Journal: Frontiers in Oncology

Article Title: Ring finger protein 128 promotes, rather than inhibits, colorectal cancer progression by regulating the Hippo signaling pathway

doi: 10.3389/fonc.2022.1031160

Figure Lengend Snippet: RNF128 ubiquitinates MST for degradation and regulates target genes of the Hippo signaling pathway. (A) Co-IP assays revealed that RNF128 interacts with MST. (B) An IP assay was used to examine the ubiquitination levels of MST. HCT116 cells were treated with MG132 (10 μM) for 6 h after transfection with si-NC and si-RNF128 for 48 h. (C, D) The expression levels of HIF-1α, Bcl-2, p-IκBα and IκBα were examined by western blotting. HCT116 cells were transfected with si-NC and si-RNF128 and treated with verteporfin (1 μM) for 16 h and PY-60 (10 μM) for 24 h. ** P <0.001.

Article Snippet: The proteasome inhibitor MG132 (MedChemExpress, HY-13259) was used to treat HCT116 cells for 6 h, the YAP inhibitor verteporfin (TargetMol, T3112) was used to treat HCT116 cells for 16 h, and the YAP agonist PY-60 (TargetMol, T9566) was used to treat HCT116 cells for 24 h.

Techniques: Co-Immunoprecipitation Assay, Ubiquitin Proteomics, Transfection, Expressing, Western Blot

Journal: Bone Research

Article Title: Lumbar instability remodels cartilage endplate to induce intervertebral disc degeneration by recruiting osteoclasts via Hippo-CCL3 signaling

doi: 10.1038/s41413-024-00331-x

Figure Lengend Snippet: Activation of YAP attenuates cartilage degradation during LSI. a Top, representative 3D Micro-CT images of the caudal endplates of L4-5 level (coronal view) from LSI mice with or without Lats-IN-1 treatment. Bottom, quantitative analysis of the total porosity and Tb. Sp. b Representative SOFG staining images of the CEP from LSI mice with or without Lats-IN-1 treatment. c Endplate scores of the caudal endplates based on ( b ). d Left, representative IF staining images of the Collagen II and Collagen X (green) of the CEP from LSI mice with or without Lats-IN-1 treatment. Right, quantitative analysis of the percentage of each area in CEP. e Gene expression of Col2a1 and Col10a1 in CEPCs treated with 12% CTS and Lats-IN-1. f WB analysis of Collgen II, Collgen X and YAP in CEPCs treated with 12% CTS and Lats-IN-1. g , h Left, IF staining of Collagen II and Collagen X in Ctrl, 12% CTS, Ctrl+Lats-IN-1 and 12% CTS+Lats-IN-1 groups. Right, quantitative analysis of the mean fluorescence intensity. F-actin (red), Collagen II and Collagen X (Green), DAPI (blue). * P < 0.05, ** P < 0.01

Article Snippet: YAP agonists Lats-IN-1 (10 μmol/L, MCE) and inhibitor Verteporfin (2 μmol/L, Sigma-Aldrich) was supplemented in culture medium during the entire course of mechanical loading., ,

Techniques: Activation Assay, Micro-CT, Staining, Gene Expression, Fluorescence

Journal: Bone Research

Article Title: Lumbar instability remodels cartilage endplate to induce intervertebral disc degeneration by recruiting osteoclasts via Hippo-CCL3 signaling

doi: 10.1038/s41413-024-00331-x

Figure Lengend Snippet: Loss of YAP triggered CCL3 expression in CEPCs to active osteoclastogenesis. a Volcano plot analysis of the genes expression change profile in CEPCs treated with 12% CTS. b ELISA analysis of CCL3 concentration in the supernatant of CEPCs treated with CTS. c IHC and quantitative analysis of CCL3 in CEP from mice with sham or LSI surgery. d WB analysis of CCL3 in CEPs from mice taken sham or LSI surgery after 8 weeks. e Transwell assay to evaluate the BMM migration under CCL3 attraction. The BMMs were seeded on the upper chamber and different concentrations of CCL3 were added into the lower chamber. Representative images are shown on the left and random cell counting manually in five random fields under ×100 magnification on the right. f WB analysis of YAP and CCL3 in CEPCs treated with 12% CTS and YAP agonists Lats-IN-1. g IF staining of CCL3 in CEPCs treated with 12% CTS and YAP agonists Lats-IN-1. h TRAP staining of BMMs after induction with the supernatant of CTS-treated CEPCs supplemented with CCL3-neutralizing antibody. i Quantative analysis of osteoclast number in ( g ). j Luciferase assay of CCL3 reporter activity to identify the active region of CCL3 promoter in HEK293T cells. k Luciferase assay of CCL3 reporter activity with transfection of control or Yap1-overexpression plasmid in HEK293T cells. l Gene expression analysis of TEADs in CEPCs treated by 12% CTS. * P < 0.05. ** P < 0.01

Article Snippet: YAP agonists Lats-IN-1 (10 μmol/L, MCE) and inhibitor Verteporfin (2 μmol/L, Sigma-Aldrich) was supplemented in culture medium during the entire course of mechanical loading., ,

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Concentration Assay, Transwell Assay, Migration, Cell Counting, Staining, Luciferase, Activity Assay, Transfection, Control, Over Expression, Plasmid Preparation, Gene Expression