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The Forkhead transcription factor FKHR (FOXO1) regulates expression of gluconeogenic factors, and plays a key role in the insulin regulation of glucose levels. Besides its role in glucose homeostasis, FKHR has been shown to be involved in pathological processes such as type 2 diabetes, cancer and organismal aging. The transcriptional output of FKHR is controlled by a two-tiered mechanism of phosphorylation and acetylation, and alterations in this balance can result in profound effects.
Figure 1: Measurement of phosphorylated and total FKHR.
CHO-K1 cells were cultured in 96-well plates and serum-starved for 16 hours. Cells were then treated with 20% FBS for 30 minutes and fixed. Total and phospho FKHR were each assayed in triplicate using the phospho and total FKHR antibodies included in the FACE FKHR Kit. Data was plotted after correction for cell number (performed through use of Crystal Violet).
Antibody Specificities
The phospho-FKHR antibody is specific for phosphorylated FKHR and was raised against a synthetic phospho-peptide corresponding to residues surrounding Thr24 of human FKHR. This antibody recognizes FKHR only when phosphorylated at Thr24 and also recognizes FKHRL1 when phosphorylated at Thr32 and AFX when phosphorylated at Thr28. The total-FKHR antibody recognizes FKHR proteins regardless of the phosphorylation state.
