Forkhead Transcription Factor FOXO3a Research Articles

Potent FOXO3a Activators from Biologically Active Compound Library for Cancer Therapeutics: Aninsilico Approach.

The forkhead transcription factor FOXO3a is a member of the FOXO subfamily, which controls a number of cellular processes including apoptosis, proliferation, cell cycle progression, DNA damage, and carcinogenesis. In addition, it reacts to a number of biological stressors such as oxidative stress and UV radiation. FOXO3a has been predominantly associated with many diseases including cancer. Recent research suggests that FOXO3a suppresses tumor growth in cancer. By cytoplasmic sequestration of the FOXO3a protein or mutation of the FOXO3a gene, FOXO3a is commonly rendered inactive in cancer cells. Furthermore, the onset and development of cancer are linked to its inactivation. In order to reduce and prevent tumorigenesis, FOXO3a needs to be activated. So, it is critical to develop new strategies to enhance FOXO3a expression for cancer therapy. Hence, the present study has beenaimed to screen small molecules targeting FOXO3a using bioinformatics tools. Molecular docking and molecular dynamic simulation studies reveal the potent FOXO3a activating small molecules such as F3385-2463, F0856-0033, and F3139-0724. These top three compounds will be subjected to further wet experiments. The findings of this study will lead us to explore the potent FOXO3a activating small moleculesfor cancer therapeutics.

Transcription Factor FOXO3a Overexpression Inhibits the Progression of Neuroblastoma by Regulating the miR-21/SPRY2/ERK Axis

Neuroblastoma is one of the most common extracranial solid tumors in children. The forkhead transcription factor FOXO3a has been implicated in the progressionof a variety of human diseases. Here, we aim to identify the effects of FOXO3a on the malignancy of neuroblastoma. Bioinformatics analysis was employed to identify differentially expressed genes related to neuroblastoma and the downstream regulator of FOXO3a. FOXO3a expression was examined in SH-SY5Y neuroblastoma cells. Interactions between FOXO3a and microRNA-21 (miR-21) were then identified using bioinformatics analysis and dual-luciferase reporter assay. After ectopic expression and depletion experiments in SH-SY5Y cells, cell malignant phenotypes were assessed by cell counting kit-8 and Transwell assays. FOXO3a-overexpressing neuroblastoma cells were xenografted into nude mice to validate the role of FOXO3a in tumor growth. Downregulated expression of FOXO3a was observed in neuroblastoma cells, with a negative correlation between FOXO3a and miR-21 expression. FOXO3a bound to the promoter region of miR-21 to downregulate its expression, resulting in inhibition of SH-SY5Y cell malignant phenotypes. Additionally, miR-21 targeted SPRY2 by binding to the 3'UTR of the mRNA encoding SPRY2, activating the extracellular signal-regulated kinase (ERK) pathway. FOXO3a disrupted the binding of miR-21 to SPRY2 and inactivated ERK to suppress the malignant phenotypes of SH-SY5Y cells as well as tumor growth invivo. In conclusion, FOXO3a may inhibit the progression of neuroblastoma by suppressing the miR-21 expression and facilitating SPRY2-dependent ERK pathway inactivation.

Novel effects of Foxo3a on immune system and cardiac remodeling after myocardial infarction

