RESUMO
Ferroptosis is a type of regulated cell death characterized by iron-dependent lipid peroxidation. A model cell system is constructed to induce ferroptosis by re-expressing the transcription factor BACH1, a potent ferroptosis inducer, in immortalized mouse embryonic fibroblasts (iMEFs). The transfer of the culture supernatant from ferroptotic iMEFs activates the proliferation of hepatoma cells and other fibroblasts and suppresses cellular senescence-like features. The BACH1-dependent secretion of the longevity factor FGF21 is increased in ferroptotic iMEFs. The anti-senescent effects of the culture supernatant from these iMEFs are abrogated by Fgf21 knockout. BACH1 activates the transcription of Fgf21 by promoting ferroptotic stress and increases FGF21 protein expression by suppressing its autophagic degradation through transcriptional Sqstm1 and Lamp2 repression. The BACH1-induced ferroptotic FGF21 secretion suppresses obesity in high-fat diet-fed mice and the short lifespan of progeria mice. The inhibition of these aging-related phenotypes can be physiologically significant regarding ferroptosis.
Assuntos
Fatores de Transcrição de Zíper de Leucina Básica , Senescência Celular , Ferroptose , Fatores de Crescimento de Fibroblastos , Obesidade , Animais , Ferroptose/genética , Fatores de Crescimento de Fibroblastos/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Obesidade/metabolismo , Obesidade/patologia , Camundongos , Longevidade , Humanos , Camundongos Endogâmicos C57BL , Fibroblastos/metabolismo , Autofagia , Dieta Hiperlipídica , Camundongos Knockout , Masculino , Proteína Sequestossoma-1/metabolismoRESUMO
BTB and CNC homology 1 (BACH1) represses the expression of genes involved in the metabolism of iron, heme and reactive oxygen species. While BACH1 is rapidly degraded when it is bound to heme, it remains unclear how BACH1 degradation is regulated under other conditions. We found that FBXO22, a ubiquitin ligase previously reported to promote BACH1 degradation, polyubiquitinated BACH1 only in the presence of heme in a highly purified reconstitution assay. In parallel to this regulatory mechanism, TANK binding kinase 1 (TBK1), a protein kinase that activates innate immune response and regulates iron metabolism via ferritinophagy, was found to promote BACH1 degradation when overexpressed in 293T cells. While TBK1 phosphorylated BACH1 at multiple serine and threonine residues, BACH1 degradation was observed with not only the wild-type TBK1 but also catalytically impaired TBK1. The BACH1 degradation in response to catalytically impaired TBK1 was not dependent on FBXO22 but involved both autophagy-lysosome and ubiquitin-proteasome pathways judging from its suppression by using inhibitors of lysosome and proteasome. Chemical inhibition of TBK1 in hepatoma Hepa1 cells showed that TBK1 was not required for the heme-induced BACH1 degradation. Its inhibition in Namalwa B lymphoma cells increased endogenous BACH1 protein. These results suggest that TBK1 promotes BACH1 degradation in parallel to the FBXO22- and heme-dependent pathway, placing BACH1 as a downstream effector of TBK1 in iron metabolism or innate immune response.
Assuntos
Fatores de Transcrição de Zíper de Leucina Básica , Proteínas F-Box , Heme , Proteínas Serina-Treonina Quinases , Proteólise , Receptores Citoplasmáticos e Nucleares , Humanos , Heme/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fosforilação , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Células HEK293 , Ubiquitinação , Linhagem Celular Tumoral , Lisossomos/metabolismo , Autofagia , Complexo de Endopeptidases do Proteassoma/metabolismoRESUMO
Ferroptosis is a regulated cell death induced by iron-dependent lipid peroxidation. The heme-responsive transcription factor BTB and CNC homology 1 (BACH1) promotes ferroptosis by repressing the transcription of genes involved in glutathione (GSH) synthesis and intracellular labile iron metabolism, which are key regulatory pathways in ferroptosis. We found that BACH1 re-expression in Bach1-/- immortalized mouse embryonic fibroblasts (iMEFs) can induce ferroptosis upon 2-mercaptoethanol removal, without any ferroptosis inducers. In these iMEFs, GSH synthesis was reduced, and intracellular labile iron levels were increased upon BACH1 re-expression. We used this system to investigate whether the major ferroptosis regulators glutathione peroxidase 4 (Gpx4) and apoptosis-inducing factor mitochondria-associated 2 (Aifm2), the gene for ferroptosis suppressor protein 1, are target genes of BACH1. Neither Gpx4 nor Aifm2 was regulated by BACH1 in the iMEFs. However, we found that BACH1 represses AIFM2 transcription in human pancreatic cancer cells. These results suggest that the ferroptosis regulators targeted by BACH1 may vary across different cell types and animal species. Furthermore, we confirmed that the ferroptosis induced by BACH1 re-expression exhibited a propagating effect. BACH1 re-expression represents a new strategy for inducing ferroptosis after GPX4 or system Xc- suppression and is expected to contribute to future ferroptosis research.
