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1.
Nat Commun ; 11(1): 4111, 2020 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-32807776

RESUMO

Mutational inactivation of VHL is the earliest genetic event in the majority of clear cell renal cell carcinomas (ccRCC), leading to accumulation of the HIF-1α and HIF-2α transcription factors. While correlative studies of human ccRCC and functional studies using human ccRCC cell lines have implicated HIF-1α as an inhibitor and HIF-2α as a promoter of aggressive tumour behaviours, their roles in tumour onset have not been functionally addressed. Herein we show using an autochthonous ccRCC model that Hif1a is essential for tumour formation whereas Hif2a deletion has only minor effects on tumour initiation and growth. Both HIF-1α and HIF-2α are required for the clear cell phenotype. Transcriptomic and proteomic analyses reveal that HIF-1α regulates glycolysis while HIF-2α regulates genes associated with lipoprotein metabolism, ribosome biogenesis and E2F and MYC transcriptional activities. HIF-2α-deficient tumours are characterised by increased antigen presentation, interferon signalling and CD8+ T cell infiltration and activation. Single copy loss of HIF1A or high levels of HIF2A mRNA expression correlate with altered immune microenvironments in human ccRCC. These studies reveal an oncogenic role of HIF-1α in ccRCC initiation and suggest that alterations in the balance of HIF-1α and HIF-2α activities can affect different aspects of ccRCC biology and disease aggressiveness.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Renais/metabolismo , Neoplasias Renais/patologia , Células 3T3 , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Western Blotting , Linfócitos T CD8-Positivos/metabolismo , Carcinoma de Células Renais/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Proliferação de Células/fisiologia , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Imuno-Histoquímica , Inflamação/genética , Inflamação/metabolismo , Neoplasias Renais/genética , Espectrometria de Massas , Camundongos , Proteômica/métodos , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de RNA , Microambiente Tumoral/genética , Microambiente Tumoral/fisiologia
2.
Mutat Res ; 785: 108319, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32800270

RESUMO

Cleft lip and palate (CL/P) is among the most common congenital malformations and affects 1 in 700 newborns. CL/P is caused by genetic and environmental factors (maternal smoking, alcohol or drug use and others). Many genes and loci were associated with cleft lip/palate but the amount of heterogeneity justifies identifying new causal genes and variants. AHRR (Aryl-Hydrocarbon Receptor Repressor) gene has recently been related to CL/P however, few functional studies analyze the genotypephenotype interaction of AHRR with CL/P. Several studies associate the molecular pathway of AHRR to CL/P which indicates this gene as a functional candidate in CL/P etiology. METHODS: Systematic Literature Review was performed using PUBMED database with the keywords cleft lip, cleft palate, orofacial cleft, AHRR and synonyms. SLR resulted in 37 included articles. RESULTS: AHRR is a positional and functional candidate gene for CL/P. In silico analysis detected interactions with other genes previously associated to CL/P like ARNT and CYP1A1. AHRR protein regulates cellular toxicity through TCDD mediated AHR pathway. Exposure to TCDD in animal embryos is AHR mediated and lead to cleft palate due to palate fusion failure and post fusion rupture. AHRR regulates cellular growth and differentiation, fundamental to lip and palatogenesis. AHRR decreases carcinogenesis and recently a higher tumor risk has been described in CL/P patients and families. AHRR is also a smoking biomarker due to changed methylation sites found in smokers DNA although folate intake may partially revert these methylation alterations. This corroborates the role of maternal smoking and lack of folate supplementation as risk factors for CL/P. CONCLUSION: This research identified the importance of AHRR in dioxin response and demonstrated an example of genetic and environmental interaction, indispensable in the development of many complex diseases.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fenda Labial/genética , Fissura Palatina/genética , Proteínas Repressoras/genética , Fumar/efeitos adversos , Motivos de Aminoácidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/química , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Biomarcadores/metabolismo , Metilação de DNA , Suplementos Nutricionais , Feminino , Ácido Fólico/metabolismo , Estudos de Associação Genética , Humanos , Recém-Nascido , Masculino , Modelos Moleculares , Domínios Proteicos , Isoformas de RNA/genética , Proteínas Repressoras/química , Proteínas Repressoras/metabolismo , Fatores de Risco
3.
Nat Commun ; 11(1): 4214, 2020 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-32843632

