RESUMEN
TANK-binding kinase 1 (TBK1) is a potential therapeutic target in multiple cancers, including clear cell renal cell carcinoma (ccRCC). However, targeting TBK1 in clinical practice is challenging. One approach to overcome this challenge would be to identify an upstream TBK1 regulator that could be targeted therapeutically in cancer specifically. In this study, we perform a kinome-wide small interfering RNA (siRNA) screen and identify doublecortin-like kinase 2 (DCLK2) as a TBK1 regulator in ccRCC. DCLK2 binds to and directly phosphorylates TBK1 on Ser172. Depletion of DCLK2 inhibits anchorage-independent colony growth and kidney tumorigenesis in orthotopic xenograft models. Conversely, overexpression of DCLK2203, a short isoform that predominates in ccRCC, promotes ccRCC cell growth and tumorigenesis in vivo. Mechanistically, DCLK2203 elicits its oncogenic signaling via TBK1 phosphorylation and activation. Taken together, these results suggest that DCLK2 is a TBK1 activator and potential therapeutic target for ccRCC.
Asunto(s)
Carcinoma de Células Renales , Neoplasias Renales , Humanos , Carcinogénesis/genética , Carcinoma de Células Renales/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Transformación Celular Neoplásica/genética , Quinasas Similares a Doblecortina , Regulación Neoplásica de la Expresión Génica , Neoplasias Renales/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismoRESUMEN
The pyruvate kinase isoforms PKM1 and PKM2 are alternatively spliced products of the PKM2 gene. PKM2, but not PKM1, alters glucose metabolism in cancer cells and contributes to tumorigenesis by mechanisms that are not explained by its known biochemical activity. We show that PKM2 gene transcription is activated by hypoxia-inducible factor 1 (HIF-1). PKM2 interacts directly with the HIF-1α subunit and promotes transactivation of HIF-1 target genes by enhancing HIF-1 binding and p300 recruitment to hypoxia response elements, whereas PKM1 fails to regulate HIF-1 activity. Interaction of PKM2 with prolyl hydroxylase 3 (PHD3) enhances PKM2 binding to HIF-1α and PKM2 coactivator function. Mass spectrometry and anti-hydroxyproline antibody assays demonstrate PKM2 hydroxylation on proline-403/408. PHD3 knockdown inhibits PKM2 coactivator function, reduces glucose uptake and lactate production, and increases O(2) consumption in cancer cells. Thus, PKM2 participates in a positive feedback loop that promotes HIF-1 transactivation and reprograms glucose metabolism in cancer cells.
Asunto(s)
Hipoxia de la Célula , Dioxigenasas/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Neoplasias/metabolismo , Piruvato Quinasa/metabolismo , Animales , Línea Celular Tumoral , Dioxigenasas/genética , Retroalimentación , Técnicas de Silenciamiento del Gen , Técnicas de Inactivación de Genes , Células HeLa , Humanos , Factor 1 Inducible por Hipoxia/metabolismo , Prolina Dioxigenasas del Factor Inducible por Hipoxia , Redes y Vías Metabólicas , Ratones , Elementos de Respuesta , Activación Transcripcional , Factores de Transcripción p300-CBP/metabolismoRESUMEN
T cell differentiation into distinct functional effector and inhibitory subsets is regulated, in part, by the cytokine environment present at the time of antigen recognition. Here, we show that hypoxia-inducible factor 1 (HIF-1), a key metabolic sensor, regulates the balance between regulatory T cell (T(reg)) and T(H)17 differentiation. HIF-1 enhances T(H)17 development through direct transcriptional activation of RORγt and via tertiary complex formation with RORγt and p300 recruitment to the IL-17 promoter, thereby regulating T(H)17 signature genes. Concurrently, HIF-1 attenuates T(reg) development by binding Foxp3 and targeting it for proteasomal degradation. Importantly, this regulation occurs under both normoxic and hypoxic conditions. Mice with HIF-1α-deficient T cells are resistant to induction of T(H)17-dependent experimental autoimmune encephalitis associated with diminished T(H)17 and increased T(reg) cells. These findings highlight the importance of metabolic cues in T cell fate determination and suggest that metabolic modulation could ameliorate certain T cell-based immune pathologies.
