Journal: Phytotherapy research : PTR
Article Title: Gossypin inhibits gastric cancer growth by direct targeting AURKA and RSK2
doi: 10.1002/ptr.6253
Figure Lengend Snippet: (A) Effect of gossypin on EGF-induced activation of various signaling proteins in JB6 cells. Serum-starved (0.1% FBS; 48 h) JB6 cells were treated with various doses of gossypin for 2 h followed by treatment with EGF for 30 min. (B) Effect of gossypin on various signaling proteins in HGC27 gastric cancer cells. Cells were treated with gossypin and then signaling pathway proteins were detected by Western blotting. (C) Gossypin inhibits phosphorylation of histone H3 in AGS gastric cancer cells. Cells were treated with gossypin for 24 h and detected by immunofluorescence assay. Red color indicates phosphorylated histone H3 at serine 10 and blue color shows DAPI. (D-E) Gossypin suppresses AURKA and RSK2 kinase activity in a dose-dependent manner. The effect of gossypin on AURKA or RSK2 activity was assessed by an in vitro kinase assay using AURKA (active, 200 ng) and MBP (AURKA substrate, 400 ng) or RSK2 (active, 200 ng) and ATF1 (RSK2 substrate, 300 ng) proteins. The effect of gossypin on kinase activities was determined by Western blotting using an antibody to detect phosphorylated serine and threonine. Band density was measured using the Image J (NIH) software program. For all data, similar results were obtained from 3 independent experiments.
Article Snippet: The active recombinant AURKA (300 ng) or RSK2 (200 ng) protein was mixed with different concentrations of gossypin in reaction buffer (Cell Signaling Technology) and incubated at room temperature for 15 min.
Techniques: Activation Assay, Western Blot, Phospho-proteomics, Immunofluorescence, Activity Assay, In Vitro, Kinase Assay, Software