Forkhead Box O Transcription Factor Research Articles

Targeting cis-regulatory elements of FOXO family is a novel therapeutic strategy for induction of leukemia cell differentiation

Differentiation therapy has been proposed as a promising therapeutic strategy for acute myeloid leukemia (AML); thus, the development of more versatile methodologies that are applicable to a wide range of AML subtypes is desired. Although the FOXOs transcription factor represents a promising drug target for differentiation therapy, the efficacy of FOXO inhibitors is limited in vivo. Here, we show that pharmacological inhibition of a common cis-regulatory element of forkhead box O (FOXO) family members successfully induced cell differentiation in various AML cell lines. Through gene expression profiling and differentiation marker-based CRISPR/Cas9 screening, we identified TRIB1, a complement of the COP1 ubiquitin ligase complex, as a functional FOXO downstream gene maintaining an undifferentiated status. TRIB1 is direct target of FOXO3 and the FOXO-binding cis-regulatory element in the TRIB1 promoter, referred to as the FOXO-responsive element in the TRIB1 promoter (FRE-T), played a critical role in differentiation blockade. Thus, we designed a DNA-binding pharmacological inhibitor of the FOXO-FRE-T interface using pyrrole-imidazole polyamides (PIPs) that specifically bind to FRE-T (FRE-PIPs). The FRE-PIPs conjugated to chlorambucil (FRE-chb) inhibited transcription of TRIB1, causing differentiation in various AML cell lines. FRE-chb suppressed the formation of colonies derived from AML cell lines but not from normal counterparts. Administration of FRE-chb inhibited tumor progression in vivo without remarkable adverse effects. In conclusion, targeting cis-regulatory elements of the FOXO family is a promising therapeutic strategy that induces AML cell differentiation.

Stress activated signalling impaired protein quality control pathways in human hypertrophic cardiomyopathy

Hypertrophic cardiomyopathy (HCM) is a complex myocardial disorder with no well-established disease-modifying therapy so far. Our study aimed to investigate how autophagy, oxidative stress, inflammation, stress signalling pathways, and apoptosis are hallmark of HCM and their contribution to the cardiac dysfunction. Demembranated cardiomyocytes from patients with HCM display increased titin-based stiffness (Fpassive), which was corrected upon antioxidant treatment. Titin as a main determinant of Fpassive was S-glutathionylated and highly ubiquitinated in HCM patients. This was associated with a shift in the balance of reduced and oxidized forms of glutathione (GSH and GSSG, respectively). Both heat shock proteins (HSP27 and α-ß crystalline) were upregulated and S-glutathionylated in HCM. Administration of HSPs in vitro significantly reduced HCM cardiomyocyte stiffness. High levels of the phosphorylated monomeric superoxide anion-generating endothelial nitric oxide synthase (eNOS), decreased nitric oxide (NO) bioavailability, decreased soluble guanylyl cyclase (sGC) activity, and high levels of 3-nitrotyrosine were observed in HCM. Many regulators of signal transduction pathways that are involved in autophagy, apoptosis, cardiac contractility, and growth including the mitogen-activated protein kinase (MAPK), protein kinase B (AKT), glycogen synthase kinase 3ß (GSK-3ß), mammalian target of rapamycin (mTOR), forkhead box O transcription factor (FOXO), c-Jun N-terminal protein kinase (JNK), and extracellular-signal-regulated kinase (ERK1/2) were modified in HCM. The apoptotic factors cathepsin, procaspase 3, procaspase 9 and caspase 12, but not caspase 9, were elevated in HCM hearts and associated with increased proinflammatory cytokines (Interleukin 6 (IL-6), interleukin 18 (IL-18), intercellular cell adhesion molecule-1 (ICAM1), vascular cell adhesion molecule-1 (VCAM1), the Toll-like receptors 2 (TLR2) and the Toll-like receptors 4 (TLR4)) and oxidative stress (3-nitrotyrosine and hydrogen peroxide (H2O2)). Here we reveal stress signalling and impaired PQS as potential mechanisms underlying the HCM phenotype. Our data suggest that reducing oxidative stress can be a viable therapeutic approach to attenuating the severity of cardiac dysfunction in heart failure and potentially in HCM and prevent its progression.

