Journal: Oncotarget

Article Title: Glycycoumarin exerts anti-liver cancer activity by directly targeting T-LAK cell-originated protein kinase

doi: 10.18632/oncotarget.11610

Figure Lengend Snippet: A. GCM inhibited TOPK in HepG2 cells. The cells were treated with various concentrations of GCM for 24 h and the changes of phospho-, total TOPK and its substrate histone H3 phosphorylation were determined by western blotting. B. GCM promoted TOPK degradation in HepG2 cells. The cells were treated with 50μM of GCM in the presence or absence of CHX and the expression of TOPK was analyzed by western blotting. C. GCM inhibited TOPK in Huh-7 cells. D. GCM inhibited TOPK in HCT-116 cells. E. GCM inhibited TOPK in DU145 cells. F. Overall inhibitory effects of TOPK knockdown on HepG2 cells measured by crystal violet staining. G. TOPK knockdown activated p53 in HepG2 cells. The cells were transfected with 5 nmol/L of TOPK siRNA or non-targeting siRNA for 36 h and the changes of p-p53, p21 and puma were analyzed by western blotting.

Article Snippet: Subsequently, GCM was docked into these three predicted TOPK structures with both Autodock4.0 and Autodock vina [ , ], and the docking parameters were kept as default.

Techniques: Phospho-proteomics, Western Blot, Expressing, Knockdown, Staining, Transfection

Journal: Oncotarget

Article Title: Glycycoumarin exerts anti-liver cancer activity by directly targeting T-LAK cell-originated protein kinase

doi: 10.18632/oncotarget.11610

Figure Lengend Snippet: A. Three predicted structures of the full-length TOPK (left) and their ATP-binding sites (right). B. The docking model of GCM / TOPK complex structure. GCM is shown in sphere representation (carbons are colored by cyan. TOPK is shown as a cartoon model, left) and in surface representation of TOPK (the location of GCM is shown in ball-stick, right). C. Binding site of TOPK with GCM. The ATP-binding site of TOPK is shown in surface representation (white: hydrophobic residues, blue: residues with positive charge, red: residues with negative charge, green: other polar residues). GCM is shown in ball-stick representation. D. The detailed Interaction between TOPK and GCM. Carbons on TOPK are colored grey. GCM is shown in ball-stick representation and key residues on TOPK are shown in stick representation. Carbons, oxygen, nitrogen and sulfur atoms are colored cyan, red, blue and yellow respectively. E. GCM binds directly to TOPK. Sepharose 4B was used for binding and pull-down assay. Lane 1, the negative control, indicating no binding between TOPK and beads alone; Lane 2, TOPK binds to GCM-Sepharose 4B beads; lane 3, input control. F. GCM inhibited TOPK activity in vitro in a dose dependent manner. An inactive GST-Histine H3 protein was used as the substrate with active TOPK and the p-histone H3 was measured by western blotting.

Article Snippet: Subsequently, GCM was docked into these three predicted TOPK structures with both Autodock4.0 and Autodock vina [ , ], and the docking parameters were kept as default.

Techniques: Binding Assay, Pull Down Assay, Negative Control, Control, Activity Assay, In Vitro, Western Blot

Journal: Oncotarget

Article Title: Glycycoumarin exerts anti-liver cancer activity by directly targeting T-LAK cell-originated protein kinase

doi: 10.18632/oncotarget.11610

Figure Lengend Snippet: GCM was given daily by i.p. injection (30 mg/kg body weight) after one week inoculation of HepG2 cells for 24 days. A. Body weight kinetics of mice. B. Inhibitory effects of GCM on tumor growth. C. Reduction of final tumor weight. D. Influences of GCM on TOPK activity and p53 signaling assessed by western blotting.

Article Snippet: Subsequently, GCM was docked into these three predicted TOPK structures with both Autodock4.0 and Autodock vina [ , ], and the docking parameters were kept as default.

Techniques: Injection, Activity Assay, Western Blot