Assuntos
Ferroptose , Fibroblastos , Animais , Humanos , Camundongos , Fibroblastos/metabolismo , Ferroptose/genética , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Ferro/metabolismo , Glutationa/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismoRESUMO
Although establishment and maintenance of mitochondria are essential for the production of massive amounts of heme in erythroblasts, mitochondria must be degraded upon terminal differentiation to red blood cells (RBCs), thus creating a biphasic regulatory process. Previously, we reported that iron deficiency in mice promotes mitochondrial retention in RBCs, suggesting that a proper amount of iron and/or heme is necessary for the degradation of mitochondria during erythroblast maturation. Because the transcription factor GATA1 regulates autophagy in erythroid cells, which involves mitochondrial clearance (mitophagy), we investigated the relationship between iron or heme and mitophagy by analyzing the expression of genes related to GATA1 and autophagy and the impact of iron or heme restriction on the amount of mitochondria. We found that heme promotes the expression of GATA1-regulated mitophagy-related genes and the induction of mitophagy. GATA1 might induce the expression of the autophagy-related genes Atg4d and Stk11 for mitophagy through a heme-dependent mechanism in murine erythroleukemia (MEL) cells and a genetic rescue system with G1E-ER-GATA1 erythroblast cells derived from Gata1-null murine embryonic stem cells. These results provide evidence for a biphasic mechanism in which mitochondria are essential for heme generation, and the heme generated during differentiation promotes mitophagy and mitochondrial disposal. This mechanism provides a molecular framework for understanding this fundamentally important cell biological process.
Assuntos
Heme , Mitofagia , Camundongos , Animais , Heme/metabolismo , Diferenciação Celular , Células Eritroides/metabolismo , Ferro/metabolismoRESUMO
The transcription factor BACH1, which is regulated by direct binding of prosthetic group heme, promotes epithelial-mesenchymal transition (EMT) and drives metastasis of diverse types of cancer cells. De-regulated target genes of BACH1 in cancer cells include those for glycolysis, oxidative phosphorylation, epithelial cell adhesion, and mesodermal cell motility. In addition, the canonical target genes of BACH1 include genes for the regulation of iron homeostasis. Importantly, cancer cells are addicted to iron. We summarize known functions of BACH1 in cancer and discuss how BACH1 may affect iron homeostasis in cancer cells to support their progression by increasing mobile iron within cells. The dependency on BACH1 for cancer progression may also confer upon cancer cells susceptibility to iron-dependent cell death ferroptosis. Finally, we discuss that the human transcription factors provide research opportunities for better understanding of cancer cell properties.
Assuntos
Ferro , Neoplasias , Humanos , Ferro/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Heme/química , Heme/metabolismo , Neoplasias/genética , Fatores de Transcrição/metabolismo , HomeostaseRESUMO
Parkinson's disease (PD) is the second most common neurodegenerative movement disorder characterized by a progressive loss of dopaminergic neurons in the substantia nigra pars compacta. Although a complex interplay of multiple environmental and genetic factors has been implicated, the etiology of neuronal death in PD remains unresolved. Various mechanisms of neuronal degeneration in PD have been proposed, including oxidative stress, mitochondrial dysfunction, neuroinflammation, α-synuclein proteostasis, disruption of calcium homeostasis, and other cell death pathways. While many drugs individually targeting these pathways have shown promise in preclinical PD models, this promise has not yet translated into neuroprotective therapies in human PD. This has consequently spurred efforts to identify alternative targets with multipronged therapeutic approaches. A promising therapeutic target that could modulate multiple etiological pathways involves drug-induced activation of a coordinated genetic program regulated by the transcription factor, nuclear factor E2-related factor 2 (Nrf2). Nrf2 regulates the transcription of over 250 genes, creating a multifaceted network that integrates cellular activities by expressing cytoprotective genes, promoting the resolution of inflammation, restoring redox and protein homeostasis, stimulating energy metabolism, and facilitating repair. However, FDA-approved electrophilic Nrf2 activators cause irreversible alkylation of cysteine residues in various cellular proteins resulting in side effects. We propose that the transcriptional repressor of BTB and CNC homology 1 (Bach1), which antagonizes Nrf2, could serve as a promising complementary target for the activation of both Nrf2-dependent and Nrf2-independent neuroprotective pathways. This review presents the current knowledge on the Nrf2/Bach1 signaling pathway, its role in various cellular processes, and the benefits of simultaneously inhibiting Bach1 and stabilizing Nrf2 using non-electrophilic small molecules as a novel therapeutic approach for PD.