Dexmedetomidine ameliorates lipopolysaccharide-induced acute lung injury by inhibiting the PI3K/Akt/FoxO1 signaling pathway.

PurposeDexmedetomidine (DEX) has been associated with inflammation, oxidative stress, and apoptosis, but its effects on lipopolysaccharide (LPS)-induced lung injury remain uncertain. The present study explored the effects of DEX on LPS-induced lung injury and studied the possible molecular mechanisms by testing the effects of the phosphoinositide-3 kinase (PI3K) inhibitor LY294002 and BEZ235.MethodsSeventy C57BL/6 mice were randomly divided into the control, LPS, LPS + DEX, LPS + LY294002, LPS + BEZ235, LPS + DEX + LY294002, and LPS + DEX + BEZ235groups. Lung samples were collected 48 h after LPS treatment.ResultsDEX significantly inhibited LPS-induced increases in the lung weight/body weight ratio and lung wet/dry weight ratio, decreased inflammatory cell infiltration, and decreased the production of proinflammatory factors, such as interleukin-1β (IL-1β), IL-6, and tumor necrosis factor α (TNF-α)in the lungs. DEX also markedly attenuated the increases in malondialdehyde 5 (MDA 5) and inositol-dependent enzyme a (IRE-a), attenuated the decrease in superoxide dismutase 1(SOD-1), reversed the low expression of B-cell lymphoma-2 (Bcl-2), and the high expressions of Bax and Caspase-3. DEX also decreased the expression of phosphorylated PI3K and phosphorylated Akt and increased the expression of phosphorylated forkhead box-O transcription factor 1 (FoxO1). More interestingly, LY294002 or BEZ235 pretreatment significantly abolished the inhibitory effects of DEX on LPS-induced lung inflammation, oxidative stress, and apoptosis.ConclusionsThese data suggest that DEX ameliorates LPS-induced acute lung injury partly through the PI3K/Akt/FoxO1 signaling pathway.

DMSO Delays Alzheimer Disease Causing Aβ-induced Paralysis in C. elegans Through Modulation of Glutamate/Acetylcholine Neurotransmission.

Background:Alzheimer’s disease (AD), a prevalent neurodegenerative disease with progressive dementia and neurotransmission (NT)-dysfunction-related complications in older adults, is known to be caused by abnormal Amyloid-β (Aβ) peptide and associated amyloid plaques in the brain. Drugs to cure AD are not in sight. Two major excitatory neurotransmitters, glutamate (Glu) and acetylcholine (ACh), and their signaling systems are implicated in AD.Objective:To determine the effect of various NT-altering compounds including fenobam, quisqualic acid, and dimethyl sulfoxide (DMSO) in the protection against Aβ toxicity. Further, to identify the potential mechanism through which the protection happens.Methods:The well-known C. elegans AD model, CL4176, in which human Aβ expression is turned on upon a temperature shift to 25 °C that leads to paralysis, was screened for protection/delay in paralysis because of Αβ toxicity. While screening the compounds, dimethyl sulfoxide (DMSO), a universal solvent used to solubilize compounds, was identified to provide protection. Aldicarb and levamisole assays were performed to identify the contribution of ACh neurotransmission in Αβ toxicity protection by DMSO.Results:One percent and two percent DMSO delayed paralysis by 48% and 90%, respectively. DMSO was dominant over one of the Glu-NT pathway-related compounds, Fenobam-Group I mGluR antagonist. But DMSO provided only 30% to 50% protection against Quisqualic acid, the Glu-agonist. DMSO (2%) delayed ACh-NT, both presynaptic acetylcholine esterase inhibitor (AchEi)-aldicarb and postsynaptic-iAChR-agonst-levamisole induced paralysis, by ∼70% in CL4176. DMSO seems to be altering Ca2+ ion permeability essential for NT as EthyleneDiamine Tetra-Acetic acid (EDTA) and DMSO provided similar aldicarb resistance either combined or alone in wildtype worms. But postsynaptic Ca2+ depletion by EDTA could reverse DMSO-induced levamisole hypersensitivity. Surprisingly, the absence of FOrkhead boXO (FOXO) transcription factor homolog, daf-16 (loss-of-function mutant), a critical transcription factor in the reduced IIS-mediated longevity in C. elegans, abolished DMSO-mediated AldR.Conclusion:DMSO and Fenobam protect against Aβ toxicity through modulation of NT.

