Fig 2: Knockdown of ECT2 expression inhibits the proliferation, migration and invasion of PC-3 cells. (A) PC-3 cells were transfected with sh-ECT2-1/2 or sh-NC, and the mRNA levels of ECT2 were examined using reverse transcription-quantitative PCR. (B) Protein expression levels of ECT2 were examined by western blot analysis. (C) A colony formation assay was conducted to examine cell proliferation. (D) Cell viability was assessed using a Cell Counting Kit-8 assay. ***P<0.001 vs. sh-NC. (E) A wound-healing assay was performed to assess cell migration. Scale bar, 100 µm. (F) A Transwell assay was conducted to evaluate cell invasion. Scale bar, 100 µm. (G) Relative protein expression levels were examined by western blot analysis. ***P<0.001. ECT2, epithelial cell transforming sequence 2; NC, negative control; sh, short hairpin RNA.

Fig 3: Impact of knockdown of ECT2 expression on proliferation, migration and invasion of PC-3 cells is weakened by ETS1 overexpression. (A) PC-3 cells were transfected with sh-ECT2/sh-NC alone or co-transfected with sh-ECT2 and oe-ETS1/oe-NC, and a Cell Counting Kit-8 assay was performed to examine cell viability. ***P<0.001 sh-ECT2 vs. sh-NC; ###P<0.001 sh-ECT2 + oe-ETS1 vs. sh-ECT2 + oe-NC. (B) A colony formation assay was conducted to examine cell proliferation. (C) A wound healing assay was performed to assess cell migration. Scale bar, 100 µm. (D) A Transwell assay was performed to evaluate cell migration. Scale bar, 100 µm. (E) Relative protein expression was examined by western blot analysis. *P<0.05 and ***P<0.001. ECT2, epithelial cell transforming sequence 2; NC, negative control; oe, overexpression vector; sh, short hairpin RNA.

Fig 4: Knockdown of ECT2 expression restricts aerobic glycolysis in PC-3 cells. The cell culture medium was harvested and the (A) lactate release and (B) glucose uptake were examined using commercial kits. (C) The ECAR and OCR of PC-3 cells were assessed using a Seahorse XFe96 Analyzer to evaluate the glycolytic capacity and maximal respiration. (D) Western blotting was performed to examine the expression levels of several critical enzymes in the aerobic glycolysis process. ***P<0.001. 2-DG, 2-deoxy-D-glucose; A&R, antimycin A/rotenone; ECAR, extracellular acidification rate; ECT2, epithelial cell transforming sequence 2; FCCP, carbonylcyanide-4-(trifluoromethoxy) phenylhydrazone; Glc, glucose; HK2, hexokinase 2; LDHA, lactate dehydrogenase A; NC, negative control; OCR, oxygen consumption rate; Oligo, oligomycin; PKM2, pyruvate kinase M2; sh, short hairpin RNA.

Fig 5: Impact of knockdown of ECT2 expression on aerobic glycolysis in PC-3 cells is weakened by ETS1 overexpression. The cell culture medium was harvested and the (A) lactate release and (B) glucose uptake were examined using commercial kits. (C) The ECAR and OCR of PC-3 cells were assessed using a Seahorse XFe96 Analyzer to evaluate the glycolytic capacity and maximal respiration. (D) Western blotting was performed to examine the expression levels of several critical enzymes in the aerobic glycolysis process. *P<0.05 and ***P<0.001. 2-DG, 2-deoxy-D-glucose; A&R, antimycin A/rotenone; ECAR, extracellular acidification rate; ECT2, epithelial cell transforming sequence 2; FCCP, carbonylcyanide-4-(trifluoromethoxy) phenylhydrazone; Glc, glucose; HK2, hexokinase 2; LDHA, lactate dehydrogenase A; NC, negative control; OCR, oxygen consumption rate; oe, overexpression vector; Oligo, oligomycin; PKM2, pyruvate kinase M2; sh, short hairpin RNA.