RESUMO
BTB and CNC homology 1 (BACH1) represses the expression of genes involved in the metabolism of iron, heme and reactive oxygen species and promotes metastasis of various cancers including pancreatic ductal adenocarcinoma (PDAC). However, it is not clear how BACH1 is regulated in PDAC cells. Knockdown of Tank binding kinase 1 (TBK1) led to reductions of BACH1 mRNA and protein amounts in AsPC-1 human PDAC cells. Gene expression analysis of PDAC cells with knockdown of TBK1 or BACH1 suggested the involvement of TBK1 and BACH1 in the regulation of iron homeostasis. Ferritin mRNA and proteins were both increased upon BACH1 knockdown in AsPC-1 cells. Flow cytometry analysis showed that AsPC-1 cells with BACH1 knockout or knockdown contained lower labile iron than control cells, suggesting that BACH1 increased labile iron by repressing the expression of ferritin genes. We further found that the expression of E-cadherin was upregulated upon the chelation of intracellular iron content. These results suggest that the TBK1-BACH1 pathway promotes cancer cell metastasis by increasing labile iron within cells.
RESUMO
Methionine adenosyltransferase (MAT) catalyzes the synthesis of S-adenosylmethionine (SAM). As the sole methyl-donor for methylation of DNA, RNA, and proteins, SAM levels affect gene expression by changing methylation patterns. Expression of MAT2A, the catalytic subunit of isozyme MAT2, is positively correlated with proliferation of cancer cells; however, how MAT2A promotes cell proliferation is largely unknown. Given that the protein synthesis is induced in proliferating cells and that RNA and protein components of translation machinery are methylated, we tested here whether MAT2 and SAM are coupled with protein synthesis. By measuring ongoing protein translation via puromycin labeling, we revealed that MAT2A depletion or chemical inhibition reduced protein synthesis in HeLa and Hepa1 cells. Furthermore, overexpression of MAT2A enhanced protein synthesis, indicating that SAM is limiting under normal culture conditions. In addition, MAT2 inhibition did not accompany reduction in mechanistic target of rapamycin complex 1 activity but nevertheless reduced polysome formation. Polysome-bound RNA sequencing revealed that MAT2 inhibition decreased translation efficiency of some fraction of mRNAs. MAT2A was also found to interact with the proteins involved in rRNA processing and ribosome biogenesis; depletion or inhibition of MAT2 reduced 18S rRNA processing. Finally, quantitative mass spectrometry revealed that some translation factors were dynamically methylated in response to the activity of MAT2A. These observations suggest that cells possess an mTOR-independent regulatory mechanism that tunes translation in response to the levels of SAM. Such a system may acclimate cells for survival when SAM synthesis is reduced, whereas it may support proliferation when SAM is sufficient.