Assessment of Functional Phosphatidylinositol 3-Kinase Pathway Activity in Cancer Tissue Using Forkhead Box-O Target Gene Expression in a Knowledge-Based Computational Model

The phosphatidylinositol 3-kinase (PI3K) pathway is commonly activated in cancer. Tumors are potentially sensitive to PI3K pathway inhibitors, but reliable diagnostic tests that assess functional PI3K activity are lacking. Because PI3K pathway activity negatively regulates forkhead box-O (FOXO) transcription factor activity, FOXO target gene expression is inversely correlated with PI3K activity. Aknowledge-based Bayesian computational model was developed to infer PI3K activity in cancer tissue samples from FOXO target gene mRNA levels and validated in cancer cell lines treated with PI3K inhibitors. However, applied to patient tissue samples, FOXO was often active in cancer types with expected active PI3K. SOD2 was differentially expressed between FOXO-active healthy and cancer tissue samples, indicating that cancer-associated cellular oxidative stress alternatively activated FOXO. To enable correct interpretation of active FOXO in cancer tissue, threshold levels for normal SOD2 expression in healthy tissue were defined above which FOXO activity is oxidative stress induced and below which PI3K regulated. In slow-growing luminal A breast cancer and low Gleason score prostate cancer, FOXO was active in a PI3K-regulated manner, indicating inactive PI3K. In aggressive luminal B, HER2, and basal breast cancer, FOXO was increasingly inactive or actively induced by oxidative stress, indicating PI3K activity. We provide a decision tree that facilitates functional PI3K pathway activity assessment in tissue samples from patients with cancer for therapy response prediction and prognosis.

The FOXO transcription factor controls insect growth and development by regulating juvenile hormone degradation in the silkworm, Bombyx mori

Forkhead box O (FOXO) functions as the terminal transcription factor of the insulin signaling pathway and regulates multiple physiological processes in many organisms, including lifespan in insects. However, how FOXO interacts with hormone signaling to modulate insect growth and development is largely unknown. Here, using the transgene-based CRISPR/Cas9 system, we generated and characterized mutants of the silkworm Bombyx mori FOXO (BmFOXO) to elucidate its physiological functions during development of this lepidopteran insect. The BmFOXO mutant (FOXO-M) exhibited growth delays from the first larval stage and showed precocious metamorphosis, pupating at the end of the fourth instar (trimolter) rather than at the end of the fifth instar as in the wild-type (WT) animals. However, different from previous reports on precocious metamorphosis caused by juvenile hormone (JH) deficiency in silkworm mutants, the total developmental time of the larval period in the FOXO-M was comparable with that of the WT. Exogenous application of 20-hydroxyecdysone (20E) or of the JH analog rescued the trimolter phenotype. RNA-seq and gene expression analyses indicated that genes involved in JH degradation but not in JH biosynthesis were up-regulated in the FOXO-M compared with the WT animals. Moreover, we identified several FOXO-binding sites in the promoter of genes coding for JH-degradation enzymes. These results suggest that FOXO regulates JH degradation rather than its biosynthesis, which further modulates hormone homeostasis to control growth and development in B. mori In conclusion, we have uncovered a pivotal role for FOXO in regulating JH signaling to control insect development.

