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Multiple elements regulate nuclear/cytoplasmic shuttling of FOXO1: characterization of phosphorylation- and

Xiangshan Zhao1, Lixia Gan, Haiyun Pan

  • 1Department of Medicine, University of Illinois at Chicago College of Medicine, and Medical Research Unit, Chicago, IL 60612, USA.

The Biochemical Journal
|December 11, 2003
PubMed

Insights

Forkhead box protein 1 (FOXO1) shuttles between the nucleus and cytoplasm via multiple phosphorylation-dependent mechanisms. These pathways, involving 14-3-3 proteins, are crucial for insulin and growth factor signaling.

Area of Science:

  • Cellular Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Forkhead box protein 1 (FOXO1) is a key transcription factor regulated by insulin and growth factors.
  • Understanding FOXO1's nuclear-cytoplasmic shuttling is vital for comprehending its cellular functions.
  • Existing knowledge identified a nuclear localization signal (NLS) and a nuclear export signal (NES) in FOXO1.

Purpose of the Study:

  • To elucidate the complex mechanisms governing FOXO1's nuclear/cytoplasmic shuttling.
  • To investigate the roles of phosphorylation sites and protein interactions in FOXO1 regulation.
  • To identify novel regulatory elements involved in FOXO1 localization.

Main Methods:

  • Utilized green fluorescent protein (GFP) fusion proteins to track FOXO1 localization.
  • Investigated the effects of specific phosphorylation sites (Thr-24, Ser-256, Ser-319, Ser-322, Ser-325, Ser-329) on FOXO1 shuttling.
  • Employed co-precipitation assays to study interactions between FOXO1 and 14-3-3 proteins.
  • Assessed the impact of leptomycin B on FOXO1 export.

Main Results:

  • Multiple mechanisms, including phosphorylation and novel elements within the DNA-binding domain (DBD), regulate FOXO1 nuclear localization.
  • Phosphorylation of Ser-319 and nearby residues, along with the NES, cooperatively drives nuclear exclusion.
  • The N-terminal region (amino acids 1-149), particularly amino acids 1-50, is sufficient for nuclear exclusion, requiring Thr-24 phosphorylation.
  • Phosphorylated FOXO1 binds 14-3-3 proteins via cooperative interactions at Thr-24 and Ser-256, potentially affecting DNA-binding and nuclear localization.

Conclusions:

  • FOXO1's cellular distribution is controlled by multiple, redundant regulatory mechanisms.
  • Phosphorylation and 14-3-3 protein binding are critical for modulating FOXO1's localization and function.
  • These intricate regulatory pathways underscore the significance of FOXO1 in insulin and growth factor signaling pathways.

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