RESUMO

Stomata are epidermal structures that modulate gas exchanges between plants and the atmosphere. The formation of stomata is regulated by multiple developmental and environmental signals, but how these signals are coordinated to control this process remains unclear. Here, we showed that the conserved energy sensor kinase SnRK1 promotes stomatal development under short-day photoperiod or in liquid culture conditions. Mutation of KIN10, the catalytic α-subunit of SnRK1, results in the decreased stomatal index; while overexpression of KIN10 significantly induces stomatal development. KIN10 displays the cell-type-specific subcellular location pattern. The nuclear-localized KIN10 proteins are highly enriched in the stomatal lineage cells to phosphorylate and stabilize SPEECHLESS, a master regulator of stomatal formation, thereby promoting stomatal development. Our work identifies a module links connecting the energy signaling and stomatal development and reveals that multiple regulatory mechanisms are in place for SnRK1 to modulate stomatal development in response to changing environments.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Estômatos de Plantas/genética , Fatores de Transcrição/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Microscopia Confocal , Mutação , Fosforilação , Fotoperíodo , Estômatos de Plantas/citologia , Estômatos de Plantas/metabolismo , Plantas Geneticamente Modificadas , Estabilidade Proteica , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/genética , Fatores de Transcrição/metabolismo
4.
Proc Natl Acad Sci U S A ; 117(32): 19376-19387, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32719140

RESUMO

Inflammatory bowel diseases (IBDs), including Crohn's disease and ulcerative colitis, are associated with dysbiosis of the gut microbiome. Emerging evidence suggests that small-molecule metabolites derived from bacterial breakdown of a variety of dietary nutrients confer a wide array of host benefits, including amelioration of inflammation in IBDs. Yet, in many cases, the molecular pathways targeted by these molecules remain unknown. Here, we describe roles for three metabolites-indole-3-ethanol, indole-3-pyruvate, and indole-3-aldehyde-which are derived from gut bacterial metabolism of the essential amino acid tryptophan, in regulating intestinal barrier function. We determined that these metabolites protect against increased gut permeability associated with a mouse model of colitis by maintaining the integrity of the apical junctional complex and its associated actin regulatory proteins, including myosin IIA and ezrin, and that these effects are dependent on the aryl hydrocarbon receptor. Our studies provide a deeper understanding of how gut microbial metabolites affect host defense mechanisms and identify candidate pathways for prophylactic and therapeutic treatments for IBDs.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Microbioma Gastrointestinal/fisiologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Receptores de Hidrocarboneto Arílico/metabolismo , Triptofano/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Células CACO-2 , Colite Ulcerativa/dietoterapia , Colite Ulcerativa/microbiologia , Colite Ulcerativa/patologia , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Modelos Animais de Doenças , Humanos , Inflamação , Mucosa Intestinal/patologia , Camundongos , Camundongos Knockout , Miosina não Muscular Tipo IIA/metabolismo , Permeabilidade , Receptores de Hidrocarboneto Arílico/genética , Triptofano/administração & dosagem
5.
PLoS Genet ; 16(7): e1008883, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32609718

RESUMO

Plant steroid hormones brassinosteroids (BRs) regulate plant growth and development at many levels. While negative regulatory factors that inhibit development and are counteracted by BRs exist in the root meristem, these factors have not been characterized. The functions of UPB1 transcription factor in BR-regulated root growth have not been established, although its role in regulating root are well documented. Here, we found that BIN2 interacts with and phosphorylates the UPB1 transcription factor consequently promoting UPB1 stability and transcriptional activity. Genetic analysis revealed that UPB1 deficiency could partially recover the short-root phenotype of BR-deficient mutants. Expression of a mutated UPB1S37AS41A protein lacking a conserved BIN2 phosphorylation sites can rescue shorter root phenotype of bin2-1 mutant. In addition, UPB1 was repressed by BES1 at the transcriptional level. The paclobutrazol-resistant protein family (PRE2/3) interacts with UPB1 and inhibits its transcriptional activity to promote root meristem development, and BIN2-mediated phosphorylation of UPB1 suppresses its interaction with PRE2/3, and subsequently impairing root meristem development. Taken together, our data elucidate a molecular mechanism by which BR promotes root growth via inhibiting BIN2-UPB1 module.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Brassinosteroides/metabolismo , Proteínas Quinases/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica de Plantas/genética , Meristema/genética , Meristema/crescimento & desenvolvimento , Fosforilação , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Transdução de Sinais/genética
6.
Proc Natl Acad Sci U S A ; 117(31): 18840-18848, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32690706

