P pkm2

Protein extraction from NCI-H2030 and NCI-H1650 cells was implemented via the RIPA solution (P0013C; Beyotime, China). Protein concentrations were tested utilizing BCA assay kits (P0011; Beyotime). The extracted proteins were denatured in boiling water for 15-minutes. Thereafter, electrophoresis was carried out utilizing 10% SDS-PAGE gel for separation of protein molecules. Following protein transference onto PVDF membranes, the proteins were blocked in 5% nonfat milk at room temperature for 2 hours. Afterwards, incubation with primary antibody of β-catenin (1/1000; ab265591; Abcam, USA), E-cadherin (1/2000; ab231303), GAPDH (1/2500; ab9485), PKM2 (1/1000; #4053; Cell Signaling Technology, USA), phosphorylated PKM2 (p-PKM2; 1/1000; #3827; Cell Signaling Technology), lactate dehydrogenase A (LDHA; 1/1000; #3582; Cell Signaling Technology), and glucose transporter 1 (GLUT1; 1/1000; #73015; Cell Signaling Technology) was implemented at 4°C lasting 18 hours, followed by incubation with IgG-HRP goat anti-rabbit (1/3000; #7074; Cell Signaling Technology) for 2 hours. Signals were developed with ECL (Sigma-Aldrich) and normalized.

The following antibodies were used for western blot and immunofluorescent studies: Girdin (AF5345, R&D Systems, Minneapolis, MN, USA); Girdin (A16132, ABclonal, Wuhan, China); PKM2 (#4053, Cell Signaling Technology, Danvers, MA, USA); p-PKM2 (#3827, Cell Signaling Technology, Danvers, MA, USA); p53 (#2524, Cell Signaling Technology, Danvers, MA, USA); GST (sc459, Santa Cruz, Santa Cruz, CA, USA); GFP (598, MBL, Tokyo, Japan); β-Actin (66009-1-Ig, Proteintech, Wuhan, China). The uncropped Western Blot figures for relevant results are shown in Figures S2–S5.

Protein extraction and Western blotting experiments were performed as described in our previous publications [9 (link), 49 (link)]. The following primary antibodies were used: antibodies against KLF4 (Cell Signaling Technology, Inc., Danvers, MA, USA and Proteintech Group, Inc, Rosemont, IL, USA), c-Myc, PTBP1, PARP, LC3B, p-PKM2, GAPDH (Cell Signaling Technology, Inc., Danvers, MA, USA), PKM1, and PKM2 (Novus Biologicals, USA). HRP-conjugated goat anti-rabbit and horse anti-mouse IgG (Cell Signaling Technology) were used as secondary antibodies. GAPDH served as an internal control.