± SD (n = 5, each group). ^**P < 0.01 vs. ECs group. (g) The effects of SPON2 knockdown on the viability of GECs were detected by the CCK-8 assay. Values represent the means ± SD (n = 5, each group). ^**P < 0.01 vs. SPON2(−)NC group. (h) The effects of SPON2 knockdown on the migration of GECs were determined by the transwell assay. Values represent the means ± SD (n = 5, each group). ^**P < 0.01 vs. SPON2(−)NC group. The scale bar represents 100 μm. (i) The effects of SPON2 knockdown on the tube formation of GECs were evaluated by the Matrigel tube formation assay. Values represent the means ± SD (n = 5, each group). ^**P < 0.01 vs. SPON2(−)NC group. The scale bar represents 100 μm