Assuntos
Metionina Adenosiltransferase , S-Adenosilmetionina , Animais , Humanos , Mamíferos/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Metionina/metabolismo , Metionina Adenosiltransferase/genética , Metionina Adenosiltransferase/metabolismo , Metilação , RNA Mensageiro/metabolismo , S-Adenosilmetionina/metabolismoRESUMO
Parkinson's disease (PD) is a progressive neurodegenerative movement disorder characterized by the loss of nigrostriatal dopaminergic neurons. Mounting evidence suggests that Nrf2 is a promising target for neuroprotective interventions in PD. However, electrophilic chemical properties of the canonical Nrf2-based drugs cause irreversible alkylation of cysteine residues on cellular proteins resulting in side effects. Bach1 is a known transcriptional repressor of the Nrf2 pathway. We report that Bach1 levels are up-regulated in PD postmortem brains and preclinical models. Bach1 knockout (KO) mice were protected against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced dopaminergic neurotoxicity and associated oxidative damage and neuroinflammation. Functional genomic analysis demonstrated that the neuroprotective effects in Bach1 KO mice was due to up-regulation of Bach1-targeted pathways that are associated with both Nrf2-dependent antioxidant response element (ARE) and Nrf2-independent non-ARE genes. Using a proprietary translational technology platform, a drug library screen identified a substituted benzimidazole as a Bach1 inhibitor that was validated as a nonelectrophile. Oral administration of the Bach1 inhibitor attenuated MPTP neurotoxicity in pre- and posttreatment paradigms. Bach1 inhibitor-induced neuroprotection was associated with the up-regulation of Bach1-targeted pathways in concurrence with the results from Bach1 KO mice. Our results suggest that genetic deletion as well as pharmacologic inhibition of Bach1 by a nonelectrophilic inhibitor is a promising therapeutic approach for PD.
Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Neuroproteção , Doença de Parkinson/terapia , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina , Idoso , Idoso de 80 Anos ou mais , Animais , Elementos de Resposta Antioxidante , Fatores de Transcrição de Zíper de Leucina Básica/antagonistas & inibidores , Fatores de Transcrição de Zíper de Leucina Básica/genética , Estudos de Casos e Controles , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Doença de Parkinson/metabolismo , RatosRESUMO
The progression of cancer involves not only the gradual evolution of cells by mutations in DNA but also alterations in the gene expression induced by those mutations and input from the surrounding microenvironment. Such alterations contribute to cancer cells' abilities to reprogram metabolic pathways and undergo epithelial-to-mesenchymal transition (EMT), which facilitate the survival of cancer cells and their metastasis to other organs. Recently, BTB and CNC homology 1 (BACH1), a heme-regulated transcription factor that represses genes involved in iron and heme metabolism in normal cells, was shown to shape the metabolism and metastatic potential of cancer cells. The growing list of BACH1 target genes in cancer cells reveals that BACH1 promotes metastasis by regulating various sets of genes beyond iron metabolism. BACH1 represses the expression of genes that mediate cell-cell adhesion and oxidative phosphorylation but activates the expression of genes required for glycolysis, cell motility, and matrix protein degradation. Furthermore, BACH1 represses FOXA1 gene encoding an activator of epithelial genes and activates SNAI2 encoding a repressor of epithelial genes, forming a feedforward loop of EMT. By synthesizing these observations, we propose a "two-faced BACH1 model", which accounts for the dynamic switching between metastasis and stress resistance along with cancer progression. We discuss here the possibility that BACH1-mediated promotion of cancer also brings increased sensitivity to iron-dependent cell death (ferroptosis) through crosstalk of BACH1 target genes, imposing programmed vulnerability upon cancer cells. We also discuss the future directions of this field, including the dynamics and plasticity of EMT.
Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/fisiologia , Transição Epitelial-Mesenquimal/fisiologia , Ferroptose , Neoplasias/patologia , Fatores de Transcrição de Zíper de Leucina Básica/química , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Progressão da Doença , Heme/metabolismo , Humanos , Metástase Neoplásica , Estresse Oxidativo , Relação Estrutura-AtividadeRESUMO
Ferroptosis regulated cell death due to the iron-dependent accumulation of lipid peroxide. Ferroptosis is known to constitute the pathology of ischemic diseases, neurodegenerative diseases, and steatohepatitis and also works as a suppressing mechanism against cancer. However, how ferroptotic cells affect surrounding cells remains elusive. We herein report the transfer phenomenon of lipid peroxidation and cell death from ferroptotic cells to nearby cells that are not exposed to ferroptotic inducers (FINs). While primary mouse embryonic fibroblasts (MEFs) and NIH3T3 cells contained senescence-associated ß-galactosidase (SA-ß-gal)-positive cells, they were decreased upon induction of ferroptosis with FINs. The SA-ß-gal decrease was inhibited by ferroptotic inhibitors and knockdown of Atg7, pointing to the involvement of lipid peroxidation and activated autophagosome formation during ferroptosis. A transfer of cell culture medium of cells treated with FINs, type 1 or 2, caused the reduction in SA-ß-gal-positive cells in recipient cells that had not been exposed to FINs. Real-time imaging of Kusabira Orange-marked reporter MEFs cocultured with ferroptotic cells showed the generation of lipid peroxide and deaths of the reporter cells. These results indicate that lipid peroxidation and its aftereffects propagate from ferroptotic cells to surrounding cells, even when the surrounding cells are not exposed to FINs. Ferroptotic cells are not merely dying cells but also work as signal transmitters inducing a chain of further ferroptosis.