Asunto(s)
Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Linfocitos T Reguladores/citología , Células Th17/citología , Animales , Secuencia de Bases , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/metabolismo , Factores de Transcripción Forkhead/metabolismo , Humanos , Factor 1 Inducible por Hipoxia/metabolismo , Interleucina-17/genética , Interleucina-17/inmunología , Células Jurkat , Ratones , Datos de Secuencia Molecular , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/metabolismo , Factor de Transcripción STAT3/metabolismo , Alineación de Secuencia , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Células Th17/inmunología , Células Th17/metabolismo , Factores de Transcripción p300-CBP/metabolismoRESUMEN
Poly(ADP-ribose) polymerase-1 (PARP-1) is a DNA damage sensor and contributes to both DNA repair and cell death processes. However, how PARP-1 signaling is regulated to switch its function from DNA repair to cell death remains largely unknown. Here, we found that PARP-1 plays a central role in alkylating agent-induced PARthanatic cancer cell death. Lysine demethylase 6B (KDM6B) was identified as a key regulator of PARthanatos. Loss of KDM6B protein or its demethylase activity conferred cancer cell resistance to PARthanatic cell death in response to alkylating agents. Mechanistically, KDM6B knockout suppressed methylation at the promoter of O6-methylguanine-DNA methyltransferase (MGMT) to enhance MGMT expression and its direct DNA repair function, thereby inhibiting DNA damage-evoked PARP-1 hyperactivation and subsequent cell death. Moreover, KDM6B knockout triggered sustained Chk1 phosphorylation and activated a second XRCC1-dependent repair machinery to fix DNA damage evading from MGMT repair. Inhibition of MGMT or checkpoint response re-sensitized KDM6B deficient cells to PARthanatos induced by alkylating agents. These findings provide new molecular insights into epigenetic regulation of PARP-1 signaling mediating DNA repair or cell death and identify KDM6B as a biomarker for prediction of cancer cell vulnerability to alkylating agent treatment.
Asunto(s)
Dacarbazina , Parthanatos , Alquilantes , ADN , Reparación del ADN , Dacarbazina/farmacología , Epigénesis Genética , Guanina/análogos & derivados , O(6)-Metilguanina-ADN Metiltransferasa/genética , O(6)-Metilguanina-ADN Metiltransferasa/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Temozolomida/farmacologíaRESUMEN
OBJECTIVE: To estimate the diagnostic value of contrast-enhanced MR angiography (CE-MRA) in identifying residual brain arteriovenous malformations (AVMs) after treatment. METHODS: We retrieved appropriate references from the electronic databases of PubMed, Web of Science, Embase, and Cochrane Library, and then evaluated the methodology quality of included references using the QUADAS-2 tool. We calculated the pooled sensitivity and specificity by applying a bivariate mixed-effects model and detected the publication bias using Deeks' funnel plot. The values of I2 were used to test heterogeneity and meta-regression analyses were performed to search for the causes of heterogeneity. RESULTS: We included 7 eligible studies containing 223 participants. Compared with a gold standard, the overall sensitivity and specificity of CE-MRA in detecting residual brain AVMs were 0.77 (95% CI 0.65-0.86) and 0.97 (95% CI 0.82-1.00), respectively. Based on the summary ROC curve, the AUC was 0.89 (95% CI 0.86-0.92). Heterogeneity could be observed in our study, especially for the specificity (I2 = 74.23%). Furthermore, there was no evidence of publication bias. CONCLUSIONS: Our study provides evidence that CE-MRA has good diagnostic value and specificity for the follow-up of treated brain AVMs. Nevertheless, considering the small sample size, heterogeneity, and many factors that may affect the diagnostic accuracy, future large-sample, prospective studies are necessary to validate the results. KEY POINTS: ⢠The pooled sensitivity and specificity of contrast-enhanced MR angiography (CE-MRA) in detecting residual arteriovenous malformations (AVMs) were 0.77 (95% CI 0.65-0.86) and 0.97 (95% CI 0.82-1.00). ⢠The four-dimensional CE-MRA showed less sensitivity than the three-dimensional CE-MRA for treated AVMs. ⢠CE-MRA is helpful to identify residual AVMs and reduce excessive DSA during follow-up.
