FoxO4 Inhibits NF-κB and Protects Mice Against Colonic Injury and Inflammation

Abstract

Aims-FoxO4 is a member of the forkhead box transcription factor O (FoxO) subfamily. FoxO proteins are involved in diverse biological processes. In this study, we examine the role of FoxO4 in intestinal mucosal immunity and inflammatory bowel disease (IBD).Methods-Foxo4-null mice were subjected to trinitrobenzene sulfonic acid (TNBS)-treatment. Microarray analysis and quantitative RT-PCR were used to identify the cytokine transcripts that were altered by Foxo4-deletion. The effects of Foxo4-deficiency on the intestinal epithelial permeability and levels of tight junction proteins were examined by permeable fluorescent dye and western blot. The molecular and cellular mechanism(s) by which FoxO4 regulates the mucosal immunity were explored through immunological and biochemical analyses. The expression level of FoxO4 in intestinal epithelial cells of patients with IBD was examined using immunohistochemistry.Results-Foxo4-null mice were more susceptible to TNBS injury induced colitis. The chemokine CCL5 is significantly up-regulated in the colonic epithelial cells of Foxo4-null mice, with increased recruitment of CD4+ intraepithelial T cells and up-regulation of cytokines INFγ and TNFα in the colon. Foxo4-deficiency also resulted in an increase in intestinal epithelial permeability and downregulation of the tight junction proteins ZO-1 and claudin-1. Mechanistically, FoxO4 inhibited the transcriptional activity of NF-κB and Foxo4-deficiency is associated with increased NF-κB activity in vivo. FoxO4 transcription is transiently repressed in response to TNBS treatment and in patients with IBD. *To whom correspondence should be addressed:, Ph: 214 648-1485, Fx: 214 648-1450, E-mail: Zhi-Ping.Liu@utsouthwestern.edu. Author contribution: Wen Zhou, Qian Cao, and Qing-Jun Zhang-acquisition of data, analysis and interpretation of data; Yan Penganalysis and interpretation of data; Diego H. Castrillon and Ronald A. DePinho-material support, drafting of the manuscript; Zhi-Ping Liu-study concept and design, drafting of the manuscript, analysis and interpretation of data, and obtained funding.Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Conclusion-These results indicate that FoxO4 is an endogenous inhibitor of NF-κB and identify a novel function of FoxO4 in the regulation of NF-κB mediated mucosal immunity.

FOXO3 as a new IKK‐ε‐controlled check‐point of regulation of IFN‐β expression

“…One of the suggested mechanisms of NF-κB inhibition is upregulation of IκB expression directly or indirectly [15,29], but this is believed to be cell-type-dependent mechanism [10,11]. Another direct physical interaction, which could prevent NF-κB from either entering the nucleus or, as demonstrated for FOXO4, or its binding to the DNA [11,15]. Our data do not support the hypothesis that FOXO3 blocks nuclear translocation of NF-κB (Supporting Information Fig.…”

“…Another possibility is the sequestration of active NF-κB complexes, as described for FOXO4 [11]. Indeed, complex formation between HA-tagged FOXO3 and FLAG-tagged p65/RelA and IRF3 were detected in 293-TLR4 cells ectopically expressing the aforementioned proteins (Supporting Information Fig.…”

“…FOXO3 was implicated in regulation of NF-κB inhibitors, IκBs [11,15], with inhibition of FOXO3 resulting in attenuated expression of IL-8 in LPS-treated intestinal epithelia [29]. It has also been proposed that FOXO3 prevents NF-κB translocation to the nucleus [15].…”

“…More recently, FOXO factors have been shown to play a role in immunity and inflammation [7][8][9][10][11]. In addition to their critical role in homeostasis of immune-relevant cells including B and T cells [7][8][9], FOXOs are associated with inflammatory diseases [12,13].…”

“…Given their importance in such critical cellular functions, their activity is tightly regulated by posttranslational modifications, mainly via the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway [4]. In response to growth factors or cytokines, FOXO proteins are phosphorylated by AKT Correspondence: Dr. Lionel Luron e-mail: luron@ciml.univ-mrs.fr at three conserved serine/threonine residues (Thr 32 , Ser 253 , and Ser 315 of FOXO3) resulting in the protein inactivation via nuclear exclusion and subsequent degradation [5,6].More recently, FOXO factors have been shown to play a role in immunity and inflammation [7][8][9][10][11]. In addition to their critical role in homeostasis of immune-relevant cells including B and T cells [7][8][9], FOXOs are associated with inflammatory diseases [12,13].…”