FoxO6, a Novel Member of the FoxO Class of Transcription Factors with Distinct Shuttling Dynamics

Abstract

Forkhead transcription factors of the FoxO-group are associated with cellular processes like cell cycle progression and DNA-repair. FoxO function is regulated by protein kinase B (PKB) via the phosphatidylinositol 3-kinase/PKB survival pathway. Phosphorylation of serine and threonine residues in specific PKB phosphorylation motifs leads to exclusion of FoxO-proteins from the nucleus, which excludes them from exerting transactivating activity. Members of the FoxO-group have three highly conserved regions containing a PKB phosphorylation motif. This study describes the cloning and characterization of a novel forkhead domain gene from mouse that appeared to be highly related to the FoxO group of transcription factors and was therefore designated FoxO6. The FoxO6 gene was mapped in region D1 on mouse chromosome 4. In humans, FOXO6 is located on chromosomal region 1p34.1. Embryonic expression of FoxO6 is most apparent in the developing brain, and FoxO6 is expressed in a specific temporal and spatial pattern. Therefore it is probably involved in regulation of specific cellular differentiation. In the adult animal FoxO6 expression is maintained in areas of the nucleus accumbens, cingulate cortex, parts of the amygdala, and in the hippocampus. Structure function analysis of FoxO6 compared with its group members shows that the overall homology is high, but surprisingly a highly conserved region containing multiple phosphorylation sites is lacking. In transfection studies, FoxO6 coupled to GFP showed an unexpected high nuclear localization after stimulation with growth factors, in contrast to the predominant cytosolic localization of FoxO1 and FoxO3. We also show that nuclear export of FoxO6 is mediated through the phosphatidylinositol 3-kinase/PKB pathway. Furthermore, we show using a chimeric approach that we can fully restore the ability of FoxO6 to shuttle between nucleus and cytosol. In conclusion, the data presented here gives a new view on regulation of FoxO-function through multiple phosphorylation events and other mechanisms involved in the nuclear exclusion of FoxO-proteins.