Asunto(s)
Malformaciones Arteriovenosas , Angiografía por Resonancia Magnética , Humanos , Angiografía por Resonancia Magnética/métodos , Estudios Prospectivos , Estudios de Seguimiento , Encéfalo , Sensibilidad y Especificidad , Angiografía de Substracción Digital/métodosRESUMEN
Dysregulated IL-17 expression is central to the pathogenesis of several inflammatory disorders, including ulcerative colitis. We have shown earlier that SUMOylation of ROR-γt, the transcription factor for IL-17, regulates colonic inflammation. In this study, we show that the expression of Ubc9, the E2 enzyme that targets ROR-γt for SUMOylation, is significantly reduced in the colonic mucosa of ulcerative colitis patients. Mechanistically, we demonstrate that hypoxia-inducible factor 1α (HIF-1α) binds to a CpG island within the Ubc9 gene promoter, resulting in its hypermethylation and reduced Ubc9 expression. CRISPR-Cas9-mediated inhibition of HIF-1α normalized Ubc9 and attenuated IL-17 expression in Th17 cells and reduced diseases severity in Rag1 -/- mice upon adoptive transfer. Collectively, our study reveals a novel epigenetic mechanism of regulation of ROR-γt that could be exploited in inflammatory diseases.
Asunto(s)
Colitis Ulcerosa/genética , Metilación de ADN/genética , Hipoxia/genética , Interleucina-17/genética , Regiones Promotoras Genéticas/genética , Enzimas Ubiquitina-Conjugadoras/genética , Animales , Colitis Ulcerosa/patología , Colon/patología , Humanos , Hipoxia/patología , Inflamación/genética , Inflamación/patología , Mucosa Intestinal/patología , Ratones , Ratones Endogámicos C57BL , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Células Th17RESUMEN
Cell death is a key feature of neurological diseases, including stroke and neurodegenerative disorders. Studies in a variety of ischemic/hypoxic mouse models demonstrate that poly(ADP-ribose) polymerase 1 (PARP-1)-dependent cell death, also named PARthanatos, plays a pivotal role in ischemic neuronal cell death and disease progress. PARthanatos has its unique triggers, processors, and executors that convey a highly orchestrated and programmed signaling cascade. In addition to its role in gene transcription, DNA damage repair, and energy homeostasis through PARylation of its various targets, PARP-1 activation in neuron and glia attributes to brain damage following ischemia/reperfusion. Pharmacological inhibition or genetic deletion of PARP-1 reduces infarct volume, eliminates inflammation, and improves recovery of neurological functions in stroke. Here, we reviewed the role of PARP-1 and PARthanatos in stroke and their therapeutic potential.
Asunto(s)
Accidente Cerebrovascular Isquémico/metabolismo , Accidente Cerebrovascular Isquémico/patología , Parthanatos/fisiología , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Animales , HumanosRESUMEN
Regulatory T (Treg) cells suppress inflammatory immune responses and autoimmunity caused by self-reactive T cells. The key Treg cell transcription factor Foxp3 is downregulated during inflammation to allow for the acquisition of effector T cell-like functions. Here, we demonstrate that stress signals elicited by proinflammatory cytokines and lipopolysaccharides lead to the degradation of Foxp3 through the action of the E3 ubiquitin ligase Stub1. Stub1 interacted with Foxp3 to promote its K48-linked polyubiquitination in an Hsp70-dependent manner. Knockdown of endogenous Stub1 or Hsp70 prevented Foxp3 degradation. Furthermore, the overexpression of Stub1 in Treg cells abrogated their ability to suppress inflammatory immune responses in vitro and in vivo and conferred a T-helper-1-cell-like phenotype. Our results demonstrate the critical role of the stress-activated Stub1-Hsp70 complex in promoting Treg cell inactivation, thus providing a potential therapeutic target for the intervention against autoimmune disease, infection, and cancer.