Assuntos
Autofagia/genética , Morte Celular/genética , Ferroptose/genética , Peroxidação de Lipídeos/genética , Animais , Humanos , CamundongosRESUMO
The chromatin protein positive coactivator 4 (PC4) has multiple functions, including chromatin compaction. However, its role in immune cells is largely unknown. We show that PC4 orchestrates chromatin structure and gene expression in mature B cells. B-cell-specific PC4-deficient mice show impaired production of antibody upon antigen stimulation. The PC4 complex purified from B cells contains the transcription factors (TFs) IKAROS and IRF4. IKAROS protein is reduced in PC4-deficient mature B cells, resulting in de-repression of their target genes in part by diminished interactions with gene-silencing components. Upon activation, the amount of IRF4 protein is not increased in PC4-deficient B cells, resulting in reduction of plasma cells. Importantly, IRF4 reciprocally induces PC4 expression via a super-enhancer. PC4 knockdown in human B cell lymphoma and myeloma cells reduces IKAROS protein as an anticancer drug, lenalidomide. Our findings establish PC4 as a chromatin regulator of B cells and a possible therapeutic target adjoining IKAROS in B cell malignancies.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Fator de Transcrição Ikaros/metabolismo , Fatores Reguladores de Interferon/metabolismo , Fatores de Transcrição/metabolismo , Animais , Linfócitos B/metabolismo , Linfócitos B/patologia , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos TransgênicosRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is among the cancers with the poorest prognoses due to its highly malignant features. BTB and CNC homology 1 (BACH1) has been implicated in RAS-driven tumor formation. We focused on the role of BACH1 in PDAC, more than 90% of which have KRAS mutation. Knockdown of BACH1 in PDAC cell lines reduced cell migration and invasion, in part, by increasing E-cadherin expression, whereas its overexpression showed opposite effects. BACH1 directly repressed the expression of FOXA1 that is known to activate the expression of CDH1 encoding E-cadherin and to inhibit epithelial-to-mesenchymal transition. BACH1 also directly repressed the expression of genes important for epithelial cell adhesion including CLDN3 and CLDN4. In a mouse orthotopic implantation model, BACH1 was required for the high metastatic ability of AsPC-1 cells. IHC analysis of clinical specimens with a newly developed anti-BACH1 mAb revealed that high expression of BACH1 is a poor prognostic factor. These results suggest that the gene regulatory network of BACH1 and downstream genes including CDH1 contribute to the malignant features of PDAC by regulating epithelial-to-mesenchymal transition. SIGNIFICANCE: Greater understanding of the gene regulatory network involved in epithelial-to-mesenchymal transition of pancreatic cancer cells will provide novel therapeutic targets and diagnostic markers.
Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Carcinoma Ductal Pancreático/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pancreáticas/genética , Animais , Antígenos CD/genética , Fatores de Transcrição de Zíper de Leucina Básica/genética , Caderinas/genética , Carcinoma Ductal Pancreático/mortalidade , Carcinoma Ductal Pancreático/patologia , Movimento Celular/genética , Claudina-3/genética , Claudina-4/genética , Feminino , Técnicas de Inativação de Genes , Redes Reguladoras de Genes , Fator 3-alfa Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Humanos , Estimativa de Kaplan-Meier , Masculino , Camundongos , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Metástase Neoplásica/genética , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Prognóstico , RNA-SeqRESUMO
Ferroptosis is an iron-dependent programmed cell death event, whose regulation and physiological significance remain to be elucidated. Analyzing transcriptional responses of mouse embryonic fibroblasts exposed to the ferroptosis inducer erastin, here we found that a set of genes related to oxidative stress protection is induced upon ferroptosis. We considered that up-regulation of these genes attenuates ferroptosis induction and found that the transcription factor BTB domain and CNC homolog 1 (BACH1), a regulator in heme and iron metabolism, promotes ferroptosis by repressing the transcription of a subset of the erastin-induced protective genes. We noted that these genes are involved in the synthesis of GSH or metabolism of intracellular labile iron and include glutamate-cysteine ligase modifier subunit (Gclm), solute carrier family 7 member 11 (Slc7a11), ferritin heavy chain 1 (Fth1), ferritin light chain 1 (Ftl1), and solute carrier family 40 member 1 (Slc40a1). Ferroptosis has also been previously shown to induce cardiomyopathy, and here we observed that Bach1-/- mice are more resistant to myocardial infarction than WT mice and that the severity of ischemic injury is decreased by the iron-chelator deferasirox, which suppressed ferroptosis. Our findings suggest that BACH1 represses genes that combat labile iron-induced oxidative stress, and ferroptosis is stimulated at the transcriptional level by BACH1 upon disruption of the balance between the transcriptional induction of protective genes and accumulation of iron-mediated damage. We propose that BACH1 controls the threshold of ferroptosis induction and may represent a therapeutic target for alleviating ferroptosis-related diseases, including myocardial infarction.
Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Ferroptose , Glutationa/metabolismo , Ferro/metabolismo , Infarto do Miocárdio/metabolismo , Sistema y+ de Transporte de Aminoácidos/genética , Sistema y+ de Transporte de Aminoácidos/metabolismo , Animais , Fatores de Transcrição de Zíper de Leucina Básica/genética , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Células Cultivadas , Ferritinas/genética , Ferritinas/metabolismo , Fibroblastos/metabolismo , Glutamato-Cisteína Ligase/genética , Glutamato-Cisteína Ligase/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/patologia , Miócitos Cardíacos/metabolismo , Estresse Oxidativo , Oxirredutases/genética , Oxirredutases/metabolismo , Ativação TranscricionalRESUMO
OBJECTIVES: Patients with rheumatoid arthritis (RA) are at an increased risk of Mycobacterium avium complex pulmonary disease (MAC-PD). We aimed to identify factors associated with MAC-PD in RA patients, and investigate their clinical significance for diagnosis of this disease. METHODS: We examined 396 patients with RA for the presence of MAC-PD, using the criteria of the American Thoracic Society and conducted three years of follow-up on these patients. Multivariate logistic analyses were employed for selecting factors associated with MAC-PD. We developed a point system based on these factors which we call MAC-PD score to improve diagnosis of MAC-PD. RESULTS: During this study, 14 out of 396 patients were newly diagnosed with MAC-PD. Multivariate analyses revealed body mass index (BMI) <18.0 kg/m2 and lymphocyte count <1500/µl were associated with MAC-PD in RA patients. Points were assigned to them and totalled to provide the MAC-PD score. Among 20 patients with high-resolution computer tomography images consistent with MAC-PD, the scores were significantly higher in 14 patients with MAC-PD than those in six patients without MAC-PD. CONCLUSION: Using these data, in the forms of the MAC-PD score, could help to identify patients who should be considered for bronchoscopy more selectively.