Abstract Background Fibroblast/myofibroblast transdifferentiation following acute cardiac injury (MI) is a major mechanism of scar formation and adverse remodeling regulated by TGFβ/SMAD3 signaling. Myofibroblasts represent phenotypically modulated cells characterized by α-smooth muscle actin (α-SMA) and constitute the main source of extracellular matrix proteins such as collagen type I in the healing infarct wound. Moreover, the recruitment of a specific monocyte/macrophages subset (Ly6Chigh and Ly6Clow) control the healing process in the infarcted wound. The forkhead transcription factor FOXO3a has recently been shown to inhibit cardiac hypertrophy by different stressors and is involved in the immune response following injury. Purpose We hypothesized that FOXO3a, a key regulator of cell differentiation, cell cycle and size as well as the stress response, might inhibit matricellular remodeling following myocardial infarction by regulating transdifferentiation of fibroblasts into myofibroblasts. Moreover, we hypothesized that Foxo3a−/− mice favor the recruitment of monocytes of the type Ly6Clow leading to an anti-inflammatory profile and thus the remodeling after injury. Methods Acute myocardial infarction was induced in FOXO3−/− and WT mice (FVB background) by permanent LAD ligation. The myocardial infarction size was determined via ECHO. Myofibroblast trans-differentiation and fibrosis markers were analyzed by QT-PCR and protein analysis. The differentiation of macrophages studied via FACS. IP/IF and Western blotting were used to test for a direct interaction between FOXO3a and SMAD3 in vitro. Results FOXO3a−/− mice showed significantly higher survival rates compared to WT littermates. Myocardial inflammation (Mac-1, LFA, CD3, CD45), injury markers such as Troponin T were similar in both groups on day 4 post-MI. Myocardial expression of alpha smooth muscle actin (ASMA) and Collagen1A1 (Col1A1), as well as numbers of cardiac CD11b+/F4–80+/Ly6clow macrophages was significantly enhanced in FOXO3a−/− mice 15 days post infarction. Moreover, Foxo3a−/− mice showed larger fibrotic areas following MI. Mechanistically, immunoprecipitation showed direct interaction of FOXO3a with SMAD3a that was enhanced following activation of the transcription factor leading to diminished SMAD3 downstream gene expression. Conclusion Our results indicate that FOXO3a acts as a direct inhibitor of TGF-β regulated myofibroblast differentiation and matrix remodeling via FOXO3a-SMAD3 interaction following MI. Moreover, a Foxo3a deficiency might lead to the recruitment of monocytes type Ly6Clow leading to an anti-inflammatory profile in the infarct wound. Therefore, FOXO3a collectively regulates cardiac hypertrophy and fibrosis/scar formation following MI. Thus, targeting the FOXO3a-SMAD3 signaling axis might be of future therapeutic interest. Funding Acknowledgement Type of funding source: Foundation. Main funding source(s): German Resaerch Foundation

β-amyloid-induced gonadotropin-releasing hormone decline involving Forkhead transcription factor FOXO3a and nuclear factor-κB

Previously, it has been demonstrated that aging is controlled by the hypothalamus, and that hypothalamus-driven programmatic aging is associated with nuclear factor-κB (NF-κB)-mediated gonadotropin-releasing hormone (GnRH) decrease. Abundant accumulation of β-amyloid (Aβ) has been observed in brains of cognitively normal elderly. However, it is unclear whether Aβ neurotoxicity is involved in aging-associated hypothalamic GnRH decline. GT1-7 cells, which are a cell line of hypothalamic GnRH neurons, were used in the current study to investigate whether and how Aβ decreased GnRH release. The results of the current study demonstrated that Aβ impaired the release of GnRH through activation of NF-κB. Mechanistic studies revealed that Aβ activated NF-κB via Forkhead box protein O3a, thereby inhibiting gnrh1 gene. The results of the present study provided novel insights into the mechanisms underlying aging-dependent hypothalamic GnRH decline.

Downregulated FOXO3a Associates With Poor Prognosis and Promotes Cell Invasion and Migration via WNT/β-catenin Signaling in Cervical Carcinoma.

Background: Emerging studies have demonstrated that the Forkhead transcription factor FOXO3a is closely correlated with the progression of multiple tumors. Nevertheless, the biological role and prognostic value of FOXO3a have yet to be fully elucidated in cervical carcinoma. This study was designed to determine the molecular mechanism and prognosis of FOXO3a in cervical carcinoma.Methods: The protein levels of FOXO3a were detected using immunohistochemistry and Western blotting. The relationships between FOXO3a expression and clinicopathological variables were analyzed. The biological mechanism of FOXO3a in cervical carcinoma cells (HeLa and CaSki) was investigated. We also explored the effect of FOXO3a on WNT/β-catenin signaling with respect to its expression and function.Results: The results demonstrated that decreased FOXO3a expression was related to increased tumor stage and grade, positive lymph node metastasis, and poor survival outcome in cervical carcinoma. Survival analysis revealed that the FOXO3a level is an independent prognostic factor for cervical carcinoma patients. Furthermore, the data indicated that the downregulation of FOXO3a expression promotes cell invasion and migration, while FOXO3a overexpression exhibited the opposite effects on cervical carcinoma. In addition, FOXO3a acted as a negative regulator of the canonical WNT/ β-catenin pathway in cervical carcinoma. Moreover, overexpression of FOXO3a also inhibited the expression of MMP2 and MMP9.Conclusion: These results reveal that FOXO3a, serving as a tumor suppressor gene, could suppress cell invasion and migration via the WNT/β-catenin signaling pathway and indicates a good prognosis in cervical carcinoma.

