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1.
Blood ; 139(14): 2089-2091, 2022 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-35389439
2.
Redox Biol ; 52: 102286, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35334247

RESUMO

BACKGROUND & AIMS: Hepatic stellate cells (HSCs) are the primary cell type in liver fibrosis, a significant global health care burden. Cytoglobin (CYGB), a globin family member expressed in HSCs, inhibits HSC activation and reduces collagen production. We studied the antifibrotic properties of globin family members hemoglobin (HB), myoglobin (MB), and neuroglobin (NGB) in comparison with CYGB. APPROACH & RESULTS: We characterized the biological activities of globins in cultured human HSCs (HHSteCs) and their effects on carbon tetrachloride (CCl4)-induced cirrhosis in mice. All globins demonstrated greater antioxidant capacity than glutathione in cell-free systems. Cellular fractionation revealed endocytosis of extracellular MB, NGB, and CYGB, but not HB; endocytosed globins localized to intracellular membranous, cytoplasmic, and cytoskeletal fractions. MB, NGB, and CYGB, but not HB, scavenged reactive oxygen species generated spontaneously or stimulated by H2O2 or transforming growth factor ß1 in HHSteCs and reduced collagen 1A1 production via suppressing COL1A1 promoter activity. Disulfide bond-mutant NGB displayed decreased heme and superoxide scavenging activity and reduced collagen inhibitory capacity. RNA sequencing of MB- and NGB-treated HHSteCs revealed downregulation of extracellular matrix-encoding and fibrosis-related genes and HSC deactivation markers. Upregulation of matrix metalloproteinase (MMP)-1 was observed following MB and NGB treatment, and MMP-1 knockdown partially reversed globin-mediated effects on secreted collagen. Importantly, administration of MB, NGB, and CYGB suppressed CCl4-induced mouse liver fibrosis. CONCLUSIONS: These findings revealed unexpected roles for MB and NGB in deactivating HSCs and inhibiting liver fibrosis development, suggesting that globin therapy may represent a new strategy for combating fibrotic liver disease.


Assuntos
Globinas , Metaloproteinase 1 da Matriz , Animais , Citoglobina , Globinas/genética , Globinas/metabolismo , Hemoglobinas , Peróxido de Hidrogênio , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/genética , Camundongos , Proteínas do Tecido Nervoso/metabolismo , Neuroglobina , Espécies Reativas de Oxigênio
3.
Blood Adv ; 6(3): 1088, 2022 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-35133406

RESUMO

Big data are used in the pursuit of precision medicine in the general population. Applying these tools to patients with sickle cell disease (SCD) is essential for ensuring that they receive the most appropriate customized therapy for their disease. For these tools to be applied, there must be a large number of willing, fully phenotyped participants in collaborative registries. Working collaboratively to respond to unmet clinical needs and the lack of a large multisite registry, SCD providers at multiple sites developed The Globin Regional Data and Discovery (GRNDaD) registry. The specific goals of GRNDaD are to (1) prospectively obtain high-quality curated data on the evolving population affected by SCD; (2) improve adherence to guideline-based care; (3) provide a platform for ongoing quality improvement across sites; (4) allow for real-time investigation of therapies, and collaborate broadly to address research questions using GRNDaD as a shared platform. GRNDaD's current strength lies in the generous participation of people living with SCD, collaborative investigators, and the opportunity to conduct quality improvement activities across a large number of sites. GRNDaD will serve as the data collection tool for the Health Resources and Services Administration Sickle Cell Treatment Demonstration Program and for the newly established National Alliance for Sickle Cell Centers (https://www.sicklecellcenters.org/). GRNDaD is a robust collaborative registry that providers and researchers will use to identify genetic markers that will help predict outcomes and lead to a better understanding of the natural history of SCD in the modern era of novel therapies.