RESUMO

Light and gravity are two key environmental factors that control plant growth and architecture. However, the molecular basis of the coordination of light and gravity signaling in plants remains obscure. Here, we report that two classes of transcription factors, PHYTOCHROME INTERACTING FACTORS (PIFs) and ELONGATED HYPOCOTYL5 (HY5), can directly bind and activate the expression of LAZY4, a positive regulator of gravitropism in both shoots and roots in Arabidopsis In hypocotyls, light promotes degradation of PIFs to reduce LAZY4 expression, which inhibits the negative gravitropism of hypocotyls. LAZY4 overexpression can partially rescue the negative gravitropic phenotype of pifq in the dark without affecting amyloplast development. Our identification of the PIFs-LAZY4 regulatory module suggests the presence of another role for PIF proteins in gravitropism, in addition to a previous report demonstrating that PIFs positively regulate amyloplast development to promote negative gravitropism in hypocotyls. In roots, light promotes accumulation of HY5 proteins to activate expression of LAZY4, which promotes positive gravitropism in roots. Together, our data indicate that light exerts opposite regulation of LAZY4 expression in shoots and roots by mediating the protein levels of PIFs and HY5, respectively, to inhibit the negative gravitropism of shoots and promote positive gravitropism of roots in Arabidopsis.


Assuntos
Proteínas de Arabidopsis , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Fatores de Transcrição de Zíper de Leucina Básica , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Gravitropismo/efeitos da radiação , Proteínas Nucleares , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/efeitos da radiação , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Luz , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo
7.
Rev Cardiovasc Med ; 21(2): 253-261, 2020 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-32706213

RESUMO

It is known that functional defects of GATA binding protein 5 (GATA5), an important member of GATA transcription factor family, could cause multiple congenital defects. However, the mechanisms of this transcription factor in cardiovascular diseases are still little known. Finding a genetic approach should help with understanding the possible roles of GATA5 in different cardiovascular diseases and purpose it as a possible therapeutic agent. Hence, this review is divided into three chapters to summarize the roles and main regulatory mechanisms of GATA5 in hypertension, arrhythmia and congenital heart disease, respectively. In each chapter, this review firstly introduces the roles of GATA5 mutations, and then discusses the main regulatory mechanisms of GATA5 in the corresponding diseases (Such as the endothelial dysfunction signaling pathway in the chapter of hypertension, GATA5-NaV1.5 signaling pathway in the chapter of arrhythmia, GATA5-HEY2 and GATA5-Nodal signaling pathway in the chapter of congenital heart disease). Additionally, based on these regulatory networks, it is also speculated that abnormal methylation of the GATA5 gene promoter may lead to cardiovascular diseases such as congenital heart disease. This conjecture is proposed to enrich the regulatory networks of GATA5 and provide a theoretical basis for diagnosis and treatment of cardiovascular diseases.


Assuntos
Doenças Cardiovasculares/metabolismo , Sistema Cardiovascular/metabolismo , Fator de Transcrição GATA5/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Doenças Cardiovasculares/genética , Doenças Cardiovasculares/fisiopatologia , Sistema Cardiovascular/fisiopatologia , Fator de Transcrição GATA5/genética , Redes Reguladoras de Genes , Predisposição Genética para Doença , Humanos , Mutação , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Canal de Sódio Disparado por Voltagem NAV1.5/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transdução de Sinais
8.
PLoS One ; 15(7): e0235922, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32673370

RESUMO

We have previously established that epigenetic regulator RING1 and YY1 binding protein (RYBP) is required for the contractility of embryonic stem (ES) cell derived cardiomyocytes (CMCs), suggesting its essential role in contractility. In order to investigate the underlying molecular events of this phenotype, we compared the transcriptomic profile of the wild type and Rybp null mutant ES cells and CMCs differentiated from these cell lines. We identified genes related to ion homeostasis, cell adhesion and sarcomeric organization affected in the Rybp null mutant CMCs, by using hierarchical gene clustering and Gene Ontology analysis. We have also demonstrated that the amount of RYBP is drastically reduced in the terminally differentiated wild type CMCs whilst it is broadly expressed in the early phase of differentiation when progenitors form. We also describe that RYBP is important for the proper expression of key cardiac transcription factors including Mesp1, Shh and Mef2c. These findings identify Rybp as a gene important for both early cardiac gene transcription and consequent sarcomere formation necessary for contractility. Since impairment of sarcomeric function and contractility plays a central role in reduced cardiac pump function leading to heart failures in human, current results might be relevant to the pathophysiology of cardiomyopathies.