Dynamic Interaction of USP14 with the Chaperone HSC70 Mediates Crosstalk between the Proteasome, ER Signaling, and Autophagy.

SummaryUSP14 is a deubiquitinating enzyme associated with the proteasome important for protein degradation. Here we show that upon proteasome inhibition or expression of the mutant W58A-USP14, association of USP14 with the 19S regulatory particle is disrupted. MS-based interactomics revealed an interaction of USP14 with the chaperone, HSC70, in neuroblastoma cells. Proteasome inhibition enhanced binding of USP14 to HSC70, and to XBP1u and IRE1α proteins, demonstrating a role in the unfolded protein response. Striatal neurons expressing mutant huntingtin exhibited reduced USP14 and HSC70 levels, whereas inhibition of HSC70 downregulated USP14. Furthermore, proteasome inhibition or use of the mutant W58A-USP14 facilitated the interaction of USP14 with the autophagy protein, GABARAP. Functionally, overexpression of W58A-USP14 increased GABARAP positive autophagosomes in striatal neurons, and this was abrogated using the HSC70 inhibitor, VER-155008. Modulation of the USP14-HSC70 axis may represent a potential therapeutic target in HD to beneficially influence multiple proteostasis pathways.

Forkhead transcription factor FOXO3a mediates interferon-γ-induced MHC II transcription in macrophages.

Macrophages are professional antigen-presenting cells relying on the expression of class II major histocompatibility complex (MHC II) genes. Interferon-γ (IFN-γ) activates MHC II transcription via the assembly of an enhanceosome centred on class II trans-activator (CIITA). In the present study, we investigated the role of the forkhead transcription factor FOXO3a in IFN- γ-induced MHC II transcription in macrophages. Knockdown of FOXO3a, but not FOXO1 or FOXO4, diminished IFN-γ-induced MHC II expression in RAW cells. On the contrary, over-expression of FOXO3a, but neither FOXO1 nor FOXO4, enhanced CIITA-mediated trans-activation of the MHC II promoter. IFN-γ treatment promoted the recruitment of FOXO3a to the MHC II promoter. Co-immunoprecipitation and RE-ChIP assays showed that FOXO3a was a component of the MHC II enhanceosome forming interactions with CIITA, RFX5, RFXB and RFXAP. FOXO3a contributed to MHC II transcription by altering histone modifications surrounding the MHC II promoter. Of interest, FOXO3a was recruited to the type IV CIITA promoter and directly activated CIITA transcription by interacting with signal transducer of activation and transcription 1 in response to IFN-γ stimulation. In conclusion, our data unveil a novel role for FOXO3a in the regulation of MHC II transcription in macrophages.

Abstract 73: Gain-of-function mutant p53 promotes the oncogenic potential of head and neck squamous cell carcinoma cell by targeting forkhead transcription factors FOXO3a and FOXM1

Abstract Many mutant p53 proteins exert oncogenic gain-of-function (GOF) properties in promoting cancer cell invasive growth and metastasis, but the mechanisms mediating these functions largely remain elusive. We show here that overexpression of the GOF mutant p53 G245D and other GOF p53 mutants enhances invasive cell growth of p53-deficient head and neck squamous cell carcinoma (HNSCC) UM-SCC-1 cells both in vitro 3D culture and in vivo orthotopic tumor mouse model. We demonstrate that the expression of the oncogenic forkhead transcription factor FOXM1 is upregulated by GOF mutant p53s in UM-SCC-1 cells and tumors. Moreover, we show that overexpression of GOF mutant p53 G245D decreases the inhibitory phosphorylation of FOXO3a, a tumor suppressive forkhead transcription factor, leading to its cytoplasmic accumulation. Consistent with the observation that FOXO3a negatively regulates FOXM1 expression, we demonstrate that overexpression FOXO3a decreases GOF mutant p53-mediated FOXM1 expression, whereas downregulation of FOXO3a partially restores decreased FOXM1 expression due to loss of GOF mutant p53. In support of the roles of FOXO3a-FOXM1 signaling axis in mutant p53's GOF signaling, we further show that overexpression of FOXO3a or downregulation of FOXM1 impairs GOF mutant p53-mediated cell invasion, whereas downregulation of FOXO3a, in part, rescues impaired cell invasion due to downregulated GOF mutant p53. Finally, given that AMP-activated protein kinase (AMPK) directly phosphorylates and promotes FOXO3a function, and that our prior demonstration that GOF mutant p53s inhibit AMPK, our current study establishes that GOF mutant p53-AMPK-FOX3a-FOXM1 signaling is one of important mechanisms through which mutant p53s gain oncogenic functions in HNSCCs. Citation Format: Noriaki Tanaka, Mei Zhao, Jiexin Zhang, Jing Wang, Ge Zhou, Jeffrey N. Myers. Gain-of-function mutant p53 promotes the oncogenic potential of head and neck squamous cell carcinoma cell by targeting forkhead transcription factors FOXO3a and FOXM1 [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; April 23-25, 2017; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(23_Suppl):Abstract nr 73.