Assuntos
Anemia Falciforme , Big Data , Anemia Falciforme/terapia , Eritrócitos Anormais , Globinas , Humanos , Medicina de Precisão , Estados Unidos
4.
Int J Biol Macromol ; 201: 216-225, 2022 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-34973267

RESUMO

The human ß-globin gene Locus Control Region (LCR), a dominant regulator of globin gene expression contains five tissue-specific DNase I-hypersensitive sites (HSs). A single nucleotide polymorphism (SNP) (A â†’ G) present in HS4 region of locus control region (LCR), have shown a notable association between the G allele and the occurrence of ß-thalassemia. This SNP site exhibiting a hairpin - duplex equilibrium manifested in A â†’ B like DNA transition has previously been reported from this laboratory. Since, DNA is a dynamic and adaptable molecule, so any change of a single base within a primary DNA sequence can produce major biological consequences commonly manifested in genetic disorders such as sickle cell anemia and ß-thalassemia. Herein, the differential behavior of sequential single base substitutions G â†’ A on the quasi-palindromic sequence (d-TGGGGGCCCCA; HPG11) has been explored. A combination of native gel electrophoresis, circular dichroism (CD), and UV-thermal denaturation (Tm) techniques have been used to investigate the structural polymorphism associated with various variants of HPG11 i.e. HPG11A2 to HPG11A5. The CD spectra confirmed that all the HPG11 variants exhibit a hairpin - duplex equilibrium. Oligomer concentration dependence on CD spectra has been correlated with A â†’ B DNA conformational transition. However, as revealed in gel electrophoresis, HPG11A2 â†’ A5 exhibit the formation of a tetramolecular structure (four-way junction) at higher oligomer concentration. UV-melting studies also supported the melting of hairpin, duplex and four-way junction structure. This polymorphism pattern may possibly be significant for DNA-protein recognition, in the process of regulation of LCR in the ß-globin gene.


Assuntos
Região de Controle de Locus Gênico , Globinas beta , Sequência de Bases , Globinas , Humanos , Família Multigênica , Polimorfismo de Nucleotídeo Único , Globinas beta/genética
5.
Proteins ; 90(5): 1152-1158, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-34982478

RESUMO

Human neuroglobin (Ngb) contains a heme group and three Cys residues (Cys46, Cys55, and Cys120) in the polypeptide chain. By introducing an additional Cys at position 15, the X-ray structure of A15C Ngb mutant was solved at a high resolution of 1.35 Å, which reveals the formation of both the native (C46C55) and the engineered (C15C120) disulfide bonds, likely playing a functional and structural role, respectively, according to the geometry analysis. Unexpectedly, 1,4-dioxane from the crystallization reagents was bound not only to the protein surface, but also to the heme distal pocket, providing insights into protein-ligand interactions for the globin and guiding the design of functional heme enzymes.


Assuntos
Globinas , Proteínas do Tecido Nervoso , Sítios de Ligação , Dissulfetos/química , Globinas/química , Globinas/genética , Globinas/metabolismo , Heme/química , Humanos , Ligantes , Proteínas do Tecido Nervoso/química , Neuroglobina , Raios X
6.
Blood ; 139(14): 2107-2118, 2022 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-35090172

RESUMO

The benign condition hereditary persistence of fetal hemoglobin (HPFH) is known to ameliorate symptoms of co-inherited ß-hemoglobinopathies, such as sickle cell disease and ß-thalassemia. The condition is sometimes associated with point mutations in the fetal globin promoters that disrupt the binding of the repressors BCL11A or ZBTB7A/LRF, which have been extensively studied. HPFH is also associated with a range of deletions within the ß-globin locus that all reside downstream of the fetal HBG2 gene. These deletional forms of HPFH are poorly understood and are the focus of this study. Numerous different mechanisms have been proposed to explain how downstream deletions can boost the expression of the fetal globin genes, including the deletion of silencer elements, of genes encoding noncoding RNA, and bringing downstream enhancer elements into proximity with the fetal globin gene promoters. Here we systematically analyze the deletions associated with both HPFH and a related condition known as δß-thalassemia and propose a unifying mechanism. In all cases where fetal globin is upregulated, the proximal adult ß-globin (HBB) promoter is deleted. We use clustered regularly interspaced short palindromic repeats-mediated gene editing to delete or disrupt elements within the promoter and find that virtually all mutations that reduce ΗΒΒ promoter activity result in elevated fetal globin expression. These results fit with previous models where the fetal and adult globin genes compete for the distal locus control region and suggest that targeting the ΗΒΒ promoter might be explored to elevate fetal globin and reduce sickle globin expression as a treatment of ß-hemoglobinopathies.