Assuntos
Artrite Reumatoide , Linfopenia , Complexo Mycobacterium avium , Infecção por Mycobacterium avium-intracellulare , Tuberculose Pulmonar , Adulto , Idoso , Artrite Reumatoide/sangue , Artrite Reumatoide/complicações , Artrite Reumatoide/diagnóstico , Índice de Massa Corporal , Broncoscopia/métodos , Correlação de Dados , Feminino , Humanos , Japão , Linfopenia/diagnóstico , Linfopenia/etiologia , Masculino , Pessoa de Meia-Idade , Complexo Mycobacterium avium/isolamento & purificação , Complexo Mycobacterium avium/patogenicidade , Infecção por Mycobacterium avium-intracellulare/sangue , Infecção por Mycobacterium avium-intracellulare/complicações , Infecção por Mycobacterium avium-intracellulare/diagnóstico , Tomografia Computadorizada por Raios X/métodos , Tuberculose Pulmonar/sangue , Tuberculose Pulmonar/complicações , Tuberculose Pulmonar/diagnósticoRESUMO
Elucidation of how the differentiation of hematopoietic stem and progenitor cells (HSPCs) is reconfigured in response to the environment is critical for understanding the biology and disorder of hematopoiesis. Here we found that the transcription factors (TFs) Bach2 and Bach1 promoted erythropoiesis by regulating heme metabolism in committed erythroid cells to sustain erythroblast maturation and by reinforcing erythroid commitment at the erythro-myeloid bifurcation step. Bach TFs repressed expression of the gene encoding the transcription factor C/EBPß, as well as that of its target genes encoding molecules important for myelopoiesis and inflammation; they achieved the latter by binding to their regulatory regions also bound by C/EBPß. Lipopolysaccharide diminished the expression of Bach TFs in progenitor cells and promoted myeloid differentiation. Overexpression of Bach2 in HSPCs promoted erythroid development and inhibited myelopoiesis. Knockdown of BACH1 or BACH2 in human CD34+ HSPCs impaired erythroid differentiation in vitro. Thus, Bach TFs accelerate erythroid commitment by suppressing the myeloid program at steady state. Anemia of inflammation and myelodysplastic syndrome might involve reduced activity of Bach TFs.
Assuntos
Anemia/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Eritropoese/fisiologia , Anemia/etiologia , Animais , Diferenciação Celular/fisiologia , Células Eritroides/citologia , Células Eritroides/metabolismo , Humanos , Infecções/complicações , Lipopolissacarídeos/toxicidade , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Síndromes Mielodisplásicas/etiologia , Síndromes Mielodisplásicas/metabolismoRESUMO
S-adenosylmethionine (SAM) is an important metabolite as a methyl-group donor in DNA and histone methylation, tuning regulation of gene expression. Appropriate intracellular SAM levels must be maintained, because methyltransferase reaction rates can be limited by SAM availability. In response to SAM depletion, MAT2A, which encodes a ubiquitous mammalian methionine adenosyltransferase isozyme, was upregulated through mRNA stabilization. SAM-depletion reduced N6-methyladenosine (m6A) in the 3' UTR of MAT2A. In vitro reactions using recombinant METTL16 revealed multiple, conserved methylation targets in the 3' UTR. Knockdown of METTL16 and the m6A reader YTHDC1 abolished SAM-responsive regulation of MAT2A. Mutations of the target adenine sites of METTL16 within the 3' UTR revealed that these m6As were redundantly required for regulation. MAT2A mRNA methylation by METTL16 is read by YTHDC1, and we suggest that this allows cells to monitor and maintain intracellular SAM levels.
Assuntos
Metionina Adenosiltransferase/genética , Metiltransferases/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Fatores de Processamento de RNA/metabolismo , Estabilidade de RNA , RNA Mensageiro/metabolismo , S-Adenosilmetionina/metabolismo , Regiões 3' não Traduzidas , Animais , Células HEK293 , Células HeLa , Humanos , Metionina Adenosiltransferase/metabolismo , Metilação , Metiltransferases/genética , Camundongos , Proteínas do Tecido Nervoso/genética , Processamento Pós-Transcricional do RNA , Fatores de Processamento de RNA/genética , RNA Mensageiro/química , RNA Mensageiro/genéticaRESUMO