Ketone bodies mimic the life span extending properties of caloric restriction.

The extension of life span by caloric restriction has been studied across species from yeast and Caenorhabditis elegans to primates. No generally accepted theory has been proposed to explain these observations. Here, we propose that the life span extension produced by caloric restriction can be duplicated by the metabolic changes induced by ketosis. From nematodes to mice, extension of life span results from decreased signaling through the insulin/insulin-like growth factor receptor signaling (IIS) pathway. Decreased IIS diminishes phosphatidylinositol (3,4,5) triphosphate (PIP3 ) production, leading to reduced PI3K and AKT kinase activity and decreased forkhead box O transcription factor (FOXO) phosphorylation, allowing FOXO proteins to remain in the nucleus. In the nucleus, FOXO proteins increase the transcription of genes encoding antioxidant enzymes, including superoxide dismutase 2, catalase, glutathione peroxidase, and hundreds of other genes. An effective method for combating free radical damage occurs through the metabolism of ketone bodies, ketosis being the characteristic physiological change brought about by caloric restriction from fruit flies to primates. A dietary ketone ester also decreases circulating glucose and insulin leading to decreased IIS. The ketone body, d-β-hydroxybutyrate (d-βHB), is a natural inhibitor of class I and IIa histone deacetylases that repress transcription of the FOXO3a gene. Therefore, ketosis results in transcription of the enzymes of the antioxidant pathways. In addition, the metabolism of ketone bodies results in a more negative redox potential of the NADP antioxidant system, which is a terminal destructor of oxygen free radicals. Addition of d-βHB to cultures of C. elegans extends life span. We hypothesize that increasing the levels of ketone bodies will also extend the life span of humans and that calorie restriction extends life span at least in part through increasing the levels of ketone bodies. An exogenous ketone ester provides a new tool for mimicking the effects of caloric restriction that can be used in future research. The ability to power mitochondria in aged individuals that have limited ability to oxidize glucose metabolites due to pyruvate dehydrogenase inhibition suggests new lines of research for preventative measures and treatments for aging and aging-related disorders. © 2017 The Authors IUBMB Life published by Wiley Periodicals, Inc. on behalf of International Union of Biochemistry and Molecular Biology, 69(5):305-314, 2017.

The impact of FOXO on dopamine and octopamine metabolism in Drosophila under normal and heat stress conditions.

ABSTRACTThe forkhead boxO transcription factor (FOXO) is a component of the insulin signalling pathway and plays a role in responding to adverse conditions, such as oxidative stress and starvation. In stressful conditions, FOXO moves from the cytosol to the nucleus where it activates gene expression programmes. Here, we show that FOXO in Drosophila melanogaster responds to heat stress as it does to other stressors. The catecholamine signalling pathway is another component of the stress response. In Drosophila, dopamine and octopamine levels rise steeply under heat, nutrition and mechanical stresses, which are followed by a decrease in the activity of synthesis enzymes. We demonstrate that the nearly twofold decline of FOXO expression in foxoBG01018 mutants results in dramatic changes in the metabolism of dopamine and octopamine and the overall response to stress. The absence of FOXO increases tyrosine decarboxylase activity, the first enzyme in octopamine synthesis, and decreases the enzymatic activity of enzymes in dopamine synthesis, alkaline phosphatase and tyrosine hydroxylase, in young Drosophila females. We identified the juvenile hormone as a mediator of FOXO regulation of catecholamine metabolism. Our findings suggest that FOXO is a possible trigger for endocrinological stress reactions.