Assuntos
Globinas , Talassemia beta , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Hemoglobina Fetal/genética , Hemoglobina Fetal/metabolismo , Expressão Gênica , Globinas/metabolismo , Humanos , Fatores de Transcrição/genética , Globinas beta/genética , Globinas beta/metabolismo , Talassemia beta/genética , Talassemia beta/terapia
7.
Chemistry ; 28(23): e202104342, 2022 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-35080290

RESUMO

Sulfite is a potent toxic substance causing harm to multi-organ in human. Despite toxicity, it is widely used as preservative, anti-browning and anti-oxidant in foods, beverages, and pharmaceuticals, which cause easy admission of sulfite in human. Sulfite is also produced endogenously during the catabolism of cysteine and methionine. In vivo, the serum sulfite level at physiological range is strictly maintained by a molybdenum dependent sulfite oxidase (SO), which catalyzes sulfite to sulfate oxidation via a two-electron oxidation pathway. The loss of SO activity causes high serum sulfite level that fosters several diseases, including asthma, neurological dysfunction, birth defects, and heart diseases. The cytotoxicity of (bi)sulfite is implicated as sulfite radicals, which are generated by mainly heme-peroxidases via a one-electron oxidation pathway. On the other hand, the toxic sulfite radicals are neutralized to sulfite by heme-globins. The enzymatic reduction of sulfite to sulfide is catalyzed by sulfite reductase, which contains an unusual metal cofactor, siroheme-[4Fe4S]-cluster. Overall, the interaction of sulfite with various metalloproteins in vivo is a close relation with human health. Therefore, this review describes the metabolic conversion of (bi)sulfite to sulfate, sulfite radical or sulfide via oxidation or reduction pathways by various metalloproteins (specially SOs, peroxidases, heme-globins, and sulfite reductases), and the potential applications of sulfite in biosensors/biofuel cells, anti-browning, and advance oxidation process.


Assuntos
Metaloproteínas , Globinas , Heme , Humanos , Hidrogênio , Metaloproteínas/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/metabolismo , Peroxidases , Sulfatos , Sulfetos , Sulfitos/metabolismo
8.
Nat Med ; 28(1): 63-70, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34980909

RESUMO

ß-Thalassemias are inherited anemias that are caused by the absent or insufficient production of the ß chain of hemoglobin. Here we report 6-8-year follow-up of four adult patients with transfusion-dependent ß-thalassemia who were infused with autologous CD34+ cells transduced with the TNS9.3.55 lentiviral globin vector after reduced-intensity conditioning (RIC) in a phase 1 clinical trial ( NCT01639690) . Patients were monitored for insertional mutagenesis and the generation of a replication-competent lentivirus (safety and tolerability of the infusion product after RIC-primary endpoint) and engraftment of genetically modified autologous CD34+ cells, expression of the transduced ß-globin gene and post-transplant transfusion requirements (efficacy-secondary endpoint). No unexpected safety issues occurred during conditioning and cell product infusion. Hematopoietic gene marking was very stable but low, reducing transfusion requirements in two patients, albeit not achieving transfusion independence. Our findings suggest that non-myeloablative conditioning can achieve durable stem cell engraftment but underscore a minimum CD34+ cell transduction requirement for effective therapy. Moderate clonal expansions were associated with integrations near cancer-related genes, suggestive of non-erythroid activity of globin vectors in stem/progenitor cells. These correlative findings highlight the necessity of cautiously monitoring patients harboring globin vectors.


Assuntos
Terapia Genética/métodos , Vetores Genéticos , Globinas/genética , Lentivirus/genética , Condicionamento Pré-Transplante/métodos , Talassemia beta/terapia , Adolescente , Adulto , Antígenos CD34/genética , Transfusão de Sangue , Feminino , Humanos , Masculino , Transdução Genética , Adulto Jovem
9.
Plant Cell Environ ; 45(1): 178-190, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34633089

RESUMO

An important and interesting feature of rice is that it can germinate under anoxic conditions. Though several biochemical adaptive mechanisms play an important role in the anaerobic germination of rice but the role of phytoglobin-nitric oxide cycle and alternative oxidase pathway is not known, therefore in this study we investigated the role of these pathways in anaerobic germination. Under anoxic conditions, deepwater rice germinated much higher and rapidly than aerobic condition and the anaerobic germination and growth were much higher in the presence of nitrite. The addition of nitrite stimulated NR activity and NO production. Important components of phytoglobin-NO cycle such as methaemoglobin reductase activity, expression of Phytoglobin1, NIA1 were elevated under anaerobic conditions in the presence of nitrite. The operation of phytoglobin-NO cycle also enhanced anaerobic ATP generation, LDH, ADH activities and in parallel ethylene levels were also enhanced. Interestingly nitrite suppressed the ROS production and lipid peroxidation. The reduction of ROS was accompanied by enhanced expression of mitochondrial alternative oxidase protein and its capacity. Application of AOX inhibitor SHAM inhibited the anoxic growth mediated by nitrite. In addition, nitrite improved the submergence tolerance of seedlings. Our study revealed that nitrite driven phytoglobin-NO cycle and AOX are crucial players in anaerobic germination and growth of deepwater rice.


