Bimodal regulation of FoxO3 by AKT and 14-3-3

• Published in: Biochimica et biophysica acta 2011-08, Vol.1813 (8), p.1453-1464
• Main Authors: Dobson, Melissa, Ramakrishnan, Gopalakrishnan, Ma, Stephanie, Kaplun, Ludmila, Balan, Vitaly, Fridman, Rafael, Tzivion, Guri
• Format: Article
• Language: English
• Subjects: 14-3-3; 14-3-3 Proteins - chemistry; 14-3-3 Proteins - genetics; 14-3-3 Proteins - metabolism; AKT; Binding Sites; Cell Line; Forkhead Box Protein O3; Forkhead domain; Forkhead Transcription Factors - chemistry; Forkhead Transcription Factors - genetics; Forkhead Transcription Factors - metabolism; FoxO; Hep G2 Cells; Humans; Models, Biological; Mutagenesis, Site-Directed; Phosphorylation; Protein Interaction Domains and Motifs; Protein phosphorylation; Proto-Oncogene Proteins c-akt - chemistry; Proto-Oncogene Proteins c-akt - genetics; Proto-Oncogene Proteins c-akt - metabolism; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Transcription factors
• ISSN: 0167-4889; 0006-3002; 1879-2596
• DOI: 10.1016/j.bbamcr.2011.05.001

FoxO3 is a member of FoxO family transcription factors that mediate cellular functions downstream of AKT. FoxO3 phosphorylation by AKT generates binding sites for 14-3-3, which in-turn regulates FoxO3 transcriptional activity and localization. We examine here the functional significance of AKT–FoxO3 interaction and further detail the mechanistic aspects of FoxO3 regulation by AKT and 14-3-3. Our data show that AKT overexpression increases the steady-state levels of FoxO3 protein in a manner dependent on AKT activity and its ability to bind FoxO3. Characterization of the AKT–FoxO3 interaction shows that the three AKT phosphorylation-site-recognition motifs (RxRxxS/T) present on FoxO3, which are required for FoxO3 phosphorylation, are dispensable for AKT binding, suggesting that AKT has a docking point on FoxO3 distinct from the phosphorylation-recognition motifs. Development of a FoxO3 mutant deficient in 14-3-3 binding (P34A), which can be phosphorylated by AKT, established that 14-3-3 binding and not AKT phosphorylation per se controls FoxO3 transcriptional activity. Intriguingly, 14-3-3 binding was found to stabilize FoxO3 by inhibiting its dephosphorylation and degradation rates. Collectively, our data support a model where both AKT and 14-3-3 positively regulate FoxO3 in addition to their established negative roles and that 14-3-3 availability could dictate the fate of phosphorylated FoxO3 toward degradation or recycling. ► AKT binding to FoxO3 is independent of the RxRxxS/T AKT phosphorylation recognition motif. ► AKT activation is required for FoxO3 binding. ► AKT expression increases FoxO3 steady-state protein levels. ► 14‐3‐3 binding stabilizes phosphorylated FoxO3. ► FoxO3 P34A mutation impairs 14‐3‐3 binding, resulting in a constitutively active form.