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1.
Nat Struct Mol Biol ; 31(2): 351-363, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38182926

RESUMO

UBR4 is a 574 kDa E3 ligase (E3) of the N-degron pathway with roles in neurodevelopment, age-associated muscular atrophy and cancer. The catalytic module that carries out ubiquitin (Ub) transfer remains unknown. Here we identify and characterize a distinct E3 module within human UBR4 consisting of a 'hemiRING' zinc finger, a helical-rich UBR zinc-finger interacting (UZI) subdomain, and an N-terminal region that can serve as an affinity factor for the E2 conjugating enzyme (E2). The structure of an E2-E3 complex provides atomic-level insight into the specificity determinants of the hemiRING toward the cognate E2s UBE2A/UBE2B. Via an allosteric mechanism, the UZI subdomain modestly activates the Ub-loaded E2 (E2∼Ub). We propose attenuated activation is complemented by the intrinsically high lysine reactivity of UBE2A, and their cooperation imparts a reactivity profile important for substrate specificity and optimal degradation kinetics. These findings reveal the mechanistic underpinnings of a neuronal N-degron E3, its specific recruitment of UBE2A, and highlight the underappreciated architectural diversity of cross-brace domains with Ub E3 activity.


Assuntos
Enzimas de Conjugação de Ubiquitina , Ubiquitina-Proteína Ligases , Humanos , Ubiquitina-Proteína Ligases/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitina/metabolismo , Enzimas Ativadoras de Ubiquitina/metabolismo , Catálise , Ubiquitinação , Proteínas de Ligação a Calmodulina/metabolismo
2.
J Clin Immunol ; 43(8): 2011-2021, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37695435

RESUMO

Autosomal recessive tyrosine kinase 2 (TYK2) deficiency is characterized by susceptibility to mycobacterial and viral infections. Here, we report a 4-year-old female with severe respiratory viral infections, EBV-driven Burkitt-like lymphoma, and infection with the neurotropic Jamestown Canyon virus. A novel, homozygous c.745C > T (p.R249*) variant was found in TYK2. The deleterious effects of the TYK2 lesion were confirmed by immunoblotting; by evaluating functional responses to IFN-α/ß, IL-10, and IL-23; and by assessing its scaffolding effect on the cell surface expression of cytokine receptor subunits. The effects of the mutation could not be pharmacologically circumvented in vitro, suggesting that alternative modalities, such as hematopoietic stem cell transplantation or gene therapy, may be needed. We characterize the first patient from Canada with a novel homozygous mutation in TYK2.


Assuntos
Encefalite Viral , Linfoma , Viroses , Feminino , Humanos , Pré-Escolar , Herpesvirus Humano 4 , TYK2 Quinase/genética , Mutação/genética
3.
Nat Chem Biol ; 16(11): 1227-1236, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32747811

RESUMO

MYCBP2 is a ubiquitin (Ub) E3 ligase (E3) that is essential for neurodevelopment and regulates axon maintenance. MYCBP2 transfers Ub to nonlysine substrates via a newly discovered RING-Cys-Relay (RCR) mechanism, where Ub is relayed from an upstream cysteine to a downstream substrate esterification site. The molecular bases for E2-E3 Ub transfer and Ub relay are unknown. Whether these activities are linked to the neural phenotypes is also unclear. We describe the crystal structure of a covalently trapped E2~Ub:MYCBP2 transfer intermediate revealing key structural rearrangements upon E2-E3 Ub transfer and Ub relay. Our data suggest that transfer to the dynamic upstream cysteine, whilst mitigating lysine activity, requires a closed-like E2~Ub conjugate with tempered reactivity, and Ub relay is facilitated by a helix-coil transition. Furthermore, neurodevelopmental defects and delayed injury-induced degeneration in RCR-defective knock-in mice suggest its requirement, and that of substrate esterification activity, for normal neural development and programmed axon degeneration.