Assuntos
Proteínas Repressoras/genética , Sarcômeros/fisiologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Diferenciação Celular , Linhagem Celular , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Canais Iônicos/genética , Canais Iônicos/metabolismo , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , RNA Mensageiro/metabolismo , Proteínas Repressoras/deficiência
9.
Biol Res ; 53(1): 25, 2020 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-32503642

RESUMO

BACKGROUND: Hypoxia inducible factor-1 (HIF-1) is considered as the most activated transcriptional factor in response to low oxygen level or hypoxia. HIF-1 binds the hypoxia response element (HRE) sequence in the promoter of different genes, mainly through the bHLH domain and activates the transcription of genes, especially those involved in angiogenesis and EMT. Considering the critical role of bHLH in binding HIF-1 to the HRE sequence, we hypothesized that bHLH could be a promising candidate to be targeted in hypoxia condition. METHODS: We inserted an inhibitory bHLH (ibHLH) domain in a pIRES2-EGFP vector and transfected HEK293T cells with either the control vector or the designed construct. The ibHLH domain consisted of bHLH domains of both HIF-1a and Arnt, capable of competing with HIF-1 in binding to HRE sequences. The transfected cells were then treated with 200 µM of cobalt chloride (CoCl2) for 48 h to induce hypoxia. Real-time PCR and western blot were performed to evaluate the effect of ibHLH on the genes and proteins involved in angiogenesis and EMT. RESULTS: Hypoxia was successfully induced in the HEK293T cell line as the gene expression of VEGF, vimentin, and ß-catenin were significantly increased after treatment of untransfected HEK293T cells with 200 µM CoCl2. The gene expression of VEGF, vimentin, and ß-catenin and protein level of ß-catenin were significantly decreased in the cells transfected with either control or ibHLH vectors in hypoxia. However, ibHLH failed to be effective on these genes and the protein level of ß-catenin, when compared to the control vector. We also observed that overexpression of ibHLH had more inhibitory effect on gene and protein expression of N-cadherin compared to the control vector. However, it was not statistically significant. CONCLUSION: bHLH has been reported to be an important domain involved in the DNA binding activity of HIF. However, we found that targeting this domain is not sufficient to inhibit the endogenous HIF-1 transcriptional activity. Further studies about the function of critical domains of HIF-1 are necessary for developing a specific HIF-1 inhibitor.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fator 1 Induzível por Hipóxia/metabolismo , Hipóxia/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Western Blotting , Expressão Gênica , Células HEK293 , Humanos , Hipóxia/genética , Fator 1 Induzível por Hipóxia/genética , Reação em Cadeia da Polimerase em Tempo Real , Ativação Transcricional/genética
10.
Proc Natl Acad Sci U S A ; 117(25): 14532-14542, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32513733

RESUMO

Implantation is initiated when an embryo attaches to the uterine luminal epithelium and subsequently penetrates into the underlying stroma to firmly embed in the endometrium. These events are followed by the formation of an extensive vascular network in the stroma that supports embryonic growth and ensures successful implantation. Interestingly, in many mammalian species, these processes of early pregnancy occur in a hypoxic environment. However, the mechanisms underlying maternal adaptation to hypoxia during early pregnancy remain unclear. In this study, using a knockout mouse model, we show that the transcription factor hypoxia-inducible factor 2 alpha (Hif2α), which is induced in subluminal stromal cells at the time of implantation, plays a crucial role during early pregnancy. Indeed, when preimplantation endometrial stromal cells are exposed to hypoxic conditions in vitro, we observed a striking enhancement in HIF2α expression. Further studies revealed that HIF2α regulates the expression of several metabolic and protein trafficking factors, including RAB27B, at the onset of implantation. RAB27B is a member of the Rab family of GTPases that allows controlled release of secretory granules. These granules are involved in trafficking MMP-9 from the stroma to the epithelium to promote luminal epithelial remodeling during embryo invasion. As pregnancy progresses, the HIF2α-RAB27B pathway additionally mediates crosstalk between stromal and endothelial cells via VEGF granules, developing the vascular network critical for establishing pregnancy. Collectively, our study provides insights into the intercellular communication mechanisms that operate during adaptation to hypoxia, which is essential for embryo implantation and establishment of pregnancy.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Hipóxia Celular/fisiologia , Implantação do Embrião/fisiologia , Vesículas Secretórias/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Comunicação Celular/fisiologia , Linhagem Celular , Embrião de Mamíferos , Endométrio/citologia , Endométrio/metabolismo , Feminino , Técnicas de Introdução de Genes , Humanos , Masculino , Camundongos , Camundongos Knockout , Gravidez , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transdução de Sinais/fisiologia , Células Estromais , Proteínas rab de Ligação ao GTP/genética
11.
Cell Prolif ; 53(7): e12833, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32525231