Repeated Glucose Deprivation/Reperfusion Induced PC-12 Cell Death through the Involvement of FOXO Transcription Factor

BackgroundCognitive impairment and brain damage in diabetes is suggested to be associated with hypoglycemia. The mechanisms of hypoglycemia-induced neural death and apoptosis are not clear and reperfusion injury may be involved. Recent studies show that glucose deprivation/reperfusion induced more neuronal cell death than glucose deprivation itself. The forkhead box O (FOXO) transcription factors are implicated in the regulation of cell apoptosis and survival, but their role in neuronal cells remains unclear. We examined the role of FOXO transcription factors and the involvement of the phosphatidylinositol 3-kinase (PI3K)/Akt and apoptosis-related signaling pathways in PC-12 cells exposed to repeated glucose deprivation/reperfusion.MethodsPC-12 cells were exposed to control (Dulbecco's Modified Eagle Medium [DMEM] containing 25 mM glucose) or glucose deprivation/reperfusion (DMEM with 0 mM glucose for 6 hours and then DMEM with 25 mM glucose for 18 hours) for 5 days. MTT assay and Western blot analysis were performed for cell viability, apoptosis, and the expression of survival signaling pathways. FOXO3/4',6-diamidino-2-phenylindole staining was done to ascertain the involvement of FOXO transcription factors in glucose deprivation/reperfusion conditions.ResultsCompared to PC-12 cells not exposed to hypoglycemia, cells exposed to glucose deprivation/reperfusion showed a reduction of cell viability, decreased expression of phosphorylated Akt and Bcl-2, and an increase of cleaved caspase-3 expression. Of note, FOXO3 protein was localized in the nuclei of glucose deprivation/reperfusion cells but not in the control cells.ConclusionRepeated glucose deprivation/reperfusion caused the neuronal cell death. Activated FOXO3 via the PI3K/Akt pathway in repeated glucose deprivation/reperfusion was involved in genes related to apoptosis.

RNA helicase HEL-1 promotes longevity by specifically activating DAF-16/FOXO transcription factor signaling in Caenorhabditis elegans

The homeostatic maintenance of the genomic DNA is crucial for regulating aging processes. However, the role of RNA homeostasis in aging processes remains unknown. RNA helicases are a large family of enzymes that regulate the biogenesis and homeostasis of RNA. However, the functional significance of RNA helicases in aging has not been explored. Here, we report that a large fraction of RNA helicases regulate the lifespan of Caenorhabditis elegans. In particular, we show that a DEAD-box RNA helicase, helicase 1 (HEL-1), promotes longevity by specifically activating the DAF-16/forkhead box O (FOXO) transcription factor signaling pathway. We find that HEL-1 is required for the longevity conferred by reduced insulin/insulin-like growth factor 1 (IGF-1) signaling (IIS) and is sufficient for extending lifespan. We further show that the expression of HEL-1 in the intestine and neurons contributes to longevity. HEL-1 enhances the induction of a large fraction of DAF-16 target genes. Thus, the RNA helicase HEL-1 appears to promote longevity in response to decreased IIS as a transcription coregulator of DAF-16. Because HEL-1 and IIS are evolutionarily well conserved, a similar mechanism for longevity regulation via an RNA helicase-dependent regulation of FOXO signaling may operate in mammals, including humans.

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.

Inhibition of polyglutamine-mediated proteotoxicity by Astragalus membranaceus polysaccharide through the DAF-16/FOXO transcription factor in Caenorhabditis elegans

Late-onset neurodegenerative diseases are characterized by progressive accumulation of aggregation-prone proteins and global disruption of the proteostasis network, e.g. abnormal polyQ (polyglutamine) aggregation in Huntington's disease. Astragalus membranaceus polysaccharide (astragalan) has recently been shown to modulate aging and proteotoxic stress pathways. Using Caenorhabditis elegans models, we now show that astragalan not only reduces polyQ aggregation, but also alleviates the associated neurotoxicity. We also reveal that astragalan can extend the adult lifespan of wild-type and polyQ nematodes, indicating a connection of its anti-aging benefit with the toxicity-suppressing effect. Further examination demonstrates that astragalan can extend the lifespan of daf-2 and age-1, but not daf-16, mutant nematodes of the insulin-like aging and stress pathway, suggesting a lifespan-regulation signalling independent of DAF (abnormal dauer formation)-2/IGF-1R (insulin-like growth factor 1 receptor), but dependent on the DAF-16/FOXO (forkhead box O) transcription factor, a pivotal integrator of divergent signalling pathways related to both lifespan regulation and stress resistance. We also show that a subset of DAF-16 downstream genes are regulated by astragalan, including the DAF-16 transcriptional target gene scl-20, which is itself constitutively up-regulated in transgenic polyQ nematodes. These findings, together with our previous work on LEA (late embryogenesis abundant) proteins and trehalose, provide a revealing insight into the potential of stress and lifespan regulators in the prevention of proteotoxic disorders.

