Forkhead Box Gene Research Articles

Forkhead transcription factors regulate mosquito reproduction

Forkhead-box (Fox) genes encode a family of transcription factors defined by a ‘winged helix’ DNA-binding domain. In this study we aimed to identify Fox factors that are expressed within the fat body of the yellow fever mosquito Aedes aegypti, and determine whether any of these are involved in the regulation of mosquito yolk protein gene expression. The Ae. aegypti genome contains 18 loci that encode putative Fox factors. Our stringent cladistic analysis has profound implications for the use of Fox genes as phylogenetic markers. Twelve Ae. aegypti Fox genes are expressed within various tissues of adult females, six of which are expressed within the fat body. All six Fox genes expressed in the fat body displayed dynamic expression profiles following a blood meal. We knocked down the ‘fat body Foxes’ through RNAi to determine whether these ‘knockdowns’ hindered amino acid-induced vitellogenin gene expression. We also determined the effect of these knockdowns on the number of eggs deposited following a blood meal. Knockdown of FoxN1, FoxN2, FoxL, and FoxO, had a negative effect on amino acid-induced vitellogenin gene expression and resulted in significantly fewer eggs laid. Our analysis stresses the importance of Fox transcription factors in regulating mosquito reproduction.

The Forkhead Transcription Factor FoxO1 Regulates Proliferation and Transdifferentiation of Hepatic Stellate Cells

The Forkhead box gene, group O (FoxO) family of Forkhead transcription factors is phopsphorylated and inactivated by the phosphatidylinositol 3-kinase (PI3K)/AKT pathway and regulates a variety of cellular functions. Hepatic stellate cells (HSCs) play a crucial role in liver fibrosis. A fibrotic stimulus causes HSCs to transdifferentiate from a quiescent phenotype to a collagen-producing myofibroblast-like phenotype and to proliferate. Mutation/deletion mutants of FoxO1 were introduced into primary rat, mouse, and immortalized human HSCs and assessed for activation, proliferation, and signal transduction. The role of FoxO1 in experimental liver fibrosis was assessed in FoxO1(+/-) and FoxO1(+/+) mice. Platelet-derived growth factor (PDGF) or insulin phosphorylates FoxO1 and induces FoxO1 translocation from the nuclei to the cytosol via the PI3K/AKT pathway in HSCs. Constitutively active FoxO1 inhibits proliferation via cell cycle arrest at the G1 phase, whereas dominant-negative FoxO1 enhances proliferation of HSCs even in the presence of the PI3K inhibitor LY294002. In addition, the phosphorylation of FoxO1 is increased during transdifferentiation of HSCs. The transdifferentiation is also inhibited by constitutively active FoxO1 and is accelerated by dominant-negative FoxO1. FoxO1 directly induces the expression of p27(kip1) and manganese superoxide dismutase (MnSOD). After bile duct ligation for 3 weeks, FoxO1(+/-) mice are more susceptible to liver fibrosis, consistent with our in vitro results. FoxO1 plays a crucial role in the transdifferentiation and proliferation of HSCs in liver fibrosis. Hyperinsulinemia inactivates FoxO1 in HSCs, resulting in HSC activation and may result in the fibrosis in nonalcoholic fatty liver disease.

Modulation of Androgen Receptor Transactivation by FoxH1. A Newly Identified Androgen Receptor Corepressor

Androgen signaling plays key roles in the development and progression of prostate cancer, and numerous ongoing studies focus on the regulation of androgen receptor (AR) transactivity to develop novel therapies for the treatment of androgen-independent prostate cancer. FoxH1, a member of the Forkhead-box (FOX) gene family of transcription factors, takes part in mediating transforming growth factor-β/activin signaling through its interaction with the Smad2·Smad4 complex. Using a series of experiments, we found that FoxH1 repressed both ligand-dependent and -independent transactivation of the AR on androgen-induced promoters. This action of FoxH1 was independent of its transactivation capacity and activin A but relieved by Smad2·Smad4. In addition, the repression of the AR by FoxH1 did not require deacetylase activity. A protein-protein interaction was identified between the AR and FoxH1 independently of dihydrotestosterone. Furthermore, a confocal microscopic analysis of LNCaP cells revealed that the interaction between the AR and FoxH1 occurred in the nucleus and that FoxH1 specifically blocked the foci formation of dihydrotestosterone-activated AR, which has been shown to be correlated with the AR transactivation potential. Taken together, our results indicate that FoxH1 functions as a new corepressor of the AR. Our observations not only strengthen the role of FoxH1 in AR-mediated transactivation but also suggest that therapeutic interventions based on AR-coregulator interactions could be designed to block both androgen-dependent and -independent growth of prostate cancer.

