Assessing the Activity of Transcription Factor FoxO1

The transcription factor FoxO1 (forkhead box O1) regulates genes that are involved in development, metabolism, cellular innovation, longevity, and stress responses. Assessment of FoxO1 activity is therefore critical to understand the regulatory network of this transcription factor. FoxO1 transactivation activity relies on its ability to bind to the promoters of target genes, which is controlled by posttranslational modifications (e.g., dephosphorylation or phosphorylation) that may promote nuclear translocation or exclusion of FoxO1. In this chapter we describe the protocols for FoxO1 activity assessment using Western blotting analysis of the posttranslational modification of FoxO1 in whole cell lysates and ELISA of DNA binding activity of FoxO1 in nuclear extracts.