Overexpression of FoxO3a is associated with glioblastoma progression and predicts poor patient prognosis.

Forkhead transcription factor FoxO3a has been reported to have ambiguous functions and distinct mechanisms in various solid tumors, including glioblastoma (GBM). Although a preliminary analysis of a small sample of patients indicated that FoxO3a aberrations in glioma might be related to aggressive clinical behavior, the clinical significance of FoxO3a in glioblastoma remains unclear. We investigated the expression of FoxO3a in a cohort of 91 glioblastoma specimens and analyzed the correlations of protein expression with patient prognosis. Furthermore, the functional impact of FoxO3a on GBM progression and the underlying mechanisms of FoxO3a regulation were explored in a series of in vitro and in vivo assays. FoxO3a expression was elevated in glioblastoma tissues, and high nuclear FoxO3a expression in human GBM tissues was associated with poor prognosis. Moreover, knockdown of FoxO3a significantly reduced the colony formation and invasion ability of GBM cells, whereas overexpression of FoxO3a promoted the colony formation and invasion ability. The results of in vivo GBM models further confirmed that FoxO3a knockdown inhibited GBM progression. More, the pro-oncogenic effects of FoxO3a in GBM were mediated by the activation of c-Myc, microtubule-associated protein 1 light chain 3 beta (LC3B) and Beclin1 in a mixed-lineage leukemia 2 (MLL2)-dependent manner. These findings suggest that high FoxO3a expression is associated with glioblastoma progression and that FoxO3a independently indicates poor prognosis in patients. FoxO3a might be a novel prognostic biomarker or a potential therapeutic target in glioblastoma.

Cordycepin induces apoptosis by caveolin-1-mediated JNK regulation of Foxo3a in human lung adenocarcinoma

Forkhead transcription factor (Foxo3a) is a downstream effector of JNK-induced tumor suppression. However, it is not clear whether the caveolin-1 (CAV1)-mediated JNK/Foxo3a pathway is involved in cancer cell apoptosis. We found that cordycepin upregulates CAV1 expression, which was accompanied by JNK phosphorylation (p-JNK) and subsequent Foxo3a translocation into the nucleus, resulting in the upregulation of Bax protein expression. Furthermore, we found that CAV1 overexpression upregulated p-JNK, whereas CAV1 siRNA downregulated p-JNK. Additionally, SP600125, a specific JNK inhibitor, significantly increased Foxo3a phosphorylation, which downregulated Foxo3a translocation into the nucleus, indicating that CAV1 mediates JNK regulation of Foxo3a. Foxo3a siRNA downregulated Bax protein and attenuated A549 apoptosis, indicating that the CAV1-mediated JNK/Foxo3a pathway induces the apoptosis of A549 lung cancer cells. Cordycepin significantly decreased tumor volume in nude mice. Taken together, these results indicate that cordycepin promotes CAV1 upregulation to enhance JNK/Foxo3a signaling pathway activation, inducing apoptosis in lung cancer cells, and support its potential as a therapeutic agent for lung cancer.

Inhibition of ROS and upregulation of inflammatory cytokines by FoxO3a promotes survival against Salmonella typhimurium.

Virulent intracellular pathogens, such as the Salmonella species, engage numerous virulence factors to subvert host defence mechanisms to induce a chronic infection that leads to typhoid or exacerbation of other chronic inflammatory conditions. Here we show the role of the forkhead transcription factor FoxO3a during infection of mice with Salmonella typhimurium (ST). Although FoxO3a signalling does not affect the development of CD8+ T cell responses to ST, FoxO3a has an important protective role, particularly during the chronic stage of infection, by limiting the persistence of oxidative stress. Furthermore, FoxO3a signalling regulates ERK signalling in macrophages, which results in the maintenance of a proinflammatory state. FoxO3a signalling does not affect cell proliferation or cell death. Thus, these results reveal mechanisms by which FoxO3a promotes host survival during infection with chronic, virulent intracellular bacteria.