RESUMO

OBJECTIVES: The current study aimed to investigate the mechanism by which exosomes secreted by CHB patients with PNALT and liver inflammation grade (≥A2) affected the development of liver cancer. MATERIALS AND METHODS: Gene expression was assessed by RT-PCR, Western blotting and immunohistochemistry. CCK-8, colony formation, transwell, scratch-wound and flow cytometry assays were used to detect cell viability, proliferation, apoptosis and metastasis. The interaction of TCF21 and HHIP was assessed by co-immunoprecipitation assay. Luciferase reporter was used to detect the combination of TCF21/HHIP and miR-25-3p. Xenograft studies in nude mice manifested tumour growth ability of miR-25-3p. Bioinformatics analyses were conducted using TargetScan, EVmiRNA, TCGA, GEO, DAVID, COEXPEDIA, UALCAN, UCSC and the Human Protein Atlas databases. RESULTS: CHB-PNALT-Exo (≥A2) promoted the proliferation and metastasis of HepG2.2.15 cells. miR-25-3p was upregulated in CHB-PNALT-Exo (≥A2). miR-25-3p overexpression promoted cell proliferation and metastasis and was related to poor survival in patients with CHB-PNALT (≥A2). The cell proliferation- and metastasis-promoting functions of CHB-PNALT-Exo (≥A2) were abolished by miR-25-3p inhibitors. TCF21 directly interacted with HHIP. Inhibition of TCF21 or HHIP promoted cell proliferation and metastasis. Knockdown of TCF21 or HHIP counteracted the effects of CHB-PNALT-Exo (≥A2) containing miR-25-3p inhibitor on cell proliferation, metastasis and the expression of Ki67, E-cadherin and caspase-3/-9. CONCLUSIONS: Transfer of miR-25-3p by CHB-PNALT-Exo promoted the development of liver cancer by inhibiting the co-expression of TCF21 and HHIP.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Transporte/genética , Exossomos/genética , Hepatite B Crônica/genética , Neoplasias Hepáticas/genética , Fígado/patologia , Glicoproteínas de Membrana/genética , MicroRNAs/genética , Adulto , Animais , Apoptose/genética , Proliferação de Células/genética , Sobrevivência Celular , Progressão da Doença , Regulação para Baixo/genética , Exossomos/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Hepatite B Crônica/patologia , Humanos , Inflamação/genética , Inflamação/patologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Regulação para Cima/genética , Adulto Jovem
12.
Proc Natl Acad Sci U S A ; 117(26): 15075-15084, 2020 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-32532919

RESUMO

Some lineage-determining transcription factors are overwhelmingly important in directing embryonic cells to a particular differentiation pathway, such as Ascl1 for nerve. They also have an exceptionally strong ability to force cells to change from an unrelated pathway to one preferred by their action. Transcription factors are believed to have a very short residence time of only a few seconds on their specific DNA or chromatin-binding sites. We have developed a procedure in which DNA containing one copy of the binding site for the neural-inducing factor Ascl1 is injected directly into a Xenopus oocyte nucleus which has been preloaded with a limiting amount of the Ascl1 transcription factor protein. This is followed by a further injection of DNA as a competitor, either in a plasmid or in chromosomal DNA, containing the same binding site but with a different reporter. Importantly, expression of the reporter provides a measure of the function of the transcription factor in addition to its residence time. The same long residence time and resistance to competition are seen with the estrogen receptor and its DNA response elements. We find that in this nondividing oocyte, the nerve-inducing factor Ascl1 can remain bound to a specific chromatin site for hours or days and thereby help to stabilize gene expression. This stability of transcription factor binding to chromatin is a necessary part of its action because removal of this factor causes discontinuation of its effect on gene expression. Stable transcription factor binding may be a characteristic of nondividing cells.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Cromatina/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Sítios de Ligação , Cromatina/genética , DNA/genética , DNA/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Oócitos/crescimento & desenvolvimento , Oócitos/metabolismo , Ligação Proteica , Xenopus laevis/embriologia , Xenopus laevis/genética , Xenopus laevis/metabolismo
13.
PLoS Genet ; 16(5): e1008782, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32421721

