Journal: Neoplasia (New York, N.Y.)

Article Title: BPGM as an intrinsic brake to constrain metastasis through phospho-epigenetic-mediated carnitine biosynthesis suppression

doi: 10.1016/j.neo.2026.101299

Figure Lengend Snippet: 2,3-BPG-CDK1-EZH2-H3K27me3 Axis: BPGM’s epigenetic circuit breaker for cellular migration. (A) Integrated functional metabolomics analysis revealed BPGM-altered metabolites clustered in methyl donor group. Bubble size: metabolites count. (B) Hypothesis of molecular mechanism underlying BPGM regulated BBOX1 expression by post transcriptional modification (PTM). (C) Silencing BPGM significantly reduced the protein level of H3K27me3, while overexpressing BPGM increased its level. Cells stably expressing shBPGM/BPGM and its control cells (shCtrl/Ctrl) were used to detect protein level by western blotting. (D) ChIP assays disclosed that the fragments of BBOX1 and MMP9 promoter precipitated by anti-H3K27me3 antibody were increased upon overexpressing BPGM. SK-HEP-1 cells stably expressing BPGM and its control cells (Ctrl) were employed to ChIP assay. The antibody precipitated DNAs were amplified by qPCR. 5 % of the total DNAs were amplified to serve as the control for DNA content. Values shown are signal of α-H3K27me3-precipitated DNA relative to the input and the mean value of the control group was normalized as 1. (E) Overexpressing BPGM significantly increased the protein level of EZH2 but decreased the protein level of p-EZH2-T 345 in tumor cells. (F) The molecular docking of 2,3-BPG and CDK1. Predicted structure of 2,3-BPG binding with CDK1. Key contact residues: Thr14, Arg127, Arg170. (G) Overexpressing BPGM significantly increased the protein level of p-CDK1-T 14 in tumor cells. Cells stably expressing BPGM (BPGM-OE) and its control cells (Ctrl) were used to detect protein level by western blotting. (H) 2,3-BPG treatment enhanced the phosphorylation of CDK1 at thr14 in tumor cells. The indicated concentration of 2,3-BPG was incubated with the lysate of trophoblasts and tumor cells for 30 minutes followed by western blotting. (I-J) RO-3306 treatment enhanced the phosphorylation of CDK1 at thr14 and reduced the phosphorylation of EZH2 at thr345 in tumor cells. The tumor cells were treated with the indicated concentration of RO-3306 for 12 hours followed by western blotting. (K) The model deciphers the role of BPGM in regulating BBOX1 and MMP9 expression. Error bar: mean ± SEM. P -values are labeled above the bar chart.

Article Snippet: The following reagents were used: l -carnitine (541-15-1, Sigma-Aldrich, Saint Louis, MO, USA); 2,3-BPG (D5764, Sigma-Aldrich); CDK1 inhibitor, RO-3306 (S7747, Selleck, Shanghai, China).

Techniques: Migration, Functional Assay, Expressing, Modification, Stable Transfection, Control, Western Blot, Amplification, Binding Assay, Phospho-proteomics, Concentration Assay, Incubation, Labeling

Journal: Scientific Reports

Article Title: Transcriptional activation of PPP1R14C by KLF7 unleashes CDK1 activity to promote lung squamous cell carcinoma

doi: 10.1038/s41598-026-39174-3

Figure Lengend Snippet: PPP1R14C promotes LUSC cell proliferation and invasion via CDK1 activation. ( A ) Overexpression of PPP1R14C activates CDK1 through T161-CDK1 phosphorylation. Western blot analysis revealed that compared to the control group, overexpression of PPP1R14C in NCI-H520 cells significantly increased the phosphorylation levels of T161-CDK1 and histone H1 (p-CDK1, P-T161(CDK1), and p-Histone H1). ( B ) PPP1R14C knockdown inhibits CDK1 activation by reducing T161-CDK1 phosphorylation. Western blot analysis showed that compared to the control group, PPP1R14C knockdown (shPPP1R14C) significantly reduced the expression levels of p-CDK1, P-T161(CDK1), and p-Histone H1. ( C ) Interaction between PPP1R14C and PP1α. Co-immunoprecipitation experiments showed that PPP1R14C antibodies pulled down PP1α, while isotype control IgG did not detect PP1α signals, indicating that PPP1R14C can directly bind to PP1α. ( D ) Overexpression of PPP1R14C inhibits the binding of CDK1 to PP1α. Co-immunoprecipitation experiments showed that PP1α could co-precipitate with CDK1 in cells expressing the empty vector, while overexpression of PPP1R14C inhibited the binding of CDK1 to PP1α. ( E ) CDK1 inhibition abolished PPP1R14C-mediated proliferation. CCK-8 proliferation assays showed that the growth advantage conferred by PPP1R14C overexpression was completely abolished by the CDK1 inhibitor RO-3306 (* p < 0.05, ** p < 0.01, **** p < 0.0001). ( F ) CDK1 activity is critical for PPP1R14C-driven clonogenicity. Colony formation experiments showed that inhibition of CDK1 with RO-3306 neutralized PPP1R14C-driven clonogenic growth (* p < 0.05, **** p < 0.0001). ( G ) CDK1 inhibition attenuated the invasive ability enhanced by PPP1R14C. Transwell invasion assays showed that blocking CDK1 with RO-3306 could inhibit the pro-invasive effect of PPP1R14C overexpression (* p < 0.05, **** p < 0.0001).

Article Snippet: The CDK1-specific inhibitor RO-3306 (MedChemExpress) was dissolved in DMSO to a 10 mM stock solution and used at a final concentration of 10 μM for in vitro assays.

Techniques: Activation Assay, Over Expression, Phospho-proteomics, Western Blot, Control, Knockdown, Expressing, Immunoprecipitation, Binding Assay, Plasmid Preparation, Inhibition, CCK-8 Assay, Activity Assay, Blocking Assay