Asunto(s)
Factores de Transcripción Forkhead/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Animales , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/metabolismo , Células Cultivadas , Citocinas/metabolismo , Inhibidores Enzimáticos , Células HEK293 , Proteínas HSP70 de Choque Térmico/genética , Humanos , Imidazoles , Inflamación/genética , Inflamación/inmunología , Lipopolisacáridos/metabolismo , Ratones , Ratones Endogámicos BALB C , Fenotipo , Piridinas , Interferencia de ARN , ARN Interferente Pequeño , Linfocitos T Colaboradores-Inductores/inmunología , Ubiquitina-Proteína Ligasas/genética , UbiquitinaciónRESUMEN
Traumatic brain injury (TBI), often induced by sports, car accidents, falls, or other daily occurrences, is a primary non-genetically related risk factor for the development of subsequent neurodegeneration and neuronal cell death. However, the molecular mechanisms underlying neurodegeneration, cell death, and neurobehavioral dysfunction following TBI remain unclear. Here, we found that poly(ADP-ribose) polymerase-1 (PARP-1) was hyperactivated following TBI and its inhibition reduced TBI-induced brain injury. Macrophage migration inhibitory factor (MIF), a newly identified nuclease involved in PARP-1-dependent cell death, was translocated from the cytosol to the nucleus in cortical neurons following TBI and promoted neuronal cell death in vivo. Genetic deletion of MIF protected neurons from TBI-induced dendritic spine loss, morphological complexity degeneration, and subsequent neuronal cell death in mice. Moreover, MIF knockout reduced the brain injury volume and improved long-term animal behavioral rehabilitation. These neuroprotective effects in MIF knockout mice were reversed by the expression of wild-type MIF but not nuclease-deficient MIF mutant. In contrast, genetic deletion of MIF did not alter TBI-induced neuroinflammation. These findings reveal that MIF mediates TBI-induced neurodegeneration, neuronal cell death and neurobehavioral dysfunction through its nuclease activity, but not its pro-inflammatory role. Targeting MIF's nuclease activity may offer a novel strategy to protect neurons from TBI.
Asunto(s)
Lesiones Traumáticas del Encéfalo/metabolismo , Oxidorreductasas Intramoleculares/fisiología , Factores Inhibidores de la Migración de Macrófagos/fisiología , Degeneración Nerviosa/metabolismo , Poli(ADP-Ribosa) Polimerasa-1/fisiología , Animales , Muerte Celular , Masculino , Ratones , Ratones NoqueadosRESUMEN
Hypoxia-inducible factors (HIFs) mediate metabolic reprogramming in response to hypoxia. However, the role of HIFs in branched-chain amino acid (BCAA) metabolism remains unknown. Here we show that hypoxia upregulates mRNA and protein levels of the BCAA transporter LAT1 and the BCAA metabolic enzyme BCAT1, but not their paralogs LAT2-4 and BCAT2, in human glioblastoma (GBM) cell lines as well as primary GBM cells. Hypoxia-induced LAT1 protein upregulation is mediated by both HIF-1 and HIF-2 in GBM cells. Although both HIF-1α and HIF-2α directly bind to the hypoxia response element at the first intron of the human BCAT1 gene, HIF-1α is exclusively responsible for hypoxia-induced BCAT1 expression in GBM cells. Knockout of HIF-1α and HIF-2α significantly reduces glutamate labeling from BCAAs in GBM cells under hypoxia, which provides functional evidence for HIF-mediated reprogramming of BCAA metabolism. Genetic or pharmacological inhibition of BCAT1 inhibits GBM cell growth under hypoxia. Together, these findings uncover a previously unrecognized HIF-dependent metabolic pathway that increases GBM cell growth under conditions of hypoxic stress.
