Journal: International Journal of Molecular Medicine

Article Title: Downregulating FGGY carbohydrate kinase domain containing promotes cell senescence by activating the p53/p21 signaling pathway in colorectal cancer

doi: 10.3892/ijmm.2025.5522

Figure Lengend Snippet: FGGY knockdown promotes cell senescence by activating the p53 pathway. The correlation between FGGY and TP53 expression was analyzed using (A) Gene Expression Omnibus (accession number: GSE39582) and (B) The Cancer Genome Atlas databases. (C) Protein levels of p53, p21 and PCNA in HCT116 cells after FGGY knockdown were assessed by western blotting. GAPDH was used as a loading control. Band intensities were semi-quantified using ImageLab software. * P<0.05 vs. sh-Ctrl. (D) Expression levels of p21 and p53 proteins in HCT116/p53 +/+ and HCT116/p53 −/− cells transduced with sh-FGGY or sh-Ctrl were assessed by western blotting. GAPDH was used as a loading control. Band intensities were semi-quantified using ImageLab software. (E) Viability of HCT116/p53 +/+ and HCT116/p53 −/− cells transduced with sh-FGGY or sh-Ctrl. Results were normalized to viability on day 1. (F) Senescence-associated β-gal staining in HCT116/p53 +/+ and HCT116/p53 −/− cells after FGGY knockdown. Representative images were taken at a magnification of ×200. Immunofluorescence images showing co-localization of FGGY in chromatin foci with the SAHF markers (G) H3k9me3 and (H) HP1γ in HCT116/p53 +/+ and HCT116/p53 −/− cells after FGGY knockdown. Representative images were taken at a magnification of ×400. * P<0.05 vs. sh-Ctrl HCT116/p53 +/+ cells. β-gal, β-galactosidase; Ctrl, control; FGGY, FGGY carbohydrate kinase domain containing; H3k9me3, trimethylation of H3K9; sh, short hairpin.

Article Snippet: The cells were subsequently permeabilized with 0.1% Triton X-100, blocked with 1% BSA (cat. no. SW3015; Beijing Solarbio Science & Technology Co., Ltd.) for 1 h at room temperature, and were incubated overnight at 4°C with primary antibodies against HP1γ (cat. no. 2619; Cell Signaling Technology, Inc.), or trimethylation of H3K9 (H3k9me3; cat. no. 49-1008; Thermo Fisher Scientific, Inc.), at a dilution of 1:200.

Techniques: Knockdown, Expressing, Gene Expression, Western Blot, Control, Software, Transduction, Staining, Immunofluorescence