Assuntos
Axônios/metabolismo , Cisteína/metabolismo , Domínios RING Finger , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Sítios de Ligação , Feminino , Técnicas de Introdução de Genes , Humanos , Lisina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL/embriologia , Camundongos Transgênicos , Modelos Moleculares , Conformação Molecular , Ligação Proteica , Conformação Proteica , Transdução de Sinais , Relação Estrutura-Atividade , Ubiquitinação
4.
Sci Rep ; 7: 42285, 2017 02 13.
Artigo em Inglês | MEDLINE | ID: mdl-28205568

RESUMO

The cellular prion protein (PrP) is essential for transmissible prion diseases, but its exact physiological function remains unclear. Better understanding the regulation of the human prion protein gene (PRNP) expression can provide insight into this elusive function. Spliced XBP1 (sXBP1) was recently shown to mediate endoplasmic reticulum (ER) stress-induced PRNP expression. In this manuscript, we identify Luman, a ubiquitous, non-canonical unfolded protein response (UPR), as a novel regulator of ER stress-induced PRNP expression. Luman activity was transcriptionally and proteolytically activated by the ER stressing drug brefeldin A (BFA) in human neurons, astrocytes, and breast cancer MCF-7 cells. Over-expression of active cleaved Luman (ΔLuman) increased PrP levels, while siRNA-mediated Luman silencing decreased BFA-induced PRNP expression. Site-directed mutagenesis and chromatin immunoprecipitation demonstrated that ΔLuman regulates PRNP expression by interacting with the ER stress response element 26 (ERSE26). Co-over-expression and siRNA-mediated silencing experiments showed that sXBP1 and ΔLuman both up-regulate ER stress-induced PRNP expression. Attempts to understand the function of PRNP up-regulation by Luman excluded a role in atorvastatin-induced neuritogenesis, ER-associated degradation, or proteasomal inhibition-induced cell death. Overall, these results refine our understanding of ER stress-induced PRNP expression and function.


Assuntos
Brefeldina A/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Estresse do Retículo Endoplasmático/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas Priônicas/genética , Elementos de Resposta/genética , Animais , Atorvastatina/farmacologia , Sequência de Bases , Caspase 3/metabolismo , Caspase 7/metabolismo , Linhagem Celular , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Degradação Associada com o Retículo Endoplasmático/efeitos dos fármacos , Humanos , Camundongos , Neuritos/efeitos dos fármacos , Neuritos/metabolismo , Neurogênese/efeitos dos fármacos , Oligopeptídeos/farmacologia , Pré-Albumina/metabolismo , Proteínas Priônicas/metabolismo , Regiões Promotoras Genéticas/genética , Especificidade por Substrato/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Regulação para Cima/genética , Proteína 1 de Ligação a X-Box/metabolismo
5.
Hum Mol Genet ; 25(5): 976-88, 2016 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-26740554

RESUMO

Similar to many proteins trafficking through the secretory pathway, cellular prion protein (PrP) partly retrotranslocates from the endoplasmic reticulum to the cytosol through the endoplasmic reticulum-associated degradation (ERAD) pathway in an attempt to alleviate accumulation of cellular misfolded PrP. Surprisingly, familial PrP mutants fail to retrotranslocate and simultaneously block normal cellular PrP retrotranslocation. That impairments in retrotranslocation of misfolded proteins could lead to global disruptions in cellular homeostasis prompted further investigations into PrP mutant retrotranslocation defects. A gain- and loss-of-function approach identified human E3 ubiquitin ligase, Hrd1, as a critical regulator of PrP retrotranslocation in mammalian cells. Expression of familial human PrP mutants, V210I(129V) and M232R(129V), not only abolished PrP retrotranslocation, but also that of Hrd1-dependent ERAD substrates, transthyretin TTR(D18G) and α1-anti-trypsin A1AT(NHK). Mutant PrP expression decreased binding immunoglobulin protein (BiP) levels by 50% and attenuated ER stress-induced BiP by increasing BiP turnover 6-fold. Overexpression of BiP with PrP mutants rescued retrotranslocation of PrP, TTR(D18G) and A1AT(NHK). PrP mutants-induced cell death was also rescued by co-expression of BiP. These results show that PrP mutants highjack the Hrd1-dependent ERAD pathway, an action that would result in misfolded protein accumulation especially in terminally differentiated neurons. This could explain the age-dependent neuronal degeneration in familial prion diseases.