RESUMO

The planar cell polarity pathway is required for heart development and whilst the functions of most pathway members are known, the roles of the jnk genes in cardiac morphogenesis remain unknown as mouse mutants exhibit functional redundancy, with early embryonic lethality of compound mutants. In this study zebrafish were used to overcome early embryonic lethality in mouse models and establish the requirement for Jnk in heart development. Whole mount in-situ hybridisation and RT-PCR demonstrated that evolutionarily conserved alternative spliced jnk1a and jnk1b transcripts were expressed in the early developing heart. Maternal zygotic null mutant zebrafish lines for jnk1a and jnk1b, generated using CRISPR-Cas9, revealed a requirement for jnk1a in formation of the proximal, first heart field (FHF)-derived portion of the cardiac ventricular chamber. Rescue of the jnk1a mutant cardiac phenotype was only possible by injection of the jnk1a EX7 Lg alternatively spliced transcript. Analysis of mutants indicated that there was a reduction in the size of the hand2 expression field in jnk1a mutants which led to a specific reduction in FHF ventricular cardiomyocytes within the anterior lateral plate mesoderm. Moreover, the jnk1a mutant ventricular defect could be rescued by injection of hand2 mRNA. This study reveals a novel and critical requirement for Jnk1 in heart development and highlights the importance of alternative splicing in vertebrate cardiac morphogenesis. Genetic pathways functioning through jnk1 may be important in human heart malformations with left ventricular hypoplasia.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Ventrículos do Coração/citologia , Proteína Quinase 8 Ativada por Mitógeno/genética , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Processamento Alternativo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Contagem de Células , Células Cultivadas , Éxons , Regulação da Expressão Gênica no Desenvolvimento , Ventrículos do Coração/embriologia , Ventrículos do Coração/metabolismo , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética
14.
PLoS Genet ; 16(5): e1008797, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32392219

RESUMO

Sun-loving plants perceive the proximity of potential light-competing neighboring plants as a reduction in the red:far-red ratio (R:FR), which elicits a suite of responses called the "shade avoidance syndrome" (SAS). Changes in R:FR are primarily perceived by phytochrome B (phyB), whereas UV-B perceived by UV RESISTANCE LOCUS 8 (UVR8) elicits opposing responses to provide a counterbalance to SAS, including reduced shade-induced hypocotyl and petiole elongation. Here we show at the genome-wide level that UVR8 broadly suppresses shade-induced gene expression. A subset of this gene regulation is dependent on the UVR8-stabilized atypical bHLH transcription regulator LONG HYPOCOTYL IN FAR-RED 1 (HFR1), which functions in part redundantly with PHYTOCHROME INTERACTING FACTOR 3-LIKE 1 (PIL1). In parallel, UVR8 signaling decreases protein levels of the key positive regulators of SAS, namely the bHLH transcription factors PHYTOCHROME INTERACTING FACTOR 4 (PIF4) and PIF5, in a COP1-dependent but HFR1-independent manner. We propose that UV-B antagonizes SAS via two mechanisms: degradation of PIF4 and PIF5, and HFR1- and PIL1-mediated inhibition of PIF4 and PIF5 function. This work highlights the importance of typical and atypical bHLH transcription regulators for the integration of light signals from different photoreceptors and provides further mechanistic insight into the crosstalk of UVR8 signaling and SAS.


Assuntos
Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/química , Proteínas Cromossômicas não Histona/genética , Proteínas de Ligação a DNA/química , Raios Ultravioleta/efeitos adversos , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Regulação para Baixo , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Estabilidade Proteica , Proteólise , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
15.
PLoS Genet ; 16(5): e1008807, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32407354

RESUMO

Pollen wall consists of several complex layers which form elaborate species-specific patterns. In Arabidopsis, the transcription factor ABORTED MICROSPORE (AMS) is a master regulator of exine formation, and another transcription factor, TRANSPOSABLE ELEMENT SILENCING VIA AT-HOOK (TEK), specifies formation of the nexine layer. However, knowledge regarding the temporal regulatory roles of TEK in pollen wall development is limited. Here, TEK-GFP driven by the AMS promoter was prematurely expressed in the tapetal nuclei, leading to complete male sterility in the pAMS:TEK-GFP (pat) transgenic lines with the wild-type background. Cytological observations in the pat anthers showed impaired callose synthesis and aberrant exine patterning. CALLOSE SYNTHASE5 (CalS5) is required for callose synthesis, and expression of CalS5 in pat plants was significantly reduced. We demonstrated that TEK negatively regulates CalS5 expression after the tetrad stage in wild-type anthers and further discovered that premature TEK-GFP in pat directly represses CalS5 expression through histone modification. Our findings show that TEK flexibly mediates its different functions via different temporal regulation, revealing that the temporal regulation of TEK is essential for exine patterning. Moreover, the result that the repression of CalS5 by TEK after the tetrad stage coincides with the timing of callose wall dissolution suggests that tapetum utilizes temporal regulation of genes to stop callose wall synthesis, which, together with the activation of callase activity, achieves microspore release and pollen wall patterning.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Pólen/fisiologia , Fatores de Transcrição/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Epigênese Genética , Regulação da Expressão Gênica de Plantas , Glucosiltransferases/metabolismo , Histonas/metabolismo , Metilação , Plantas Geneticamente Modificadas/fisiologia , Pólen/genética , Regiões Promotoras Genéticas
16.
Proc Natl Acad Sci U S A ; 117(22): 12230-12238, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32414920