Assuntos
Germinação/fisiologia , Óxido Nítrico/metabolismo , Oryza/crescimento & desenvolvimento , Oryza/metabolismo , Proteínas de Plantas/metabolismo , Anaerobiose , Etilenos/metabolismo , Fermentação , Globinas/metabolismo , Proteínas Mitocondriais/metabolismo , Nitrato Redutase/metabolismo , Nitritos/metabolismo , Oryza/genética , Oxirredutases/metabolismo , Proteínas de Plantas/genética , Espécies Reativas de Oxigênio/metabolismo , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Trealose/metabolismo
10.
Mol Biol Rep ; 49(1): 249-258, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34743275

RESUMO

BACKGROUND: CDKN1A gene encoding p21 is an important tumour supressor involved in the pathogenesis of cancers. A few studies have been devoted to the association between CDKN1A single nucleotide polymorphisms (SNPs) and esophageal cancer (EC) in China, India and Iran. The aim of this case-control study was to investigate the association of CDKN1A polymorphisms with EC risk in the Turkey population for the first time. METHODS: In the present study, CDKN1A SNPs (rs1801270 C > T, rs1059234 C > A and rs3176352 C > G) were genotyped with the use of TaqMan SNP genotyping assays in 102 patients and 119 controls. RESULTS: The genotypes and alleles of CDKN1A SNPs were not significantly different among patients and controls. However, TT-genotype and T-allele of the rs1059234, the rs1801270 CC-genotype and rs3176352 G-allele were significantly associated with EC risk for ≤ 55 age (p < 0.05). In those over 55 age, CC-genotype and C-allele of the rs1059234 was significantly associated with EC (p < 0.05). The rs1059234 T-carriers had a higher risk of high globulin level (p = 0.017) and low albumin/globulin ratio (p = 0.019) when compared to non-T carriers (CC). The rs3176352 CC-genotype carriers had a higher risk of esophageal adenocarcinoma (EAC) subtype when compared to CG-genotype carriers and CG-genotype carriers had a higher risk of squamous cell carcinoma (ESCC) subtype (OR/95% CI = 4.00/1.06-15.08, p = 0.04). The rs3176352 CC-genotype is also a risk factor for the higher BMI (p = 0.04) and the higher CA-19-9 level (p = 0.009). CONCLUSION: Our study suggests that the CDKN1A polymorphisms may play an important role in EC risk in relation to age. Future studies are needed to validate our findings.


Assuntos
Adenocarcinoma/genética , Carcinoma de Células Escamosas/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Neoplasias Esofágicas/genética , Polimorfismo de Nucleotídeo Único , Adenocarcinoma/metabolismo , Adulto , Fatores Etários , Idoso , Antígenos Glicosídicos Associados a Tumores/metabolismo , Carcinoma de Células Escamosas/metabolismo , Estudos de Casos e Controles , China , Neoplasias Esofágicas/metabolismo , Feminino , Predisposição Genética para Doença , Globinas/metabolismo , Humanos , Índia , Irã (Geográfico) , Masculino , Pessoa de Meia-Idade
11.
Physiol Rev ; 102(2): 859-892, 2022 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-34486392