Amyloid-β induced astrocytosis and astrocyte death: Implication of FoxO3a-Bim-caspase3 death signaling.

Astrocytes, the main element of the homeostatic system in the brain, are affected in various neurological conditions including Alzheimer's disease (AD). A common astrocytic reaction in pathological state is known as astrocytosis which is characterized by a specific change in astrocyte shape due to cytoskeletal remodeling, cytokine secretion and cellular proliferation. Astrocytes also undergo apoptosis in various neurological conditions or in response to toxic insults. AD is pathologically characterized by progressive deposition of amyloid-β (Aβ) in senile plaques, intraneuronal neurofibrillary tangles, synaptic dysfunction and neuron death. Astrocytosis and astrocyte death have been reported in AD brain as well as in response to Aβ in vitro. However, how astrocytes undergo both proliferation and death in response to Aβ remains elusive. In this study, we used primary cultures of cortical astrocytes and exposed them to various doses of oligomeric Aβ. We found that cultured astrocytes proliferate and manifest all signs of astrocytosis at a low dose of Aβ. However, at high dose of Aβ the activated astrocytes undergo apoptosis. Astrocytosis was also noticed in vivo in response to Aβ in the rat brain. Next, we investigated the mechanism of astrocyte apoptosis in response to a high dose of Aβ. We found that death of astrocyte induced by Aβ requires a set of molecules that are instrumental for neuron death in response to Aβ. It involves activation of Forkhead transcription factor Foxo3a, induction of its pro-apoptotic target Bim and activation of its downstream molecule, caspase3. Hence, this study demonstrates that the concentration of Aβ decides whether astrocytes do proliferate or undergo apoptosis via a mechanism that is required for neuron death.

Antiproliferative and pro-apoptotic effects of three fungal exocellular β-glucans in MCF-7 breast cancer cells is mediated by oxidative stress, AMP-activated protein kinase (AMPK) and the Forkhead transcription factor, FOXO3a

Fungal β-d-glucans of the (1→3)-type are known to exhibit direct antitumor effects, and can also indirectly decrease tumor proliferation through immunomodulatory responses. The underlying molecular mechanisms involved in decreasing tumor formation, however, are not well understood. In this study, we examined the antiproliferative role and mechanism of action of three different fungal exocellular β-glucans in MCF-7 breast cancer cells. The β-glucans were obtained from Botryosphaeria rhodina MAMB-05 [two botryosphaerans; (1→3)(1→6)-β-d-glucan; one produced on glucose, the other on fructose] and Lasiodiplodia theobromae MMPI [lasiodiplodan; (1→6)-β-d-glucan, produced on glucose]. Using the cell proliferation-MTT assay, we showed that the β-glucans exhibited a time- and concentration-dependent antiproliferative activity (IC50, 100μg/ml). Markers of cell cycle, apoptosis, necrosis and oxidative stress were analyzed using flow cytometry, RT-PCR and Western blotting. Exposure to β-glucans increased apoptosis, necrosis, oxidative stress, mRNA expression of p53, p27 and Bax; the activity of AMP-activated protein-kinase, Forkhead transcription factor FOXO3a, Bax and caspase-3; and decreased the activity of p70S6K in MCF-7 cells. In the presence of hydrogen peroxide, the fungal β-glucans increased oxidative stress, which was associated with reduced cell viability. We showed that these β-glucans exhibited an antiproliferative effect that was associated with apoptosis, necrosis and oxidative stress. This study demonstrated for the first time that the apoptosis induced by β-glucans was mediated by AMP-activated protein-kinase and Forkhead transcription factor, FOXO3a. Our findings provide novel mechanistic insights into their antiproliferative roles, and compelling evidence that these β-glucans possess a broad range of biomodulatory properties that may prove useful in cancer treatment.

Hydrogen sulfide upregulates KATP channel expression in vascular smooth muscle cells of spontaneously hypertensive rats.