RESUMO

Tibetans have adapted to the chronic hypoxia of high altitude and display a distinctive suite of physiologic adaptations, including augmented hypoxic ventilatory response and resistance to pulmonary hypertension. Genome-wide studies have consistently identified compelling genetic signatures of natural selection in two genes of the Hypoxia Inducible Factor pathway, PHD2 and HIF2A The product of the former induces the degradation of the product of the latter. Key issues regarding Tibetan PHD2 are whether it is a gain-of-function or loss-of-function allele, and how it might contribute to high-altitude adaptation. Tibetan PHD2 possesses two amino acid changes, D4E and C127S. We previously showed that in vitro, Tibetan PHD2 is defective in its interaction with p23, a cochaperone of the HSP90 pathway, and we proposed that Tibetan PHD2 is a loss-of-function allele. Here, we report that additional PHD2 mutations at or near Asp-4 or Cys-127 impair interaction with p23 in vitro. We find that mice with the Tibetan Phd2 allele display augmented hypoxic ventilatory response, supporting this loss-of-function proposal. This is phenocopied by mice with a mutation in p23 that abrogates the PHD2:p23 interaction. Hif2a haploinsufficiency, but not the Tibetan Phd2 allele, ameliorates hypoxia-induced increases in right ventricular systolic pressure. The Tibetan Phd2 allele is not associated with hemoglobin levels in mice. We propose that Tibetans possess genetic alterations that both activate and inhibit selective outputs of the HIF pathway to facilitate successful adaptation to the chronic hypoxia of high altitude.


Assuntos
Adaptação Fisiológica , Proteínas de Ligação a DNA/metabolismo , Prolina Dioxigenases do Fator Induzível por Hipóxia/genética , Prolina Dioxigenases do Fator Induzível por Hipóxia/metabolismo , Prolina Dioxigenases do Fator Induzível por Hipóxia/fisiologia , Hipóxia/fisiopatologia , Mutação com Perda de Função , Alelos , Altitude , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas de Ligação a DNA/genética , Humanos , Camundongos , Camundongos Knockout , Fenótipo , Seleção Genética , Tibet
17.
Nat Commun ; 11(1): 2588, 2020 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-32444594

RESUMO

The lysine acetyltransferases type 3 (KAT3) family members CBP and p300 are important transcriptional co-activators, but their specific functions in adult post-mitotic neurons remain unclear. Here, we show that the combined elimination of both proteins in forebrain excitatory neurons of adult mice resulted in a rapidly progressing neurological phenotype associated with severe ataxia, dendritic retraction and reduced electrical activity. At the molecular level, we observed the downregulation of neuronal genes, as well as decreased H3K27 acetylation and pro-neural transcription factor binding at the promoters and enhancers of canonical neuronal genes. The combined deletion of CBP and p300 in hippocampal neurons resulted in the rapid loss of neuronal molecular identity without de- or transdifferentiation. Restoring CBP expression or lysine acetylation rescued neuronal-specific transcription in cultured neurons. Together, these experiments show that KAT3 proteins maintain the excitatory neuron identity through the regulation of histone acetylation at cell type-specific promoter and enhancer regions.


Assuntos
Encéfalo/citologia , Lisina Acetiltransferases/metabolismo , Neurônios/fisiologia , Acetilação , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Encéfalo/fisiologia , Elementos Facilitadores Genéticos , Epigenoma , Feminino , Regulação da Expressão Gênica , Lisina Acetiltransferases/genética , Masculino , Proteínas de Membrana/metabolismo , Camundongos Knockout , Neurônios/citologia , Fosfoproteínas/metabolismo , Fatores de Transcrição de p300-CBP/metabolismo
18.
Nat Commun ; 11(1): 2608, 2020 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-32451418

RESUMO

IL-22 has dual functions during tumorigenesis. Short term IL-22 production protects against genotoxic stress, whereas uncontrolled IL-22 activity promotes tumor growth; therefore, tight regulation of IL-22 is essential. TGF-ß1 promotes the differentiation of Th17 cells, which are known to be a major source of IL-22, but the effect of TGF-ß signaling on the production of IL-22 in CD4+ T cells is controversial. Here we show an increased presence of IL-17+IL-22+ cells and TGF-ß1 in colorectal cancer compared to normal adjacent tissue, whereas the frequency of IL-22 single producing cells is not changed. Accordingly, TGF-ß signaling in CD4+ T cells (specifically Th17 cells) promotes the emergence of IL-22-producing Th17 cells and thereby tumorigenesis in mice. IL-22 single producing T cells, however, are not dependent on TGF-ß signaling. We show that TGF-ß, via AhR induction, and PI3K signaling promotes IL-22 production in Th17 cells.