Asunto(s)
Aminoácidos de Cadena Ramificada/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/deficiencia , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Sistemas CRISPR-Cas/genética , Hipoxia de la Célula , Proliferación Celular , Células Cultivadas , Regulación Neoplásica de la Expresión Génica , Técnicas de Inactivación de Genes , Glioblastoma/metabolismo , Glioblastoma/patología , Ácido Glutámico/metabolismo , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/deficiencia , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Transportador de Aminoácidos Neutros Grandes 1/genética , Transportador de Aminoácidos Neutros Grandes 1/metabolismo , Unión Proteica , Transaminasas/antagonistas & inhibidores , Transaminasas/genética , Transaminasas/metabolismoRESUMEN
Bacteriocins derived from lactic acid bacteria (LAB) are well recognized as promising food preservative due to high safety and potent antibacterial activity against foodborne pathogens and spoilage bacteria. In this study, an antimicrobial agent-producing strain FZU63 from Chinese sauerkraut was identified as Lactobacillus coryniformis based on physio-biochemical characterization and 16S rDNA sequence analysis. In addition, a bacteriocin was purified from the culture supernatant of L. coryniformis FZU63, and its molecular mass was determined as 1493.709 Da. Moreover, the amino acid sequence of the bacteriocin was predicted to be RQQPMTLDYRW-NH2 using nanoliter/microliter liquid chromatography combined with triple quadrupole-linear ion trap tandem mass spectrometry and was named as Lactocin 63. Furthermore, Lactocin 63 displays potent antimicrobial activity against the tested Gram-positive and negative bacteria based on the results of determining MICs. Subsequently, the action mode of Lactocin 63 against Shewanella putrefaciens was investigated. The results demonstrated that Lactocin 63 targets and is adsorbed onto the bacterial cell wall and membrane and then disrupts cytoplasmic membrane, which is leading to leakage of cytoplasm according to the results of flow cytometry analysis and the observation of cellular ultra-structure using confocal laser microscopy and atomic force microscopy. Collectively, these results are helpful and providing the theoretical base for developing and applying LAB-derived bacteriocins as promising bio-preservatives to combat foodborne pathogens and spoilage bacteria in seafood industries.Key points⢠A bacteriocin-producing strain Lactobacillus coryniformis was isolated.⢠A novel bacteriocin produced by Lactobacillus coryniformis FZU63 was characterized.⢠Action mechanism of the bacteriocin against S. putrefaciens was elucidated in vitro.
Asunto(s)
Antiinfecciosos , Bacteriocinas , Shewanella putrefaciens , Antibacterianos/farmacología , Bacteriocinas/genética , Bacteriocinas/farmacología , LactobacillusRESUMEN
Fengycin, as a lipopeptide produced by Bacillus subtilis, displays potent activity against filamentous fungi, including Aspergillus flavus and Soft-rot fungus, which exhibits a wide range of potential applications in food industries, agriculture, and medicine. To better clarify the regulatory mechanism of fructose on fengycin biosynthesis, the iTRAQ-based proteomic analysis was utilized to investigate the differentially expressed proteins of B. amyloliquefaciens fmb-60 cultivated in ML (without fructose) and MLF (with fructose) medium. The results indicated that a total of 811 proteins, including 248 proteins with differential expression levels (162 which were upregulated (fold > 2) and 86, which were downregulated (fold < 0.5) were detected, and most of the proteins are associated with cellular metabolism, biosynthesis, and biological regulation process. Moreover, the target genes' relative expression was conducted using quantitative real-time PCR to validate the proteomic analysis results. Based on the results of proteome analysis, the supposed pathways of fructose enhancing fengycin biosynthesis in B. amyloliquefaciens fmb-60 can be summarized as improvement of the metabolic process, including cellular amino acid and amide, fatty acid biosynthesis, peptide and protein, nucleotide and nucleobase-containing compound, drug/toxin, cofactor, and vitamin; reinforcement of peptide/protein translation, modification, biological process, and response to a stimulus. In conclusion, this study represents a comprehensive and systematic investigation of the fructose mechanism on improving fengycin biosynthesis in B. amyloliquefaciens, which will provide a road map to facilitate the potential application of fengycin or its homolog in defending against filamentous fungi.