ATP-binding cassette subfamily B member 10 (Abcb10) is a mitochondrial ATP-binding cassette (ABC) transporter that complexes with mitoferrin1 and ferrochelatase to enhance heme biosynthesis in developing red blood cells. Reductions in Abcb10 levels have been shown to reduce mitoferrin1 protein levels and iron import into mitochondria, resulting in reduced heme biosynthesis. As an ABC transporter, Abcb10 binds and hydrolyzes ATP, but its transported substrate is unknown. Here, we determined that decreases in Abcb10 did not result in protoporphyrin IX accumulation in morphant-treated zebrafish embryos or in differentiated Abcb10-specific shRNA murine Friend erythroleukemia (MEL) cells in which Abcb10 was specifically silenced with shRNA. We also found that the ATPase activity of Abcb10 is necessary for hemoglobinization in MEL cells, suggesting that the substrate transported by Abcb10 is important in mediating increased heme biosynthesis during erythroid development. Inhibition of 5-aminolevulinic acid dehydratase (EC 4.2.1.24) with succinylacetone resulted in both 5-aminolevulinic acid (ALA) accumulation in control and Abcb10-specific shRNA MEL cells, demonstrating that reductions in Abcb10 do not affect ALA export from mitochondria and indicating that Abcb10 does not transport ALA. Abcb10 silencing resulted in an alteration in the heme biosynthesis transcriptional profile due to repression by the transcriptional regulator Bach1, which could be partially rescued by overexpression of Alas2 or Gata1, providing a mechanistic explanation for why Abcb10 shRNA MEL cells exhibit reduced hemoglobinization. In conclusion, our findings rule out that Abcb10 transports ALA and indicate that Abcb10's ATP-hydrolysis activity is critical for hemoglobinization and that the substrate transported by Abcb10 provides a signal that optimizes hemoglobinization.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Regulação Enzimológica da Expressão Gênica , Heme/biossíntese , Proteínas de Peixe-Zebra/metabolismo , Transportadores de Cassetes de Ligação de ATP/antagonistas & inibidores , Transportadores de Cassetes de Ligação de ATP/química , Transportadores de Cassetes de Ligação de ATP/genética , Motivos de Aminoácidos , Substituição de Aminoácidos , Animais , Fatores de Transcrição de Zíper de Leucina Básica/antagonistas & inibidores , Fatores de Transcrição de Zíper de Leucina Básica/genética , Embrião não Mamífero/enzimologia , Embrião não Mamífero/metabolismo , Proteínas de Grupos de Complementação da Anemia de Fanconi , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Camundongos , Microinjeções , Morfolinos/metabolismo , Mutação , Interferência de RNA , RNA Interferente Pequeno , Peixe-Zebra , Proteínas de Peixe-Zebra/antagonistas & inibidores , Proteínas de Peixe-Zebra/genéticaRESUMO
The capacity of the liver to regenerate is likely to be encoded as a plasticity of molecular networks within the liver. By applying a combination of comprehensive analyses of the epigenome, transcriptome, and proteome, we herein depict the molecular landscape of liver regeneration. We demonstrated that histone H3 Lys-4 was trimethylated at the promoter regions of many loci, among which only a fraction, including cell-cycle-related genes, were transcriptionally up-regulated. A cistrome analysis guided by the histone methylation patterns and the transcriptome identified FOXM1 as the key transcription factor promoting liver regeneration, which was confirmed in vitro using a hepatocarcinoma cell line. The promoter regions of cell-cycle-related genes and Foxm1 acquired higher levels of trimethylated histone H3 Lys-4, suggesting that epigenetic regulations of these key regulatory genes define quiescence and regeneration of the liver cells. A quantitative proteome analysis of the regenerating liver revealed that conditional protein degradation also mediated regeneration-specific protein expression. These sets of informational resources should be useful for further investigations of liver regeneration.
Assuntos
Regulação da Expressão Gênica , Histonas/química , Regeneração Hepática , Fígado/metabolismo , RNA Mensageiro/metabolismo , Motivos de Aminoácidos , Animais , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Epigênese Genética , Proteína Forkhead Box M1/metabolismo , Neoplasias Hepáticas/metabolismo , Lisina/química , Masculino , Metilação , Camundongos , Camundongos Endogâmicos C57BL , Regiões Promotoras Genéticas , Proteoma/metabolismo , TranscriptomaRESUMO
Hematopoietic stem cell and multipotent progenitor (MPP) commitment can be tuned in response to an infection so that their differentiation is biased toward myeloid cells. Here, we find that Bach2, which inhibits myeloid differentiation in common lymphoid progenitors, represses a cohort of myeloid genes and activates those linked to lymphoid function. Bach2 repressed both Cebpb and its target Csf1r, encoding C/EBPß and macrophage colony-stimulating factor receptor (M-CSFr), respectively, whereas C/EBPß repressed Bach2 and activated Csf1r. Bach2 and C/EBPß further bound to overlapping regulatory regions at their myeloid target genes, suggesting the presence of a gene regulatory network (GRN) with mutual repression between these factors and a feedforward loop leading to myeloid gene regulation. Lipopolysaccharide reduced the expression of Bach2, resulting in enhanced myeloid differentiation. The Bach2-C/EBPß GRN pathway thus tunes MPP commitment to myeloid and lymphoid lineages both under normal conditions and after infection.