Modulation of Androgen Receptor Transactivation by FoxH1: A NEWLY IDENTIFIED ANDROGEN RECEPTOR COREPRESSOR

Androgen signaling plays key roles in the development and progression of prostate cancer, and numerous ongoing studies focus on the regulation of androgen receptor (AR) transactivity to develop novel therapies for the treatment of androgen-independent prostate cancer. FoxH1, a member of the Forkhead-box (FOX) gene family of transcription factors, takes part in mediating transforming growth factor-beta/activin signaling through its interaction with the Smad2.Smad4 complex. Using a series of experiments, we found that FoxH1 repressed both ligand-dependent and -independent transactivation of the AR on androgen-induced promoters. This action of FoxH1 was independent of its transactivation capacity and activin A but relieved by Smad2.Smad4. In addition, the repression of the AR by FoxH1 did not require deacetylase activity. A protein-protein interaction was identified between the AR and FoxH1 independently of dihydrotestosterone. Furthermore, a confocal microscopic analysis of LNCaP cells revealed that the interaction between the AR and FoxH1 occurred in the nucleus and that FoxH1 specifically blocked the foci formation of dihydrotestosterone-activated AR, which has been shown to be correlated with the AR transactivation potential. Taken together, our results indicate that FoxH1 functions as a new corepressor of the AR. Our observations not only strengthen the role of FoxH1 in AR-mediated transactivation but also suggest that therapeutic interventions based on AR-coregulator interactions could be designed to block both androgen-dependent and -independent growth of prostate cancer.

Human FOX gene family (Review)

Human Forkhead-box (FOX) gene family consists of at least 43 members, including FOXA1, FOXA2, FOXA3, FOXB1, FOXC1, FOXC2, FOXD1, FOXD2, FOXD3, FOXD4, FOXD5 (FOXD4L1), FOXD6 (FOXD4L3), FOXE1, FOXE2, FOXE3, FOXF1, FOXF2, FOXG1 (FOXG1B), FOXH1, FOXI1, FOXJ1, FOXJ2, FOXJ3, FOXK1, FOXK2, FOXL1, FOXL2, FOXM1, FOXN1, FOXN2 (HTLF), FOXN3 (CHES1), FOXN4, FOXN5 (FOXR1), FOXN6 (FOXR2), FOXO1 (FOXO1A), FOXO2 (FOXO6), FOXO3 (FOXO3A), FOXO4 (MLLT7), FOXP1, FOXP2, FOXP3, FOXP4, and FOXQ1. FOXE3-FOXD2 (1p33), FOXQ1-FOXF2-FOXC1 (6p25.3), and FOXF1-FOXC2-FOXL1 (16q24.1) loci are FOX gene clusters within the human genome. Members of FOX subfamilies A-G, I-L and Q were grouped into class 1 FOX proteins, while members of FOX subfamilies H and M-P were grouped into class 2 FOX proteins. C-terminal basic region within the FOX domain was the common feature of class 1 FOX proteins. FOXH1 and FOXO1 mRNAs are expressed in human embryonic stem (ES) cells. FOXC1, FOXC2, FOXE1, FOXE3, FOXL2, FOXN1, FOXP2 and FOXP3 genes are mutated in human congenital disorders. FOXA1 gene is amplified and over-expressed in esophageal and lung cancer. FOXM1 gene is up-regulated in pancreatic cancer and basal cell carcinoma due to the transcriptional regulation by Sonic Hedgehog (SHH) pathway. FOXO1 gene is fused to PAX3 or PAX7 genes in rhabdomyosarcoma. FOXO3 and FOXO4 genes are fused to MLL gene in hematological malignancies. Deregulation of FOX family genes leads to congenital disorders, diabetes mellitus, or carcinogenesis. Expression profiles, genetic alterations and epigenetic changes of FOX family genes as well as binding proteins and target genes of FOX family transcription factors should be comprehensively investigated to develop novel therapeutics and preventives for human diseases.

Profiling the molecular difference between Patched- and p53-dependent rhabdomyosarcoma.

Rhabdomyosarcoma (RMS) is a highly malignant tumor that is histologically related to skeletal muscle, yet genetic and molecular lesions underlying its genesis and progression remain largely unknown. In this study we have compared the molecular profiles of two different mouse models of RMS, each associated with a defined primary genetic defect known to play a role in rhabdomyosarcomagenesis in man. We report that RMS of heterozygous Patched1 (Ptch1) mice show less aggressive growth and a greater degree of differentiation than RMS of heterozygous p53 mice. By means of cDNA microarray analysis we demonstrate that RMS in Ptch1 mutants predominantly express a number of myogenic markers, including myogenic differentiation 1, myosin heavy chain, actin, troponin and tropomyosin, as well as genes associated with Hedgehog/Patched signaling like insulin-like growth factor 2, forkhead box gene Foxf1 and the growth arrest and DNA-damage-inducible gene Gadd45a. In sharp contrast, RMS in p53 mutants display higher expression levels of cell cycle-associated genes like cyclin B1, cyclin-dependent kinase 4 and the proliferation marker Ki-67. These results demonstrate that different causative mutations lead to distinct gene expression profiles in RMS, which appear to reflect their different biological characteristics. Our results provide a first step towards a molecular classification of different forms of RMS. If the described differences can be confirmed in human RMS our results will contribute to a new molecular taxonomy of this cancer, which will be critical for gene mutation- and expression-specific therapy.