Asunto(s)
Bacillus amyloliquefaciens/metabolismo , Proteínas Bacterianas/metabolismo , Fructosa/metabolismo , Lipopéptidos/metabolismo , Proteómica/métodosRESUMEN
Hypoxia-inducible factor 1 (HIF-1) is a master transcriptional regulator in response to hypoxia and its transcriptional activity is crucial for cancer cell mobility. Here we present evidence for a novel epigenetic mechanism that regulates HIF-1 transcriptional activity and HIF-1-dependent migration of glioblastoma cells. The lysine methyltransferases G9a and GLP directly bound to the α subunit of HIF-1 (HIF-1α) and catalyzed mono- and di-methylation of HIF-1α at lysine (K) 674 in vitro and in vivo. K674 methylation suppressed HIF-1 transcriptional activity and expression of its downstream target genes PTGS1, NDNF, SLC6A3, and Linc01132 in human glioblastoma U251MG cells. Inhibition of HIF-1 by K674 methylation is due to reduced HIF-1α transactivation domain function but not increased HIF-1α protein degradation or impaired binding of HIF-1 to hypoxia response elements. K674 methylation significantly decreased HIF-1-dependent migration of U251MG cells under hypoxia. Importantly, we found that G9a was downregulated by hypoxia in glioblastoma, which was inversely correlated with PTGS1 expression and survival of patients with glioblastoma. Therefore, our findings uncover a hypoxia-induced negative feedback mechanism that maintains high activity of HIF-1 and cell mobility in human glioblastoma.
Asunto(s)
Autoantígenos/metabolismo , Epigénesis Genética , Regulación Neoplásica de la Expresión Génica , Glioblastoma/genética , Proteínas de la Matriz de Golgi/metabolismo , Antígenos de Histocompatibilidad/metabolismo , N-Metiltransferasa de Histona-Lisina/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Transcripción Genética , Hipoxia de la Célula , Línea Celular , Movimiento Celular , Glioblastoma/metabolismo , Glioblastoma/fisiopatología , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/antagonistas & inhibidores , Subunidad alfa del Factor 1 Inducible por Hipoxia/química , Lisina/metabolismo , Metilación , Elementos de RespuestaRESUMEN
MCM proteins are components of a DNA helicase that plays an essential role in DNA replication and cell proliferation. However, MCM proteins are present in excess relative to origins of replication, suggesting they may serve other functions. Decreased proliferation is a fundamental physiological response to hypoxia in many cell types, and hypoxia-inducible factor 1 (HIF-1) has been implicated in this process. Here, we demonstrate that multiple MCM proteins bind directly to the HIF-1α subunit and synergistically inhibit HIF-1 transcriptional activity via distinct O(2)-dependent mechanisms. MCM3 inhibits transactivation domain function, whereas MCM7 enhances HIF-1α ubiquitination and proteasomal degradation. HIF-1 activity decreases when quiescent cells re-enter the cell cycle, and this effect is MCM dependent. Exposure to hypoxia leads to MCM2-7 downregulation in diverse cell types. These studies reveal a function of MCM proteins apart from their DNA helicase activity and establish a direct link between HIF-1 and the cell-cycle machinery.
Asunto(s)
Proteínas de Ciclo Celular/fisiología , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Proteínas Nucleares/fisiología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/antagonistas & inhibidores , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Línea Celular , Regulación de la Expresión Génica , Células HEK293 , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/antagonistas & inhibidores , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Ratones , Células 3T3 NIH , Oxígeno/metabolismo , Transcripción Genética , UbiquitinaciónRESUMEN
The hypoxia-inducible factor (HIF) is a heterodimeric transcription factor governing a transcriptional program in response to reduced O2 availability in metazoans. It contributes to physiology and pathogenesis of many human diseases through its downstream target genes. Emerging studies have shown that the transcriptional activity of HIF is highly regulated at multiple levels and the epigenetic regulators are essential for HIF-mediated transactivation. In this review, we will discuss the comprehensive regulation of HIF transcriptional activity by different types of epigenetic regulators.
