anti vasp  (Proteintech)

Journal: Cell Death & Disease

Article Title: IGF2BP3 regulates EMP1 stability in an m 6 A-dependent manner and activates the TGF-β pathway to promote pancreatic cancer invasion

doi: 10.1038/s41419-025-08155-1

Figure Lengend Snippet: Immunoprecipitation assays obtain silver-stained bands of proteins binding to EMP1-FLAG ( A ); and LC-MS/MS analysis to demonstrate the binding sites of EMP1 and VASP ( B ); In PANC-1 and AsPC-1, Co-IP experiments depict the interactions between EMP1 and VASP ( C ); Immunofluorescence staining of PANC-1 and AsPC-1 demonstrating colocalization of EMP1 and VASP , red ( VASP ), green ( EMP1 ) and blue ( DAPI ) ( D ); Analysis of IGF2BP3 , EMP1 , with VASP correlation in the TCGA-PAAD dataset ( E , F ); Diagram of docking patterns of VASP and SMAD7 proteins ( G ); Immunofluorescence staining of PANC-1 and AsPC-1 demonstrating colocalization of VASP and SMAD7 , red ( VASP ), green ( SMAD7 ) and blue ( DAPI ) ( H ); In PANC-1 and AsPC-1, Co-IP demonstrates the interactions of VASP and SMAD7 ( I) and EMP1 expression affects this binding relation ( J); Western blot detection of EMP1-VASP-SMAD7 axis regulation of TGF-β/Smads signaling pathway in PANC-1 and AsPC-1 ( K) . Scale: 20 μm. ***, P < 0.001.

Article Snippet: Membranes were then incubated with IGF2BP3 (Abcam, ab179807, US), GAPDH (Proteintech, 60004-1-Ig, China), EMP1 (Abcam, ab230445, US), VASP (Proteintech,13472-1-AP, China), SMAD2 (Proteintech,12570-1-AP, China), SMAD3 (Proteintech,66516-1-1 g, China), SMAD7 (Proteintech,25840-1-AP, China), phospho-SMAD2 (CST,138D4, US), and phospho-SMAD3 (CST, 9520S, US) at 4 °C overnight.

Techniques: Immunoprecipitation, Staining, Binding Assay, Liquid Chromatography with Mass Spectroscopy, Co-Immunoprecipitation Assay, Immunofluorescence, Expressing, Western Blot

Journal: Cell Death & Disease

Article Title: IGF2BP3 regulates EMP1 stability in an m 6 A-dependent manner and activates the TGF-β pathway to promote pancreatic cancer invasion

doi: 10.1038/s41419-025-08155-1

Figure Lengend Snippet: METTL3 mediates the modification of m 6 A on the EMP1 RAN, and IGF2BP3 promotes the stabilization of EMP1 RNA based on the recognition of the modification site of m 6 A on EMP1 , and elevated EMP1 protein levels, which increased EMP1 and VASP binding. Meanwhile, the combination of EMP1 and VASP enhanced the function of VASP , which is to impede the inhibition of the pathway of TGF-β signaling by SMAD7 , ultimately leading to the sustained activation of TGF-β in pancreatic cancer. And the effect of METTL3/IGF2BP3/EMP1 on the microenvironment of pancreatic cancer was also demonstrated, including: ( CD8 + T cell, CD68+ Macrophage, α-SMA+ fibroblast, CD20 + B cell and CD11c+ Dendritic cell).

Article Snippet: Membranes were then incubated with IGF2BP3 (Abcam, ab179807, US), GAPDH (Proteintech, 60004-1-Ig, China), EMP1 (Abcam, ab230445, US), VASP (Proteintech,13472-1-AP, China), SMAD2 (Proteintech,12570-1-AP, China), SMAD3 (Proteintech,66516-1-1 g, China), SMAD7 (Proteintech,25840-1-AP, China), phospho-SMAD2 (CST,138D4, US), and phospho-SMAD3 (CST, 9520S, US) at 4 °C overnight.

Techniques: Modification, Binding Assay, Inhibition, Activation Assay

Journal: PLOS One

Article Title: Integrated bioinformatics and experimental validation identify ATF3 as a key gene in secondary brain damage after intracerebral hemorrhage

doi: 10.1371/journal.pone.0328530

Figure Lengend Snippet: (A) Evolutionary-conserved ATF3 binding motif identified by JASPAR (Matrix ID MA0606.1) within the VASP promoter (−1.5 kb upstream of TSS). Position frequency matrix conservation score >85% across mammals. (B) Reanalysis of GSE24265 microarray data showing VASP upregulation (linear models for microarray data [LIMMA], adj.p = 0.017). (C) ATF3 gain-of-function model: Western blot of HT-22 cells 48 h post-lentiviral ATF3 overexpression (pLVX-EF1α vector). Blots: Rabbit anti-ATF3 (CST 33593, 1:1,000), mouse anti-VASP (BD 611175, 1:500), β-actin control (Sigma A2228, 1:5,000). (D/E) Densitometric quantification of ATF3 (D) and VASP (E) protein levels (n = 3 biological replicates; unpaired t-test, **p = 0.0007 vs. EV control). (F) ATF3 loss-of-function: shRNA-mediated knockdown in HT-22 cells. Blots: 72 h post-transduction; scramble shRNA control (Addgene #1864). (G/H) The quantification of ATF3 (G) and VASP (H) suppression (n = 3; two-way ANOVA, **p < 0.0001 vs. scramble). (I) Dual-luciferase reporter assay of VASP promoter activity. Left: Wild-type vs. ΔATF3-binding mutant constructs. Right: Firefly/Renilla ratio (n = 6; **p = 0.0004, ordinary one-way ANOVA).

Article Snippet: For immunostaining, cells were incubated with either a mouse monoclonal ATF3 antibody (Abcam, ab254268) or a rabbit polyclonal VASP antibody (Proteintech, 13472-1-AP, China) as primary antibodies.

Techniques: Binding Assay, Microarray, Western Blot, Over Expression, Plasmid Preparation, Control, shRNA, Knockdown, Transduction, Luciferase, Reporter Assay, Activity Assay, Mutagenesis, Construct