The study was designed to investigate whether H2S could upregulate expression of KATP channels in vascular smooth muscle cells (VSMCs), and by this mechanism enhances vasorelaxation in spontaneously hypertensive rats (SHR). Blood pressure, vascular structure, and vasorelaxation were analyzed. Plasma H2S was detected using polarographic sensor. SUR2B and Kir6.1 expressions were detected in VSMCs of SHR and in A7r5 cells as well as primarily cultured ASMCs using real-time PCR, western blot, immunofluorescence, and confocal imaging. Nuclear translocation of forkhead transcription factors FOXO1 and FOXO3a in ASMCs was detected using laser confocal microscopy, and their binding activity with SUR2B and Kir6.1 promoters was examined by chromatin immunoprecipitation. SHR developed hypertension at 18weeks. They showed downregulated vascular SUR2B and Kir6.1 expressions in association with a decreased plasma H2S level. H2S donor, however, could upregulate vascular SUR2B and Kir6.1 expressions, causing a left shift of the vasorelaxation curve to pinacidil and lowered tail artery pressure in the SHR. Also, H2S antagonized endothelin-1 (ET-1)-inhibited KATP expression in A7r5 cells and cultured ASMCs. Mechanistically, H2S inhibited ET-1-stimulated p-FOXO1 and p-FOXO3a expressions (inactivated forms), but increased their nuclear translocation and the ET-1-inhibited binding of FOXO1 and FOXO3a with Kir6.1 and SUR2B promoters in ASMCs. Hence, H2S promotes vasorelaxation of SHR, at least in part, through upregulating the expression of KATP subunits by inhibiting phosphorylation of FOXO1 and FOXO3a, and stimulating FOXO1 and FOXO3a nuclear translocation and their binding activity with SUR2B and Kir6.1 promoters. H2S increased vascular SUR2B and Kir6.1 expression of SHR, promoting vasorelaxation. H2S antagonized ET-1-inhibited KATP expression in A7r5 cells and cultured ASMCs. H2S inhibited ET-1-induced FOXO1 and FOXO3a phosphorylation in ASMCs. H2S promoted FOXO1 and FOXO3a nuclear translocation and binding with target gene promoters.

Forkhead Transcription Factor FOXO3a Levels Are Increased in Huntington Disease Because of Overactivated Positive Autofeedback Loop

Huntington disease (HD) is a fatal autosomal dominant neurodegenerative disorder caused by an increased number of CAG repeats in the HTT gene coding for huntingtin. Decreased neurotrophic support and increased mitochondrial and excitotoxic stress have been reported in HD striatal and cortical neurons. The members of the class O forkhead (FOXO) transcription factor family, including FOXO3a, act as sensor proteins that are activated upon decreased survival signals and/or increased cellular stress. Using immunocytochemical screening in mouse striatal Hdh(7/7) (wild type), Hdh(7/109) (heterozygous for HD mutation), and Hdh(109/109) (homozygous for HD mutation) cells, we identified FOXO3a as a differentially regulated transcription factor in HD. We report increased nuclear FOXO3a levels in mutant Hdh cells. Additionally, we show that treatment with mitochondrial toxin 3-nitropropionic acid results in enhanced nuclear localization of FOXO3a in wild type Hdh(7/7) cells and in rat primary cortical neurons. Furthermore, mRNA levels of Foxo3a are increased in mutant Hdh cells compared with wild type cells and in 3-nitropropionic acid-treated primary neurons compared with untreated neurons. A similar increase was observed in the cortex of R6/2 mice and HD patient post-mortem caudate tissue compared with controls. Using chromatin immunoprecipitation and reporter assays, we demonstrate that FOXO3a regulates its own transcription by binding to the conserved response element in Foxo3a promoter. Altogether, the findings of this study suggest that FOXO3a levels are increased in HD cells as a result of overactive positive feedback loop.

Forkhead transcription factor FOXO3a potently induces smooth muscle cell apoptosis and promotes extracellular matrix degradation through activation of matrix metalloproteinases

Forkhead transcription factor FOXO3a potently induces smooth muscle cell apoptosis and promotes extracellular matrix degradation through activation of matrix metalloproteinases

MiR-155 regulates lymphocyte homeostasis in LAT mutant mice (LYM4P.749)

