Investigating fundamental signal transduction mechanisms impacted by FOXO transcription factors on NOTCH and JAK/STAT Pathways in stem cell contexts

ABSTRACT We propose a high impact collaborative project to overcome barriers to understanding the roles and regulation of Forkhead box subfamily O transcription factors FOXO -1, -3, and -4 (FOXO) in stem cell contexts. Stem cells repopulate the lining of the intestinal tract, muscle, and blood lineages, yet also drive devastating diseases such

as cancer. Evolutionarily conserved, partially redundant FOXO transcription factors are needed for stem cell maintenance in a litany of contexts such as embryonic, cancer, mesenchymal, hematopoietic, and neural cell lineages, as well as direct specification of subsequent lineages in a context-dependent manner. Canonically,

high PI3K output leads to cytoplasmic/inactive FOXO factors. Stem cells, however, are fundamentally rewired to have both high PI3K Pathway activity and nuclear/active FOXO factors. Prior work shows that FOXO factors directly bind to and activate stem genes in both embryonic stem cells and the poor prognosis cancer glioblastoma

multiforme (GBM) to drive stem cell fate. However, the precise mechanisms utilized by FOXO transcription factors in driving stemness, hindering differentiation, and determining cell fate are incompletely understood. Our novel, preliminary insights indicate that nuclear-localized FOXOs engage both the NOTCH and JAK/STAT

signaling pathways. Our preliminary evidence demonstrates a strong loss in the gene expression of NOTCH Pathway components NOTCH1 and NOTCH3, as well as their targets including HES1, in FOXO4 loss-of-function contexts. Our preliminary data indicates that FOXO factors share the ability to promote NOTCH output.

Motivated by our preliminary findings and published work (by others), we propose to examine whether FOXO - 1, -3, and -4 redundantly drive NOTCH1 and NOTCH3 gene expression in GBM and myoblasts. Our preliminary work with FOXO transcription factor disruption mutants shows that FOXO4 acts uniquely to hinder JAK/STAT3

activation and expression of targets IGF1, PDGF and TH. Of note, FOXO3 disruption led to a loss in STAT3 activation, underscoring unique contributions of FOXO factors to the JAK/STAT Pathway. We propose to dissect the ability of FOXO factors to differentially regulate the JAK/STAT Pathway in GBM and myoblast settings. Our

long-term goal is to delineate FOXO-driven fundamental biological processes that aid in stem cell maintenance and cell fate by promoting the NOTCH Pathway and differentially regulating the JAK/STAT Pathway. The proposed work will integrally involve student researchers (our group has successfully sent eleven students to

doctoral studies) and will promote the research environment at UTRGV, the second-largest Hispanic serving institution in the United States.