human stub1 chip  (Cell Signaling Technology Inc)

Journal: Frontiers in Immunology

Article Title: Targeting the BAG2/CHIP axis promotes gastric cancer apoptosis by blocking apoptosome assembly

doi: 10.3389/fimmu.2025.1578416

Figure Lengend Snippet: FIIN-2 blocks the BAG2-CHIP interaction. (A) Interactions between CHIP and different BAG2 deletion mutants analyzed via co-immunoprecipitation assays. WT, wild type; IP, immunoprecipitation; WCL, whole-cell-lysates. (B) Interactions between BAG2 and different CHIP deletion mutants were examined using co-immunoprecipitation assays. (C) The bound conformation of BAG2 and CHIP as predicted by the Cluspro algorithm. BAG2 is displayed in yellow, and CHIP is displayed in green. (D) This schematic diagram shows the amino acids that interact between BAG2 and CHIP. On the binding surface, the CHIP residue bonds are highlighted in green, while those of BAG2 are in yellow. (E) Interactions between BAG2 and different CHIP deletion mutants containing residues on the binding surface of the mode were analyzed via co-immunoprecipitation assays. (F) Flow diagram of BAG2-CHIP complex inhibitor screening. (G) Computational modeling showcases the interactions between FIIN-2 and CHIP. CHIP is displayed in green, and FIIN-2 is displayed in pink. (H) Microscale thermophoresis (MST) was utilized to ascertain the kinetic constant (Kd) for the interaction between FIIN-2 and CHIP. (I) Co-immunoprecipitation assays of the BAG2-CHIP interaction in cells treated with FIIN-2 at the indicated concentrations in HGC-27. IP, immunoprecipitation; WCL, whole-cell lysates. (J) Western blotting was conducted to assess HSP70 expression levels in cells post-treatment with different FIIN-2 concentrations in HGC-27. (K) An in vitro ubiquitination assay was performed to determine the impact of FIIN-2 (C = 10 μM) on HSP70 ubiquitination, using specified recombinant proteins in HGC-27.

Article Snippet: CHIP human recombinant was purchased from OriGene (cat. TP300310, China).

Techniques: Immunoprecipitation, Binding Assay, Residue, ChIP-chip, Microscale Thermophoresis, Western Blot, Expressing, In Vitro, Ubiquitin Proteomics, Recombinant

Journal: Journal of cellular and molecular medicine

Article Title: Aryl Hydrocarbon Receptor Alleviates Hepatic Fibrosis by Inducing Hepatic Stellate Cell Ferroptosis.

doi: 10.1111/jcmm.70278

Figure Lengend Snippet: FIGURE 1 | AHR directly regulates Mrp1 transcription in mHSCs. (A–C) The expression of Ahr, Mrp1 or Cyp1a1 was detected by QPCR. (D) There are two potential AHR exogenous response elements on the Mrp1 promoter sequence. (E) Promoter sequences containing XREL1 and XREL2 were cloned onto PGL3 plasmids, and mutant plasmids were constructed. (F) Double luciferase assay to detect luciferase activity. (G) EMSA detects AHR binding to specific elements XREL1 and XREL2. (H) CHIP detects AHR binding to specific elements XREL1 and XREL2. Data are expressed as means ± SD; *p < 0.05, **p < 0.01 and ***p < 0.001; Student's t-test or one-way ANOVA.

Article Snippet: ChIP assays were performed using the ChIP Assay kit (Beyotime, P2078). mHSCs were treated with YH439 (5 μM) for 24 h. Subsequently, mHSCs were sonicated and then immunoprecipitated with the antibody against AHR (1:100 dilution BOSTER Cat# A00225- 4, RRID: AB_3095576) with IgG (1:100 dilution Proteintech Cat# 30000- 0- AP RRID AB_2819035) as a negative control.

Techniques: Expressing, Sequencing, Clone Assay, Mutagenesis, Construct, Luciferase, Activity Assay, Binding Assay