Assuntos
Degradação Associada com o Retículo Endoplasmático/genética , Proteínas de Choque Térmico/genética , Neurônios/metabolismo , Príons/metabolismo , Ubiquitina-Proteína Ligases/genética , Animais , Morte Celular , Linhagem Celular Tumoral , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Chaperona BiP do Retículo Endoplasmático , Regulação da Expressão Gênica , Proteínas de Choque Térmico/metabolismo , Humanos , Mutação , Neuroglia/metabolismo , Neuroglia/patologia , Neurônios/patologia , Príons/genética , Dobramento de Proteína , Transporte Proteico , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Receptores de Albumina/genética , Receptores de Albumina/metabolismo , Transdução de Sinais , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Ubiquitina-Proteína Ligases/metabolismo , alfa 1-Antitripsina/genética , alfa 1-Antitripsina/metabolismo
6.
Cell Signal ; 26(8): 1649-56, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24726894

RESUMO

Hypoxia-inducible transcription factor-1 (HIF-1) plays a decisive role in cell survival and adaptation to hypoxic stress by controlling the expression of genes involved in oxygen homeostasis. HIF-1 activity is fine-tuned through specific post-translational modifications of its essential HIF-1α subunit. Among these modifications, phosphorylation is important for HIF-1 transcriptional activity. Studies have shown that the mitogen-activated protein kinases, p42/p44 MAPKs, directly phosphorylate HIF-1α and increase HIF-1-mediated transcription. Pin1, a peptidyl-prolyl cis/trans isomerase, targets a number of proteins containing a phosphorylated Ser/Thr-Pro motif. Pin1 isomerization causes a change in target protein conformation which can modify their activity. Here, we identify Pin1 as an important HIF-1α partner. Immunoprecipitation and pull-down studies show that Pin1 interacts with HIF-1α. We demonstrate that the interaction between Pin1 and HIF-1α is regulated through p42/p44 MAPK pathway activation. By performing proteolysis studies, our results indicate that Pin1 catalytic activity generates a conformational change in HIF-1α. Finally, our work shows that Pin1 is required for gene-specific HIF-1 transcriptional activity. Our results indicate that the prolyl isomerase Pin1 regulates HIF-1 transcriptional activity by interacting with HIF-1α and promoting conformational changes in a p42/p44 MAPK phosphorylation-dependent manner.


Assuntos
Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Peptidilprolil Isomerase/metabolismo , Animais , Linhagem Celular , Células HEK293 , Células HeLa , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Peptidilprolil Isomerase de Interação com NIMA , Peptidilprolil Isomerase/antagonistas & inibidores , Peptidilprolil Isomerase/genética , Fosforilação , Domínios e Motivos de Interação entre Proteínas , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transcrição Gênica
7.
J Biol Chem ; 288(28): 20378-91, 2013 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-23737521

RESUMO

Understanding the regulatory mechanisms mediating PRNP gene expression is highly relevant to elucidating normal cellular prion protein (PrP) function(s) and the transmissibility of prion protein neurodegenerative diseases. Here, luciferase reporter assays showed that an endoplasmic reticulum stress element (ERSE)-like element, CCAAT-N26-CCACG in the human PRNP promoter, is regulated by ER stress and X-box-binding protein 1 (XBP1) but not by activating transcription factor 6 α (ATF6α). Bioinformatics identified the ERSE-26 motif in 37 other human genes in the absence of canonical ERSE sites except for three genes. Several of these genes are associated with a synaptic function or are involved in oxidative stress. Brefeldin A, tunicamycin, and thapsigargin ER stressors induced gene expression of PRNP and four randomly chosen ERSE-26-containing genes, ERLEC1, GADD45B, SESN2, and SLC38A5, in primary human neuron cultures or in the breast carcinoma MCF-7 cell line, although the level of the response depends on the gene analyzed, the genetic background of the cells, the cell type, and the ER stressor. Overexpression of XBP1 increased, whereas siRNA knockdown of XBP1 considerably reduced, PRNP and ERLEC1 mRNA levels in MCF-7 cells. Taken together, these results identify a novel ER stress regulator, which implicates the ER stress response in previously unrecognized cellular functions.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Estresse do Retículo Endoplasmático , Regulação da Expressão Gênica , Elementos de Resposta , Fatores de Transcrição/metabolismo , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Sistemas de Transporte de Aminoácidos Neutros/genética , Antígenos de Diferenciação/genética , Sequência de Bases , Western Blotting , Brefeldina A/farmacologia , Células Cultivadas , Proteínas de Ligação a DNA/genética , Células HEK293 , Humanos , Lectinas/genética , Células MCF-7 , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Proteínas Nucleares/genética , Motivos de Nucleotídeos/genética , Proteínas Priônicas , Príons/genética , Regiões Promotoras Genéticas/genética , Interferência de RNA , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Transcrição de Fator Regulador X , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tapsigargina/farmacologia , Fatores de Transcrição/genética , Tunicamicina/farmacologia , Proteína 1 de Ligação a X-Box
8.
Breast Cancer Res ; 15(2): R22, 2013 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-23497519