DAF-16/Forkhead Box O Transcription Factor: Many Paths to a Single Fork(head) in the Road

The Caenorhabditis elegans Forkhead box O transcription factor (FOXO) homolog DAF-16 functions as a central mediator of multiple biological processes such as longevity, development, fat storage, stress resistance, and reproduction. In C. elegans, similar to other systems, DAF-16 functions as the downstream target of a conserved, well-characterized insulin/insulin-like growth factor (IGF)-1 signaling pathway. This cascade is comprised of an insulin/IGF-1 receptor, which signals through a conserved PI 3-kinase/AKT pathway that ultimately downregulates DAF-16/FOXO activity. Importantly, studies have shown that multiple pathways intersect with the insulin/IGF-1 signaling pathway and impinge on DAF-16 for their regulation. Therefore, in C. elegans, the single FOXO family member, DAF-16, integrates signals from several pathways and then regulates its many downstream target genes.

Muscle-Specific F-Box Only Proteins Facilitate BK Channel β 1 Subunit Downregulation in Vascular Smooth Muscle Cells of Diabetes Mellitus

activity of the large conductance Ca(2+)-activated K(+) (BK) channels is profoundly modulated by its β(1) subunit (BK-β(1)). However, BK-β(1) expression is downregulated in diabetic vessels. The ubiquitin-proteasome system (UPS) is a major mechanism of intracellular protein degradation. Whether UPS participates in BK-β(1) downregulation in diabetic vessels is unknown. we hypothesize that UPS facilitates vascular BK-β(1) degradation in diabetes. using patch clamp and molecular biological approaches, we found that BK-β(1)-mediated channel activation and BK-β(1) protein expression were reduced in aortas of streptozotocin-induced diabetic rats and in human coronary arterial smooth muscle cells (CASMCs) cultured in high glucose. This was accompanied by upregulation of F-box only protein (FBXO)-9 and FBXO-32 (atrogin-1), the key components of the Skp1-Cullin-F-box (SCF) type ubiquitin ligase complex. BK-β(1) expression was suppressed by the FBXO activator doxorubicin but enhanced by FBXO-9 small interfering RNA or by the proteasome inhibitor MG-132. Cotransfection of atrogin-1 in HEK293 cells significantly reduced Flag-hSlo-β(1) expression by 2.16-fold, compared with expression of Flag-hSlo-β(1)V146A (a mutant without the PDZ-binding motif). After cotransfection with atrogin-1, the ubiquitination of Flag-hSlo-β(1) was increased by 1.91-fold, compared with that of hSlo-β(1)V146A, whereas cotransfection with atrogin-1ΔF (a nonfunctional mutant without the F-box motif) had no effect. Moreover, inhibition of Akt signaling attenuated the phosphorylation of forkhead box O transcription factor (FOXO)-3a and enhanced atrogin-1 expression, which in turn suppressed BK-β(1) protein levels in human CASMCs. downregulation of vascular BK-β(1) expression in diabetes and in high-glucose culture conditions was associated with FOXO-3a/FBXO-dependent increase in BK-β(1) degradation.