RESUMO

Globin proteins exist in every cell type of the vasculature, from erythrocytes to endothelial cells, vascular smooth muscle cells, and peripheral nerve cells. Many globin subtypes are also expressed in muscle tissues (including cardiac and skeletal muscle), in other organ-specific cell types, and in cells of the central nervous system (CNS). The ability of each of these globins to interact with molecular oxygen (O2) and nitric oxide (NO) is preserved across these contexts. Endothelial α-globin is an example of extraerythrocytic globin expression. Other globins, including myoglobin, cytoglobin, and neuroglobin, are observed in other vascular tissues. Myoglobin is observed primarily in skeletal muscle and smooth muscle cells surrounding the aorta or other large arteries. Cytoglobin is found in vascular smooth muscle but can also be expressed in nonvascular cell types, especially in oxidative stress conditions after ischemic insult. Neuroglobin was first observed in neuronal cells, and its expression appears to be restricted mainly to the CNS and the peripheral nervous system. Brain and CNS neurons expressing neuroglobin are positioned close to many arteries within the brain parenchyma and can control smooth muscle contraction and thus tissue perfusion and vascular reactivity. Overall, reactions between NO and globin heme iron contribute to vascular homeostasis by regulating vasodilatory NO signals and scavenging reactive species in cells of the mammalian vascular system. Here, we discuss how globin proteins affect vascular physiology, with a focus on NO biology, and offer perspectives for future study of these functions.


Assuntos
Fenômenos Fisiológicos Cardiovasculares , Citoglobina/metabolismo , Células Endoteliais/metabolismo , Globinas/metabolismo , Animais , Humanos , Mioglobina/metabolismo , Neuroglobina/metabolismo
12.
Anat Rec (Hoboken) ; 305(2): 254-264, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34358403

RESUMO

Bilirubin encephalopathy (BE) is a neurological syndrome in newborns, mainly caused by neuronal injury due to excessive oxidative stress produced by unconjugated bilirubin (UCB). Neuroglobin (NGB) can protect the brain by removing oxidative stress species, but its expression and significance in BE are not clear. To address this question, the neonatal BE model was established by injecting UCB into the cerebellomedullary cistern of 7-day-old SD rats. Rats were divided into a sham and BE 6 hr group, BE 12 hr group, BE 24 hr group, and BE 7 d group according to UCB action times. Hematoxylin/eosin and Nissl staining, and electron microscopy were employed to observe the pathological and ultrastructural changes of nerve cells in each group. Immunofluorescence staining was used to detect NGB expression sites and cell types. Western blotting and quantitative PCR served to detect NGB expression and test the mitochondrial apoptosis signal pathway. The results confirm that UCB can lead to pathological damage and ultrastructural changes in rats' temporal cortex, increasing the expression of apoptosis-related proteins Bax, Bcl-2, Cyt c, Caspase-3, and neuronal NGB. UCB promotes NGB expression with an increase in action time and reach a peak at 12 hr. In summary, brain damage induced by UCB will cause an increase in NGB expression, the increasing NGB can inhibit neuron apoptosis in early BE phases. Therefore, promoting the expression of endogenous NGB, to act as a neuroprotective agent may be a potential treatment strategy for BE.


Assuntos
Globinas , Kernicterus , Animais , Globinas/genética , Globinas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neuroglobina , Ratos , Ratos Sprague-Dawley , Lobo Temporal/metabolismo
13.
Mol Aspects Med ; 84: 101055, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34876274

RESUMO

Neuroglobin is expressed in vertebrate brain and belongs to a branch of the globin family that diverged early in evolution. Sequence conservation and presence in nervous cells of several taxa suggests a relevant role in the nervous system, with tight structural restraints. Twenty years after its discovery, a rich scientific literature provides convincing evidence of the involvement of neuroglobin in sustaining neuron viability in physiological and pathological conditions however, a full and conclusive picture of its specific function, or set of functions is still lacking. The difficulty of unambiguously assigning a precise mechanism and biochemical role to neuroglobin might arise from the participation to one or more cell mechanism that redundantly guarantee the functioning of the highly specialized and metabolically demanding central nervous system of vertebrates. Here we collect findings and hypotheses arising from recent biochemical, biophysical, structural, in cell and in vivo experimental work on neuroglobin, aiming at providing an overview of the most recent literature. Proteins are said to have jobs and hobbies, it is possible that, in the case of neuroglobin, evolution has selected for it more than one job, and support to cover for its occasional failings. Disentangling the mechanisms and roles of neuroglobin is thus a challenging task that might be achieved by considering data from different disciplines and experimental approaches.


