# Dynamics of the FOXO transcription factor network

> **NIH NIH R01** · UNIVERSITY OF ARIZONA · 2021 · $57,372

## Abstract

Summary of funded parent award
The FOXO family of transcription factors are evolutionarily conserved regulators of homeostasis whose
activities are linked to both increased lifespan and tumor suppression. Consistent with their role in
maintaining cellular homeostasis, FOXO activity is upregulated by diverse types of cellular stress including
nutrient/growth factor deprivation, DNA damage and oxidative stress. Control of FOXO activity is
predominantly achieved through post-translational modifications that control nuclear-cytoplasmic shuttling
of FOXO proteins. In the nucleus, FOXOs upregulate genes in multiple, often conflicting pathways
including cell-cycle arrest, apoptosis, autophagy and ROS scavenger genes. How cells control FOXO
activity to ensure that their response is appropriate for a given stress is an open question. To address this
question we used CRISPR/Cas9 gene editing to fluorescently tag two FOXO proteins, Foxo1 and Foxo3a,
at the endogenous locus of different cell lines. We use these lines to test the hypothesis that input/output
specificity of the FOXO pathway is achieved through a dynamic control mechanism where different FOXO
nuclear/cytoplasmic shuttling dynamics dictate separate cellular responses. Our hypothesis is inspired by
similarities between the FOXO pathway and other transcription factors that use dynamic control
mechanisms for input/output specificity including p53 and NF-ΚB. In addition, our preliminary data
supports a role for FOXO dynamics in controlling cell fate. We found the single-cell dynamics of Foxo1
and Foxo3a shuttling change with different stimuli. Moreover, for the same stimulus we observed different
dynamics for each isoform. In Aim 1 we explore the shuttling dynamics of Foxo1 and Foxo3a in response
to serum starvation. We combine reverse phase protein arrays and RNA-seq to determine how time-
dependent changes in key regulators control the dynamics of each isoform and how this is translated into
different gene expression patterns. In Aim 2 we measure the dynamics of Foxo1 and Foxo3a shuttling as
well as cell death in response to EGFR and Akt inhibitors. Previous experiments have shown that both
Foxo1 and Foxo3a are required for cell death in response to EGFR inhibitors. We determine the dynamics
of each isoform associated with cell death and develop transcriptional reporters to determine how
dynamics are decoded by cells in terms of transcriptional output. In Aim 3 we develop an optogenetic
system to control Foxo1 shuttling dynamics with light. We use this system to determine whether specific
dynamic patterns of Foxo1 shuttling are sufficient to induce cell death and use RNA-seq to determine how
changes in dynamics alter target gene expression. The experiments performed in this study will address a
critical gap in our knowledge of how FOXO dynamics are controlled over time to enact specific outcomes.
More broadly, our work will help elucidate how cell signaling circuits sense and respond to different
...

## Key facts

- **NIH application ID:** 10298949
- **Project number:** 3R01GM130864-03S1
- **Recipient organization:** UNIVERSITY OF ARIZONA
- **Principal Investigator:** Andrew Luther Paek
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $57,372
- **Award type:** 3
- **Project period:** 2019-01-01 → 2023-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10298949

## Citation

> US National Institutes of Health, RePORTER application 10298949, Dynamics of the FOXO transcription factor network (3R01GM130864-03S1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10298949. Licensed CC0.

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