Identification and characterization of a novel human FOXK1 gene in silico

We describe here the identification and characterization of a human forkhead box gene, FOXK1, that encodes predicted proteins most homologous to the mouse myocyte nuclear factor (MNF)/forkhead box K1 (FoxK1). Human FOXK1 is located at the chromosomal position 7p22. Two transcript variants, FOXK1a and FOXK1b, containing the divergent 5'-ends are identified. Sequence comparison indicated that the human FOXK1 proteins share 66-74% amino acid identity (99% identity in the forkhead domain) with their mouse ortholog. The in silico expression analysis showed FOXK1 expression in immature tissues of brain, eye, heart, lung and thymus. In adults, FOXK1 was predominantly expressed in many malignant tissues, in particular in the tumors of brain, colon and lymph node. This gene is therefore implicated in the process of both normal and neoplastic development.

Identification and characterization of human FOXN5 and rat Foxn5 genes in silico

Forkhead-box (FOX) genes are implicated in embryogenesis through transcriptional regulation depending on SHH-GLI pathway, TGF-beta pathway etc., and also in carcinogenesis through gene amplification, retroviral integration and chromosomal translocation. FOXN1, FOXN2 (HTLF), FOXN3 (CHES1) and FOXN4 constitute the FOXN family. Here, we identified and characterized the FOXN5 gene, a novel member of FOXN gene family, by using bioinformatics. IMAGE5167039 (BC028191.1) was the representative cDNA derived from human FOXN5 gene. Rat Foxn5 gene, consisting of six exons, was identified within rat genome sequence CH230-26K11 (AC107575.5). Complete coding sequence of rat Foxn5 cDNA was determined by assembling nucleotide sequences of rat Foxn5 exons. Human FOXN5 (292 aa) and rat Foxn5 (296 aa) showed 77.4% total-amino-acid identity. Codon 173-254 of FOXN5 was the Forkhead domain. FOXN5 gene, consisting of six exons, was linked to BCL9L gene at human chromosome 11q23.3. FOXN5 is a candidate tumor suppressor gene (TSG), just like ARHGAP20 (KIAA1391), BTG4, SNF1LK2 (SIK2), DIXDC1 (KIAA1735) genes at 11q23.1, TTC12 (TPARM) gene at 11q23.2, IGSF4, DSCAML1, LL5A (PHLDB1), BCL9L, RNF26, and MFRP genes at 11q23.3. This is the first report on the human FOXN5 and rat Foxn5 genes.

Regulation of insulin-like growth factor-dependent myoblast differentiation by Foxo forkhead transcription factors.

Insulin-like growth factors promote myoblast differentiation through phosphoinositol 3-kinase and Akt signaling. Akt substrates required for myogenic differentiation are unknown. Forkhead transcription factors of the forkhead box gene, group O (Foxo) subfamily are phosphorylated in an insulin-responsive manner by phosphatidylinositol 3-kinase–dependent kinases. Phosphorylation leads to nuclear exclusion and inactivation. We show that a constitutively active Foxo1 mutant inhibits differentiation of C2C12 cells and prevents myotube differentiation induced by constitutively active Akt. In contrast, a transcriptionally inactive mutant Foxo1 partially rescues inhibition of C2C12 differentiation mediated by wortmannin, but not by rapamycin, and is able to induce aggregation-independent myogenic conversion of teratocarcinoma cells. Inhibition of Foxo expression by siRNA resulted in more efficient differentiation, associated with increased myosin expression. These observations indicate that Foxo proteins are key effectors of Akt-dependent myogenesis.

Predicted Protein Structure of MedakaFoxA3and Its Expression in Polster

The forkhead box genes (Fox genes) are expressed in many tissues during embryogenesis and their products play a key role in organogenesis. We cloned two Fox genes from an embryonic cDNA library of medaka (Oryzias latipes). One was MeHNF3β, a homologue of HNF3β (FoxA2). The other was Mefkh1 (medaka-forkhead1), related to HNF3γ (FoxA3). We found that the expression pattern of FoxA3 differs in some aspects between medaka and zebrafish. Mefkh1 is expressed in the dorsal marginal zone, embryonic shield, polster, gut and slightly axial mesoderm, whereas zebrafish FoxA3 is expressed in the yolk syncytial layer (YSL), axial mesoderm, polster, gut and posterior neural crest cells (Odenthal and Nüsslein-Volhard, 1998, Dev Genes Evol 208, 245–258). In zebrafish, the level of expression of FoxA3 in the axial mesoderm is similar to that of FoxA2 (axial); in medaka, FoxA3 (Mefkh1) expression is mainly detected in the polster, and FoxA2 (MeHNF3β) is expressed in the axial mesoderm. This indicates that the combined functioning of FoxA2 and FoxA3 in the organogenesis of the polster and axial mesoderm differs between medaka and zebrafish.