Assuntos
Globinas , Proteínas do Tecido Nervoso , Animais , Encéfalo/metabolismo , Globinas/química , Globinas/genética , Humanos , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Neuroglobina/metabolismo , Neurônios/metabolismo
14.
STAR Protoc ; 2(4): 100995, 2021 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-34950881

RESUMO

We have developed a protocol for barcoded cDNA libraries of 48 samples to study gene expression across tissues in the domestic dog, Canis familiaris, by modifying the Single-Cell Tagged Reverse Transcription (STRT) protocol (Islam et al., 2012, 2014). The cDNA reads represent mRNA 5' ends, enabling the study of transcription start sites (TSS). Our modifications include longer UMIs for molecular counting and Globin-Lock® to deplete globin mRNAs that are abundant in blood and blood-rich tissues dominating all reads.


Assuntos
Biblioteca Gênica , Globinas/genética , RNA-Seq/métodos , Transcriptoma/genética , Animais , Cães , Globinas/análise , Globinas/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
15.
PLoS Negl Trop Dis ; 15(9): e0009811, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34591853

RESUMO

Despite recent evidence suggesting that adult trematodes require oxygen for the generation of bioenergy and eggshells, information on the molecular mechanism by which the parasites acquire oxygen remains largely elusive. In this study, the structural and expressional features of globin genes identified in Clonorchis sinensis, a carcinogenic trematode parasite that invades the hypoxic biliary tracts of mammalian hosts, were investigated to gain insight into the molecules that enable oxygen metabolism. The number of globin paralogs substantially differed among parasitic platyhelminths, ranging from one to five genes, and the C. sinensis genome encoded at least five globin genes. The expression of these Clonorchis genes, named CsMb (CsMb1-CsMb3), CsNgb, and CsGbX, according to their preferential similarity patterns toward respective globin subfamilies, exponentially increased in the worms coinciding with their sexual maturation, after being downregulated in early juveniles compared to those in metacercariae. The CsMb1 protein was detected throughout the parenchymal region of adult worms as well as in excretory-secretory products, whereas the other proteins were localized exclusively in the sexual organs and intrauterine eggs. Stimuli generated by exogenous oxygen, nitric oxide (NO), and nitrite as well as co-incubation with human cholangiocytes variously affected globin gene expression in live C. sinensis adults. Together with the specific histological distributions, these hypoxia-induced patterns may suggest that oxygen molecules transported by CsMb1 from host environments are provided to cells in the parenchyma and intrauterine eggs/sex organs of the worms for energy metabolism and/or, more importantly, eggshell formation by CsMb1 and CsMb3, respectively. Other globin homologs are likely to perform non-respiratory functions. Based on the responsive expression profile against nitrosative stress, an oxygenated form of secreted CsMb1 is suggested to play a pivotal role in parasite survival by scavenging NO generated by host immune cells via its NO dioxygenase activity.


Assuntos
Carcinógenos/metabolismo , Clonorchis sinensis/genética , Clonorchis sinensis/metabolismo , Hipóxia , Mioglobina/genética , Mioglobina/metabolismo , Estresse Nitrosativo , Animais , Bile , Expressão Gênica/efeitos dos fármacos , Globinas/genética , Humanos , Metacercárias , Nitratos/farmacologia , Óxido Nítrico/metabolismo , Nitritos/farmacologia , Oxirredução , Oxigênio/farmacologia , Filogenia , Trematódeos/genética , Trematódeos/metabolismo , Peixe-Zebra
16.
Cell Rep ; 36(13): 109759, 2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34592153

RESUMO

Elevated levels of fetal globin protect against ß-hemoglobinopathies, such as sickle cell disease and ß-thalassemia. Two zinc-finger (ZF) repressors, BCL11A and ZBTB7A/LRF, bind directly to the fetal globin promoter elements positioned at -115 and -200, respectively. Here, we describe X-ray structures of the ZBTB7A DNA-binding domain, consisting of four adjacent ZFs, in complex with the -200 sequence element, which contains two copies of four consecutive C:G base pairs. ZF1 and ZF2 recognize the 5' C:G quadruple, and ZF4 contacts the 3' C:G quadruple. Natural non-coding DNA mutations associated with hereditary persistence of fetal hemoglobin (HPFH) impair ZBTB7A DNA binding, with the most severe disruptions resulting from mutations in the base pairs recognized by ZF1 and ZF2. Our results firmly establish ZBTB7A/LRF as a key molecular regulator of fetal globin expression and inform genome-editing strategies that inhibit repressor binding and boost fetal globin expression to treat hemoglobinopathies.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Globinas/genética , Globinas/metabolismo , Regiões Promotoras Genéticas/genética , Fatores de Transcrição/metabolismo , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Hemoglobina Fetal/genética , Edição de Genes/métodos , Humanos , Fatores de Transcrição/genética , Dedos de Zinco/fisiologia , Talassemia beta/genética
18.
Nat Commun ; 12(1): 4922, 2021 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-34389729