RESUMO

INTRODUCTION: High prion protein (PrP) levels are associated with breast, colon and gastric cancer resistance to treatment and with a poor prognosis for the patients. However, little is known about the underlying molecular mechanism(s) regulating human PrP gene (PRNP) expression in cancers. Because endoplasmic reticulum (ER) stress is associated with solid tumors, we investigated a possible regulation of PRNP gene expression by ER stress. METHODS: Published microarray databases of breast cancer tissues and breast carcinoma cell lines were analyzed for PrP mRNA and ER stress marker immunoglobulin heavy chain binding protein (BiP) levels. Breast cancer tissue microarrays (TMA) were immunostained for BiP and PrP. Breast carcinoma MCF-7, MDA-MB-231, HS578T and HCC1500 cells were treated with three different ER stressors - Brefeldin A, Tunicamycin, Thapsigargin - and levels of PrP mRNA or protein assessed by RT-PCR and Western blot analyses. A human PRNP promoter-luciferase reporter was used to assess transcriptional activation by ER stressors. Site-directed mutagenesis identified the ER stress response elements (ERSE). Chromatin immunoprecipitation (ChIP) analyses were done to identify the ER stress-mediated transcriptional regulators. The role of cleaved activating transcription factor 6α (ΔATF6α) and spliced X-box protein-1 (sXBP1) in PRNP gene expression was assessed with over-expression or silencing techniques. The role of PrP protection against ER stress was assessed with PrP siRNA and by using Prnp null cell lines. RESULTS: We find that mRNA levels of BiP correlated with PrP transcript levels in breast cancer tissues and breast carcinoma cell lines. PrP mRNA levels were enriched in the basal subtype and were associated with poor prognosis in breast cancer patients. Higher PrP and BiP levels correlated with increasing tumor grade in TMA. ER stress was a positive regulator of PRNP gene transcription in MCF-7 cells and luciferase reporter assays identified one ER stress response element (ERSE) conserved among primates and rodents and three primate-specific ERSEs that regulated PRNP gene expression. Among the various transactivators of the ER stress-regulated unfolded protein response (UPR), ATF6α and XBP1 transactivated PRNP gene expression, but the ability of these varied in different cell types. Functionally, PrP delayed ER stress-induced cell death. CONCLUSIONS: These results establish PRNP as a novel ER stress-regulated gene that could increase survival in breast cancers.


Assuntos
Neoplasias da Mama/patologia , Estresse do Retículo Endoplasmático , Regulação Neoplásica da Expressão Gênica , Príons/metabolismo , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Apoptose , Western Blotting , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Citometria de Fluxo , Humanos , Técnicas Imunoenzimáticas , Luciferases/metabolismo , Proteínas Priônicas , Príons/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição de Fator Regulador X , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise Serial de Tecidos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Células Tumorais Cultivadas , Proteína 1 de Ligação a X-Box
9.
J Cardiovasc Pharmacol ; 54(1): 31-7, 2009 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19528817

