Lysates prepared from Hek293 cells transfected with wild type GFP-FOXO3 alone mutant S207A GFP-FOXO3 co-transfected with wild type MST1, or wild-type FOXO3 co-transfected with wild-type active MST1, were resolved on a 10% polyacrylamide gel and transferred to PVDF. The membrane was blocked with a 3% BSA-TBST buffer for one hour at room temperature, and then incubated with the FOXO3 [pS207] antibody for two hours at room temperature in 3% BSA-TBST buffer, following prior incubation with: no peptide, the non-phosphopeptide corresponding to the phosphopeptide immunogen, a generic phosphoserine containing peptide, or the phosphopeptide immunogen. After washing, the membrane was incubated with goat F(ab')2 anti-rabbit IgG HRP conjugate (Cat. ##ALI4404) and signals were detected using the Pierce SuperSignal™ method. The data show that MST1 induced the phosphorylation of FOXO3 at serine 207 and that the signal was abolished in cells over-expressing mutant S207A FOXO3 or in the presence of inactive MST1 kinase. The data also show that the signal is blocked only by the phosphopeptide corresponding to FOXO3 [pS207], indicating again its specificity.
|Tested species reactivity||Human, Mouse|
|Published species reactivity||Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||The antiserum was produced against a chemically synthesized phosphopeptide derived from the region of human FOXO3 that contains serine 207.|
|Purification||Antigen affinity chromatography|
|Storage buffer||Dulbecco's PBS, pH 7.3, with 1mg/ml BSA, 50% glycerol|
|Contains||0.05% sodium azide|
|Tested Applications||Dilution *|
|Western Blot (WB)||Assay Dependent|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Western Blot (WB)||See 1 publications below|
FOXO3 is a ubiquitously expressed 75 kDa protein member of a subfamily of the forkhead homeotic gene family of transcription factors and shuttles between the cytoplasm and nucleus. FOXO transcription factors are key players of cell fate decisions, metabolism, stress resistance, tumor suppression and are regulated by growth factors, oxidative stress or nutrient deprivation. FOXO3 is involved with mTOR in the regulation of autophagy in skeletal muscle, and activates protein degradation in atrophying muscle cells. FOXO3 has also been implicated in several neurodegenerative disorders including aging, neuromuscular disease, systemic lupus erythmatosus, stroke and diabetic complications.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
|Rat||Not Cited||A conserved MST-FOXO signaling pathway mediates oxidative-stress responses and extends life span.||Lehtinen MK,Yuan Z,Boag PR,Yang Y,Villén J,Becker EB,DiBacco S,de la Iglesia N,Gygi S,Blackwell TK,Bonni A||Cell (125:987)||2006|