Asunto(s)
Epigénesis Genética , Histona Desacetilasa 1/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Hipoxia/genética , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Factores de Transcripción p300-CBP/genética , Animales , Células HeLa , Histona Desacetilasa 1/metabolismo , Humanos , Hipoxia/metabolismo , Hipoxia/patología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Oxígeno/farmacología , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estabilidad Proteica , Transcripción Genética , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Factores de Transcripción p300-CBP/metabolismoRESUMEN
Hypoxia is a hallmark of the tumor microenvironment and contributes to tumor malignant phenotypes. Hypoxia-inducible factor (HIF) is a master regulator of intratumoral hypoxia and controls hypoxia-mediated pathological processes in tumors, including angiogenesis, metabolic reprogramming, epigenetic reprogramming, immune evasion, pH homeostasis, cell migration/invasion, stem cell pluripotency, and therapy resistance. In this book chapter, we reviewed the causes and types of intratumoral hypoxia, hypoxia detection methods, and the oncogenic role of HIF in tumorigenesis and chemo- and radio-therapy resistance.
Asunto(s)
Subunidad alfa del Factor 1 Inducible por Hipoxia , Neoplasias/patología , Hipoxia Tumoral , Microambiente Tumoral , Hipoxia de la Célula , Humanos , Neovascularización PatológicaRESUMEN
Atherosclerosis is a complicated process comprising inflammation, accumulation of collagen matrix and aberrant DNA methylation. SMAD7 is known to play an important role in fibrosis and inflammation. In recent years, increasing research has concentrated on the connection between DNA methylation and atherosclerosis. The current study was designed to investigate methylation status of some specific gene with a focus on SMAD7 in atherosclerosis and elucidate their relationship. We found that SMAD7 expression was decreased and its promoter region was markedly methylated in atherosclerotic plaques when compared with normal artery walls. Using MALDI-TOF MS, increased DNA methylation levels of SMAD7 promoter at CpG unit 5.8.15.16 were found in peripheral blood of atherosclerosis patients relative to matched normal controls, respectively. Correlation analysis revealed that mean DNA methylation levels of SMAD7 promoter of CpG unit 5.8.15.16 were positively associated with homocysteine levels (râ¯=â¯0.724, pâ¯<â¯.001) and carotid plaque scores(râ¯=â¯0.790, pâ¯<â¯.001). SMAD7 promoter is hyper-methylated both in human atherosclerotic plaques and atherosclerosis patients, which is positively associated with homocysteine levels and carotid plaque scores. Thus, methylated SMAD7 may be a novel predicted marker and therapeutics target for atherosclerosis.
Asunto(s)
Aterosclerosis/diagnóstico , Aterosclerosis/genética , Metilación de ADN , Proteína smad7/genética , Anciano , Aterosclerosis/patología , Secuencia de Bases , Islas de CpG , Femenino , Marcadores Genéticos/genética , Humanos , Masculino , Placa Aterosclerótica/diagnóstico , Placa Aterosclerótica/genética , Placa Aterosclerótica/patología , Pronóstico , Regiones Promotoras Genéticas , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
Extracellular vesicles such as exosomes and microvesicles (MVs) are shed by cancer cells, are detected in the plasma of cancer patients, and promote cancer progression, but the molecular mechanisms regulating their production are not well understood. Intratumoral hypoxia is common in advanced breast cancers and is associated with an increased risk of metastasis and patient mortality that is mediated in part by the activation of hypoxia-inducible factors (HIFs). In this paper, we report that exposure of human breast cancer cells to hypoxia augments MV shedding that is mediated by the HIF-dependent expression of the small GTPase RAB22A, which colocalizes with budding MVs at the cell surface. Incubation of naïve breast cancer cells with MVs shed by hypoxic breast cancer cells promotes focal adhesion formation, invasion, and metastasis. In breast cancer patients, RAB22A mRNA overexpression in the primary tumor is associated with decreased overall and metastasis-free survival and, in an orthotopic mouse model, RAB22A knockdown impairs breast cancer metastasis.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Exosomas/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Proteínas de Unión al GTP rab/metabolismo , Animales , Neoplasias de la Mama/genética , Hipoxia de la Célula/genética , Línea Celular Tumoral , Matriz Extracelular/metabolismo , Femenino , Adhesiones Focales/metabolismo , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/secundario , Ratones , Ratones SCID , Invasividad Neoplásica , Metástasis de la Neoplasia , Transporte de Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Análisis de Supervivencia , Proteínas de Unión al GTP rab/deficiencia , Proteínas de Unión al GTP rab/genéticaRESUMEN