RESUMO

This study was designed to evaluate the effect of long-term pretreatment with celecoxib, a cyclooxygenase-2 inhibitor, on myocardial infarct size. Celecoxib (3 mg/kg/day i.p; n = 16) or vehicle (DMSO 50%; EtOH 15%; distilled water, n = 16) was administered chronically to male Sprague-Dawley rats through ALZET osmotic pumps for 28 days. Under anaesthesia, the animals were then subjected to left anterior descending coronary artery occlusion for 40 minutes, followed by 24-hour reperfusion. The results show that myocardial infarct size in celecoxib-treated rats was significantly reduced compared to the control group (37.5 +/- 2.5% versus 48.0 +/- 2.6% of the area at risk, P < 0.05, n = 10 per group). Accumulation of neutrophils, estimated by myeloperoxidase levels, indicated an increase in the ischemic area without any significant difference between groups. No significant difference was observed between the treated and vehicle groups in terms of plasma prostaglandin E2 and tumour necrosis factor-alpha. Apoptosis, evaluated by Bax/Bcl-2 and terminal dUTP nick-end labelled-positive cells, was significantly decreased in the subendocardial layer of the ischemic area in celecoxib-treated rats. This study indicates that pretreatment with celecoxib can reduce infarct size by a mechanism, which may involve apoptosis inhibition.


Assuntos
Inibidores de Ciclo-Oxigenase 2/uso terapêutico , Isquemia/patologia , Infarto do Miocárdio/patologia , Traumatismo por Reperfusão Miocárdica/patologia , Pirazóis/uso terapêutico , Sulfonamidas/uso terapêutico , Animais , Celecoxib , Modelos Animais de Doenças , Infusões Parenterais , Masculino , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley
10.
Neurosci Lett ; 453(1): 73-6, 2009 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-19429019

RESUMO

The systemic administration of atorvastatin has been shown to be neuroprotective after spinal cord injury (SCI), by decreasing the inflammatory response at the lesion site and by reducing neuronal and oligodendrocyte apoptosis. The latter effect spares white matter at the injury site and improves locomotion. The aim of this study was to confirm the neuroprotective efficacy of atorvastatin as well as its early action in limiting apoptosis with its administration post-SCI. Female Sprague-Dawley rats received an intra peritoneal injection of: (1) statin/saline (5mg/kg) at 2h after the contusion injury; (2) physiological saline at 2h post-SCI; or (3) physiological saline without injury. Statin-treated rats showed significant (p<0.05) improvement in locomotion at week 4 post-SCI compared to vehicle-treated animals. Explaining this outcome, caspase-3 activity decreased by 50% (p<0.05), and the histological TUNEL method revealed a decrease of approximately 20% in apoptotic cells at the injury site (p<0.01) at 4h post-SCI in atorvastatin-treated rats in comparison to vehicle-treated controls. These data demonstrate that atorvastatin is effective after experimental spinal cord contusion injury in preventing early apoptosis at the injury site within 2h post-administration.


Assuntos
Apoptose/efeitos dos fármacos , Ácidos Heptanoicos/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Pirróis/uso terapêutico , Recuperação de Função Fisiológica/efeitos dos fármacos , Traumatismos da Medula Espinal/tratamento farmacológico , Análise de Variância , Animais , Atorvastatina , Caspase 3/metabolismo , Feminino , Marcação In Situ das Extremidades Cortadas , Atividade Motora/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Traumatismos da Medula Espinal/patologia , Vértebras Torácicas
11.
Bull Cancer ; 93(4): 349-56, 2006 Apr.
Artigo em Francês | MEDLINE | ID: mdl-16627239

RESUMO

The transcription factor hypoxia-inducible factor 1 (HIF-1) regulates the expression of more than 70 genes in response to hypoxic stress. Composed of two subunits, HIF-1 activity is modulated by the availability of the HIF-1alpha protein subunit. The stability and transcriptional activity of this extremely labile protein is affected by post-translational modifications. Hypoxia and non-hypoxic stimuli allow the formation of an active HIF-1 complex in many types of human cancers. However, the exact implication of HIF-1 activation in tumor progression is still not precisely understood. The HIF-1 dependent genic products are involved in tumoral angiogenesis, in the metabolic switch to anaerobic glycolysis and in prosurvival, proliferative and apoptotic mechanisms. In this review, we will focus on the hypoxic and non-hypoxic stimuli leading to HIF-1 activation and in its implication in tumor processes. We will highlight the most recent developments in molecular and cellular signaling that are upstream and downstream of HIF-1.