Induction of Manganese Superoxide Dismutase by Nuclear Translocation and Activation of SIRT1 Promotes Cell Survival in Chronic Heart Failure

Oxidative stress plays a pivotal role in chronic heart failure. SIRT1, an NAD(+)-dependent histone/protein deacetylase, promotes cell survival under oxidative stress when it is expressed in the nucleus. However, adult cardiomyocytes predominantly express SIRT1 in the cytoplasm, and its function has not been elucidated. The purpose of this study was to investigate the functional role of SIRT1 in the heart and the potential use of SIRT1 in therapy for heart failure. We investigated the subcellular localization of SIRT1 in cardiomyocytes and its impact on cell survival. SIRT1 accumulated in the nucleus of cardiomyocytes in the failing hearts of TO-2 hamsters, postmyocardial infarction rats, and a dilated cardiomyopathy patient but not in control healthy hearts. Nuclear but not cytoplasmic SIRT1-induced manganese superoxide dismutase (Mn-SOD), which was further enhanced by resveratrol, and increased the resistance of C2C12 myoblasts to oxidative stress. Resveratrol's enhancement of Mn-SOD levels depended on the level of nuclear SIRT1, and it suppressed the cell death induced by antimycin A or angiotensin II. The cell-protective effects of nuclear SIRT1 or resveratrol were canceled by the Mn-SOD small interfering RNA or SIRT1 small interfering RNA. The oral administration of resveratrol to TO-2 hamsters increased Mn-SOD levels in cardiomyocytes, suppressed fibrosis, preserved cardiac function, and significantly improved survival. Thus, Mn-SOD induced by resveratrol via nuclear SIRT1 reduced oxidative stress and participated in cardiomyocyte protection. SIRT1 activators such as resveratrol could be novel therapeutic tools for the treatment of chronic heart failure.

Structural and functional characterisation of the fork head transcription factor-encoding gene, Hc-daf-16, from the parasitic nematode Haemonchus contortus (Strongylida)

Despite their phylogenetic diversity, parasitic nematodes share attributes of longevity and developmental arrest (=hypobiosis) with free-living nematodes at key points in their life cycles, particularly in larval stages responsible for establishing infection in the host. Insulin-like signalling plays crucial roles in the regulation of life span and arrest (=dauer formation) in the free-living nematode, Caenorhabditis elegans. Insulin-like signalling in C. elegans negatively regulates the fork head boxO (FoxO) transcription factor encoded by daf-16, which is linked to initiating a dauer-specific pattern of gene expression. Orthologues of daf-16 have been identified in several species of parasitic nematode. Although function has been demonstrated for an orthologue from the parasitic nematode Strongyloides stercoralis (Rhabditida), the functional capabilities of homologues/orthologues in bursate nematodes (Strongylida) are unknown. In the present study, we used a genomic approach to determine the structures of two complete daf-16 orthologues (designated Hc-daf-16.1 and Hc-daf-16.2) and their transcripts in the parasitic nematode Haemonchus contortus, and assessed their function(s) using C. elegans as a genetic surrogate. Unlike the multiple isoforms of Ce-DAF-16 and Ss-DAF-16, which are encoded by a single gene and produced by alternative splicing, mRNAs encoding the proteins Hc-DAF-16.1 and Hc-DAF-16.2 are transcribed from separate and distinct loci. Both orthologues are transcribed in all developmental stages and both sexes of H. contortus, and the inferred proteins (603 and 556 amino acids) each contain a characteristic, highly conserved fork head domain. In spite of distinct differences in genomic organisation compared with orthologues in C. elegans and S. stercoralis, genetic complementation studies demonstrated here that Hc-daf-16.2, but not Hc-daf-16.1, could restore daf-16 function to a C. elegans strain carrying a null mutation at this locus. These findings are consistent with previous results for S. stercoralis and demonstrate functional conservation of the daf-16b orthologue between key parasitic nematodes from two different taxonomic orders and C. elegans. We conclude from these experiments that the fork head transcription factor DAF-16 and, by inference, other insulin-like signalling elements, are conserved in H. contortus, a parasitic nematode of paramount economic importance. We demonstrate that functionality is sufficiently conserved in Hc-DAF-16.2 that it can replace Ce-DAF-16 in promoting dauer arrest in C. elegans.