Abstract LAT Y136F mutant mice carry a tyrosine-to-phenylalanine mutation in the LAT adapter gene that leads to a lymphoproliferative disease (LPD) initiated by CD4+ helper T cells. In a screen to detect microRNAs (miRs) specifically regulated in LAT Y136F CD4+ T cells, we found that levels of miR-155 were higher in these T cells than in other proliferating CD4+ T cells from wild type C57BL/6 mice. In order to reveal a potential role of miR-155 in LAT Y136F LPD, we crossed LAT Y136F mice to miR-155-deficient mice yielding double mutant (DM) mice. This resulted in a delay and decreased severity of LPD in DM mice. DM CD4+ T cells showed equal or higher proliferation rates than CD4+ T cells from age-matched LAT Y136F mice, however levels of apoptosis in DM CD4+ T cells were higher. One direct target downregulated by miR-155 is the forkhead transcription factor FOXO3a. In DM T cells, increased expression of FOXO3a led to accumulation of the pro-apoptotic factor Bim and enhanced apoptosis. In addition, elevated Pak1 kinase activity in DM T cells led to activation of Jnk leading to phosphorylation of FOXO3a, FOXO3a nuclear translocation and increased FOXO3a activity. Consequently, direct and indirect effects of miR-155 on FOXO3a led to an increase in Bim-mediated apoptosis and decreased LPD in DM mice. Investigation of downstream targets of miR-155 has led to new information on signaling molecules that control LPD and lymphocyte homeostasis.

The Small GTPase RALA Controls c-Jun N-terminal Kinase-mediated FOXO Activation by Regulation of a JIP1 Scaffold Complex

FOXO (forkhead box O) transcription factors are tumor suppressors and increase the life spans of model organisms. Cellular stress, in particular oxidative stress caused by an increase in levels of reactive oxygen species (ROS), activates FOXOs through JNK-mediated phosphorylation. Importantly, JNK regulation of FOXO is evolutionarily conserved. Here we identified the pathway that mediates ROS-induced JNK-dependent FOXO regulation. Following increased ROS, RALA is activated by the exchange factor RLF (RalGDS-like factor), which is in complex with JIP1 (C-Jun-amino-terminal-interacting protein 1) and JNK. Active RALA consequently regulates assembly and activation of MLK3, MKK4, and JNK onto the JIP1 scaffold. Furthermore, regulation of FOXO by RALA and JIP1 is conserved in C. elegans, where both ral-1 and jip-1 depletion impairs heat shock-induced nuclear translocation of the FOXO orthologue DAF16.

Aldose reductase inhibition enhances TRAIL-induced human colon cancer cell apoptosis through AKT/FOXO3a-dependent upregulation of death receptors

One of the major problems associated with the chemotherapy of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) that selectively kills tumor cells is decreased drug resistance. This warranted the development of safe novel pharmacological agents that could sensitize the tumor cells to TRAIL. Herein, we examined the role of aldose reductase (AR) in sensitizing cancer cells to TRAIL and potentiating TRAIL-induced apoptosis of human colon cancer cells. We demonstrate that AR inhibition potentiates TRAIL-induced cytotoxicity in cancer cells by upregulation of both death receptor (DR)-5 and DR4. Knockdown of DR5 and DR4 significantly (>85%) reduced the sensitizing effect of the AR inhibitor fidarestat on TRAIL-induced apoptosis. Further, AR inhibition also downregulates cell survival proteins (Bcl-xL, Bcl-2, survivin, XIAP, and FLIP) and upregulates the expression of proapoptotic proteins such as Bax and alters mitochondrial membrane potential, leading to cytochrome c release, caspases-3 activation, and PARP cleavage. We found that AR inhibition regulates AKT/PI3K-dependent activation of forkhead transcription factor FOXO3a. Knockdown of FOXO3a significantly (>80%) abolished AR inhibition-induced upregulation of DR5 and DR4 and apoptosis in colon cancer cells. Overall, our results show that fidarestat potentiates TRAIL-induced apoptosis through downregulation of cell survival proteins and upregulation of death receptors via activation of the AKT/FOXO3a pathway.

Role and regulation of the forkhead transcription factors FOXO3a and FOXM1 in carcinogenesis and drug resistance

The FOXO3a and FOXM1 forkhead transcription factors are key players in cancer initiation, progression, and drug resistance. Recent research shows that FOXM1 is a direct transcriptional target of FOXO3a, a vital downstream effector of the PI3K-AKT-FOXO signaling cascade. In addition, FOXM1 and FOXO3a also antagonize each other's activity by competitively binding to the same target genes, which are involved in chemotherapeutic drug sensitivity and resistance. Understanding the role and regulation of the FOXO-FOXM1 axis will provide insight into chemotherapeutic drug action and resistance in patients, and help to identify novel therapeutic approaches as well as diagnostic and predictive biomarkers.