Both preclinical and clinical studies suggest that brief cycles of ischemia and reperfusion in the arm or leg may protect the heart against injury following prolonged coronary artery occlusion and reperfusion, a phenomenon known as remote ischemic preconditioning. Recent studies in mice indicate that increased plasma interleukin-10 (IL-10) levels play an important role in remote ischemic preconditioning induced by clamping the femoral artery for 5 min followed by 5 min of reperfusion for a total of three cycles. In this study, we demonstrate that remote ischemic preconditioning increases plasma IL-10 levels and decreases myocardial infarct size in wild-type mice but not in littermates that are heterozygous for a knockout allele at the locus encoding hypoxia-inducible factor (HIF) 1α. Injection of a recombinant adenovirus encoding a constitutively active form of HIF-1α into mouse hind limb muscle was sufficient to increase plasma IL-10 levels and decrease myocardial infarct size. Exposure of C2C12 mouse myocytes to cyclic hypoxia and reoxygenation rapidly increased levels of IL-10 mRNA, which was blocked by administration of the HIF-1 inhibitor acriflavine or by expression of short hairpin RNA targeting HIF-1α or HIF-1ß. Chromatin immunoprecipitation assays demonstrated that binding of HIF-1 to the Il10 gene was induced when myocytes were subjected to cyclic hypoxia and reoxygenation. Taken together, these data indicate that HIF-1 activates Il10 gene transcription and is required for remote ischemic preconditioning.
Asunto(s)
Factor 1 Inducible por Hipoxia/metabolismo , Interleucina-10/metabolismo , Precondicionamiento Isquémico Miocárdico/métodos , Infarto del Miocardio/metabolismo , Acriflavina/farmacología , Animales , Translocador Nuclear del Receptor de Aril Hidrocarburo/antagonistas & inhibidores , Translocador Nuclear del Receptor de Aril Hidrocarburo/genética , Translocador Nuclear del Receptor de Aril Hidrocarburo/metabolismo , Hipoxia de la Célula , Línea Celular , Factor 1 Inducible por Hipoxia/antagonistas & inhibidores , Factor 1 Inducible por Hipoxia/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/antagonistas & inhibidores , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Immunoblotting , Interleucina-10/sangre , Interleucina-10/genética , Ratones , Ratones Noqueados , Mioblastos Esqueléticos/citología , Mioblastos Esqueléticos/efectos de los fármacos , Mioblastos Esqueléticos/metabolismo , Infarto del Miocardio/genética , Infarto del Miocardio/prevención & control , Miocardio/metabolismo , Miocardio/patología , Fosforilación , Unión Proteica , Proteínas Proto-Oncogénicas c-akt/metabolismo , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Increased catalytic activity of CBS (cystathionine ß-synthase) was recently shown to mediate vasodilation of the cerebral microcirculation, which is initiated within minutes of the onset of acute hypoxia. To test whether chronic hypoxia was a stimulus for increased CBS expression, U87-MG human glioblastoma and PC12 rat phaeochromocytoma cells were exposed to 1% or 20% O2 for 24-72 h. CBS mRNA and protein expression were increased in hypoxic cells. Hypoxic induction of CBS expression was abrogated in cells transfected with vector encoding shRNA targeting HIF (hypoxia-inducible factor) 1α or 2α. Exposure of rats to hypobaric hypoxia (0.35 atm; 1 atm=101.325 kPa) for 3 days induced increased CBS mRNA, protein and catalytic activity in the cerebral cortex and cerebellum, which was blocked by administration of the HIF inhibitor digoxin. HIF-binding sites, located 0.8 and 1.2 kb 5' to the transcription start site of the human CBS and rat Cbs genes respectively, were identified by ChIP assays. A 49-bp human sequence, which encompassed an inverted repeat of the core HIF-binding site, functioned as a hypoxia-response element in luciferase reporter transcription assays. Thus HIFs mediate tissue-specific CBS expression, which may augment cerebral vasodilation as an adaptive response to chronic hypoxia.