Stimulation of Movement in a Quiescent, Hibernation-Like Form ofCaenorhabditis elegansby Dopamine Signaling

One of the characteristics of animals in hibernation is reduced behavioral activity. The Caenorhabditis elegans dauer state is a hibernation-like state of diapause that displays a dramatic reduction in spontaneous locomotion. A similar dauer-like quiescent state is produced in adults by relatively strong mutations in the insulin/IGF-1 receptor homolog daf-2. In this study, we show that mutations affecting the neurotransmitter dopamine, which regulates voluntary movement in many organisms, can stimulate movement in dauers and dauer-like quiescent adults. Surprisingly, the movement of quiescent animals is stimulated by conditions that reduce dopamine signaling and also by conditions predicted to increase dopamine signaling. Reducing dopamine signaling is likely to stimulate movement by activating a foraging response also seen in nondauers after withdrawal of food. In contrast, the stimulation of movement by increased dopamine is much more pronounced in quiescent daf-2(-) dauer and dauer-like adult animals than in nondauaer animals. This altered response to dopamine is primarily attributable to activity of the FOXO (forkhead box O) transcription factor DAF-16 in neurons. We suggest that dauers and dauer-like quiescent adults may have underlying changes in the dopamine system that enable them to respond differently to environmental stimulation.

Daily Variations in Dietary Lysine Content Alter the Expression of Genes Related to Proteolysis in Chicken Pectoralis major Muscle

Amino acids are known to be anabolic factors that affect protein metabolism, but the response of animals to daily amino acid changes is little understood. We aimed to test the effects of feeding birds with alternations of diets varying in lysine content on the expression of genes related to proteolysis in chicken muscle. Cyclic feeding programs with 2 diets, each given for 24 h during 48-h cycles, were carried out from 10 d of age. Three programs were used: 1) control treatment with continuous distribution of a complete diet containing standard medium lysine level (ML; 11.9 g/kg); 2) alternation of diets with high (HL) and low (LL) lysine levels; 3) alternation of ML and LL diets, where LL = 70%, ML = 100%, HL = 130% of standard lysine level. The Pectoralis major muscles were sampled after 2 wk of cyclic feeding. Measurements included the expression patterns of 6 genes involved in proteolysis, and mammalian target of rapamycin and Forkhead box-O transcription factor (FoxO) signaling. Cathepsin B, m-calpain, and E3 ubiquitin ligases Muscle Ring Finger-1 and Muscle Atrophy F box were significantly overexpressed in chickens transiently fed the LL diet, whereas the mRNA levels of 20S proteasome C2 subunit and ubiquitin remained unchanged. Modifications of E3 ubiquitin ligase expression can be partly explained by significant changes in FoxO phosphorylation with cyclic dietary treatments. Our results suggest timing-sensitive regulation of proteolysis in chicken muscle according to dietary treatment and a high metabolism capacity to compensate for changes in amino acid supply, which might be used for nutritional purposes.

Forkhead box-O transcription factor: critical conductors of cancer's fate

Cells have evolved elaborated mechanisms to coordinate the cellular answer of either survival or apoptosis. Recent concepts of human carcinogenesis have suggested disturbances in these cellular relays as a potential link to cellular dedifferentiation and uncontrolled proliferation. Forkhead box-O transcription factors (FOXOs) play an important role in tumour suppression by regulating the expression of genes involved in stress resistance, DNA damage repair, cell cycle arrest and apoptosis. The specific regulation of FOXO function is tightly controlled by posttranslational modifications such as phosphorylation, acetylation and ubiquitination. Loss of FOXO function has recently been identified in several human cancers. In this review, we will give an overview about recent progress in the understanding of function and regulation of FOXOs, as well as their role in carcinogenesis. Furthermore, we will discuss a potential clinical use of FOXOs by therapeutically restoring their tumour suppressive properties.