RESUMO

CRISPR-Cas9 is a promising technology for gene therapy. However, the ON-target genotoxicity of CRISPR-Cas9 nuclease due to DNA double-strand breaks has received little attention and is probably underestimated. Here we report that genome editing targeting globin genes induces megabase-scale losses of heterozygosity (LOH) from the globin CRISPR-Cas9 cut-site to the telomere (5.2 Mb). In established lines, CRISPR-Cas9 nuclease induces frequent terminal chromosome 11p truncations and rare copy-neutral LOH. In primary hematopoietic progenitor/stem cells, we detect 1.1% of clones (7/648) with acquired megabase LOH induced by CRISPR-Cas9. In-depth analysis by SNP-array reveals the presence of copy-neutral LOH. This leads to 11p15.5 partial uniparental disomy, comprising two Chr11p15.5 imprinting centers (H19/IGF2:IG-DMR/IC1 and KCNQ1OT1:TSS-DMR/IC2) and impacting H19 and IGF2 expression. While this genotoxicity is a safety concern for CRISPR clinical trials, it is also an opportunity to model copy-neutral-LOH for genetic diseases and cancers.


Assuntos
Sistemas CRISPR-Cas , Edição de Genes/métodos , Globinas/genética , Células-Tronco Hematopoéticas/metabolismo , Perda de Heterozigosidade/genética , Deleção de Sequência , Células Cultivadas , Deleção Cromossômica , Cromossomos Humanos Par 11/genética , Metilação de DNA , Expressão Gênica , Células HEK293 , Células-Tronco Hematopoéticas/citologia , Humanos , Fator de Crescimento Insulin-Like II/genética , Polimorfismo de Nucleotídeo Único , RNA Longo não Codificante/genética
19.
Nat Commun ; 12(1): 4991, 2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34404810

RESUMO

Key mechanisms of fetal hemoglobin (HbF) regulation and switching have been elucidated through studies of human genetic variation, including mutations in the HBG1/2 promoters, deletions in the ß-globin locus, and variation impacting BCL11A. While this has led to substantial insights, there has not been a unified understanding of how these distinct genetically-nominated elements, as well as other key transcription factors such as ZBTB7A, collectively interact to regulate HbF. A key limitation has been the inability to model specific genetic changes in primary isogenic human hematopoietic cells to uncover how each of these act individually and in aggregate. Here, we describe a single-cell genome editing functional assay that enables specific mutations to be recapitulated individually and in combination, providing insights into how multiple mutation-harboring functional elements collectively contribute to HbF expression. In conjunction with quantitative modeling and chromatin capture analyses, we illustrate how these genetic findings enable a comprehensive understanding of how distinct regulatory mechanisms can synergistically modulate HbF expression.


Assuntos
Edição de Genes , Hemoglobinas/genética , Hemoglobinas/metabolismo , Sistemas CRISPR-Cas , Cromatina , Cromossomos , Proteínas de Ligação a DNA/metabolismo , Hemoglobina Fetal/genética , Hemoglobina Fetal/metabolismo , Expressão Gênica , Globinas , Humanos , Mutação , Proteínas Repressoras , Fatores de Transcrição/metabolismo , Globinas beta/genética
20.
Genes (Basel) ; 12(6)2021 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-34208174

RESUMO

There are many co-regulated genes in eukaryotic cells. The coordinated activation or repression of such genes occurs at specific stages of differentiation, or under the influence of external stimuli. As a rule, co-regulated genes are dispersed in the genome. However, there are also gene clusters, which contain paralogous genes that encode proteins with similar functions. In this aspect, they differ significantly from bacterial operons containing functionally linked genes that are not paralogs. In this review, we discuss the reasons for the existence of gene clusters in vertebrate cells and propose that clustering is necessary to ensure the possibility of selective activation of one of several similar genes.


Assuntos
Evolução Molecular , Família Multigênica , Animais , Caderinas/genética , Caderinas/metabolismo , Células Eritroides/metabolismo , Globinas/genética , Globinas/metabolismo , Humanos
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