Assuntos
Colite/complicações , Neoplasias do Colo/etiologia , Interleucinas/biossíntese , Células Th17/imunologia , Fator de Crescimento Transformador beta/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Carcinogênese/imunologia , Diferenciação Celular , Colite/imunologia , Neoplasias do Colo/imunologia , Neoplasias do Colo/patologia , Neoplasias Colorretais/etiologia , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Feminino , Humanos , Interleucina-17/genética , Interleucina-17/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fosfatidilinositol 3-Quinases/metabolismo , Receptores de Hidrocarboneto Arílico/genética , Receptores de Hidrocarboneto Arílico/metabolismo , Transdução de Sinais/imunologia , Células Th17/patologia , Fator de Crescimento Transformador beta1/metabolismo
19.
Physiol Plant ; 169(3): 452-466, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32412656

RESUMO

Under photoperiodic conditions, Arabidopsis thaliana seedling growth is inhibited in long days (LDs), but promoted under the extended nights of short days (SDs). This behavior is partly implemented by phytochrome (phy)-imposed oscillations in the abundance of the growth-promoting, phy-interacting bHLH transcription factors PHY-INTERACTING FACTOR 1 (PIF1), PIF3, PIF4 and PIF5 (PIF quartet or PIFq). However, the observation that a pifq mutant is still stimulated to elongate when given a phy-inactivating end-of-day far-red pulse (EODFR), suggests that additional factors are involved in the phy-mediated suppression of growth during the subsequent dark period. Here, by combining growth-analysis of pif7 single- and higher-order mutants with gene expression analysis under SD, LD, SD-EODFR, and LD-EODFR, we show that PIF7 promotes growth during the dark hours of SD, by regulating growth-related gene expression. Interestingly, the relative contribution of PIF7 in promoting growth is stronger under EODFR, whereas PIF3 role is more important under SD, suggesting that PIF7 is a prominent target of phy-suppression. Indeed, we show that phy imposes phosphorylation and inactivation of PIF7 during the light hours in SD, and prevents full dephosphorylation during the night. This repression can be lifted with an EODFR, which correlates with increased PIF7-mediated gene expression and elongation. In addition, our results suggest that PIF7 function might involve heterodimerization with PIF3. Furthermore, our data indicate that a pifqpif7 quintuple mutant is largely insensitive to photoperiod for hypocotyl elongation. Collectively, the data suggest that PIF7, together with the PIFq, is required for the photoperiodic regulation of seasonal growth.


Assuntos
Proteínas de Arabidopsis , Arabidopsis/genética , Proteínas de Ligação a DNA , Fitocromo/genética , Proteínas de Arabidopsis/fisiologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Ligação a DNA/fisiologia , Regulação da Expressão Gênica de Plantas , Hipocótilo/genética , Luz , Fotoperíodo
20.
PLoS One ; 15(5): e0232986, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32407419

RESUMO

Here we focus on the highly conserved MYB-bHLH-WD repeat (MBW) transcriptional complex model in eggplant, which is pivotal in the transcriptional regulation of the anthocyanin biosynthetic pathway. Through a genome-wide approach performed on the recently released Eggplant Genome (cv. 67/3) previously identified, and reconfirmed by us, members belonging to the MBW complex (SmelANT1, SmelAN2, SmelJAF13, SmelAN1) were functionally characterized. Furthermore, a regulatory R3 MYB type repressor (SmelMYBL1), never reported before, was identified and characterized as well. Through a qPCR approach, we revealed specific transcriptional patterns of candidate genes in different plant tissue/organs at two stages of fruit development. Two strategies were adopted for investigating the interactions of bHLH partners (SmelAN1, SmelJAF13) with MYB counterparts (SmelANT1, SmelAN2 and SmelMYBL1): Yeast Two Hybrid (Y2H) and Bimolecular Fluorescent Complementation (BiFC) in A. thaliana mesophylls protoplast. Agro-infiltration experiments highlighted that N. benthamiana leaves transiently expressing SmelANT1 and SmelAN2 showed an anthocyanin-pigmented phenotype, while their co-expression with SmelMYBL1 prevented anthocyanin accumulation. Our results suggest that SmelMYBL1 may inhibits the MBW complex via the competition with MYB activators for bHLH binding site, although this hypothesis requires further elucidation.


Assuntos
Antocianinas/biossíntese , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Solanum melongena/genética , Solanum melongena/metabolismo , Sequência de Aminoácidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Genes Reguladores , Família Multigênica , Filogenia , Plantas Geneticamente Modificadas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Tabaco/genética , Tabaco/metabolismo
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