Assuntos
Translocador Nuclear Receptor Aril Hidrocarboneto/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias/fisiopatologia , Translocador Nuclear Receptor Aril Hidrocarboneto/metabolismo , Hipóxia Celular , Progressão da Doença , Humanos , Consumo de Oxigênio , Transcrição Gênica
12.
J Biol Chem ; 280(35): 31132-40, 2005 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-15994306

RESUMO

The hypoxia-inducible factor (HIF) is a key player in a transcriptional pathway that controls the hypoxic response of mammalian cells. Post-translational modification of the alpha subunit of HIF determines its half-life and activity. Among the multiple reported modifications, acetylation, by an acetyltransferase termed arrest-defective-1 protein (ARD1), has been reported to decrease HIF-1alpha stability and therefore impact on hypoxic gene expression. In contrast, we report that both overexpression and silencing of ARD1 had no impact on the stability of HIF-1alpha or -2alpha and that cells silenced for ARD1 maintained hypoxic nuclear localization of HIF-1alpha. In addition, we show that the ARD1 mRNA and protein levels are not regulated by hypoxia in several human tumor cell lines, including cervical adenocarcinoma HeLa cells, fibrosarcoma HT1080 cells, adenovirus-transformed human kidney HEK293 cells, and human breast cancer MCF-7 cells. Using two model systems ((a) wild-type and HIF-1alpha-null mouse embryo fibroblasts and (b) HeLa cells silenced for HIF-1alpha or -2alpha by RNA interference), we demonstrate that the level of expression of the ARD1 protein is independent of HIF-1alpha and -2alpha. We also demonstrate that ARD1 is a stable, predominantly cytoplasmic protein expressed in a broad range of tissues, tumor cell lines, and endothelial cells. Taken together, our findings demonstrate that ARD1 has limited, if any, impact on the HIF signaling pathway.


Assuntos
Acetiltransferases/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica , Inativação Gênica , Hipóxia , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Acetiltransferases/genética , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Humanos , Fator 1 Induzível por Hipóxia , Subunidade alfa do Fator 1 Induzível por Hipóxia , Camundongos , Camundongos Knockout , Acetiltransferase N-Terminal A , Acetiltransferase N-Terminal E , Proteínas Nucleares/genética , Interferência de RNA , Estabilidade de RNA , Transdução de Sinais/fisiologia , Distribuição Tecidual , Fatores de Transcrição/genética
13.
Int J Biochem Cell Biol ; 37(3): 535-40, 2005 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-15618010

RESUMO

Oxygen availability is crucial for cellular metabolism. Hypoxia-inducible factor 1 (HIF-1) is the major oxygen homeostasis regulator. Under normoxic conditions, HIF-1 is rapidly degraded by the proteasome. However, under hypoxic conditions, HIF-1 is stabilized and permits the activation of genes essential to cellular adaptation to low oxygen conditions. These genes include the vascular endothelial growth factor (VEGF), erythropoietin and glucose transporter-1. There is increasing evidence showing that HIF-1 is also implicated in biological functions requiring its activation under normoxic conditions. Amongst others, growth factors and vascular hormones are implicated in this normoxic activation. In this review, we will focus on differences between hypoxic and non-hypoxic induction and activation of HIF-1. We will also discuss the biological functions of HIF-1 associated with these two induction pathways. The clear understanding of both HIF-1 activation mechanisms could have a major impact in cancer and vascular disease.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica , Proteínas Nucleares/metabolismo , Transativadores/fisiologia , Fatores de Transcrição/metabolismo , Ativação Transcricional , Animais , Hipóxia Celular , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Eritropoetina/genética , Transportador de Glucose Tipo 1 , Humanos , Fator 1 Induzível por Hipóxia , Subunidade alfa do Fator 1 Induzível por Hipóxia , Modelos Biológicos , Proteínas de Transporte de Monossacarídeos/genética , Proteínas Nucleares/química , Proteínas Nucleares/genética , Oxigênio/fisiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Estrutura Terciária de Proteína , Fatores de Transcrição/química , Fatores de Transcrição/genética , Transcrição Gênica , Fatores de Crescimento do Endotélio Vascular/genética
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