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1.
J Biochem ; 171(6): 673-693, 2022 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-35325168

RESUMO

Gram-negative intracellular pathogen Vibrio parahaemolyticus manifests its infection through a series of effector proteins released into the host via the type III secretion system. Most of these effector proteins alter signalling pathways of the host to facilitate survival and proliferation of bacteria inside host cells. Here, we report V. parahaemolyticus (serotype O3:K6) infection-induced histone deacetylation in host intestinal epithelial cells, particularly deacetylation of H3K9, H3K56, H3K18 and H4K16 residues. We found a putative NAD+-dependent deacetylase, vp1524 (vpCobB) of V. parahaemolyticus, was overexpressed during infection. Biochemical assays revealed that Vp1524 is a functional NAD+-dependent Sir2 family deacetylase in vitro, which was capable of deacetylating acetylated histones. Furthermore, we observed that vp1524 is expressed and localized to the nuclear periphery of the host cells during infection. Consequently, Vp1524 translocated to nuclear compartments of transfected cells, deacetylated histones, specifically causing deacetylation of those residues (K56, K16, K18) associated with V. parahaemolyticus infection. This infection induced deacetylation resulted in transcriptional repression of several host genes involved in epigenetic regulation, immune response, autophagy etc. Thus, our study shows that a V. parahaemolyticus lysine deacetylase Vp1524 is secreted inside the host cells during infection, modulating host gene expression through histone deacetylation.


Assuntos
Histona Desacetilases do Grupo III/metabolismo , Vibrio parahaemolyticus , Epigênese Genética , Histonas/metabolismo , Imunidade , NAD/genética , NAD/metabolismo , Vibrio parahaemolyticus/genética , Vibrio parahaemolyticus/metabolismo
2.
Eur J Pharmacol ; 914: 174666, 2022 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-34861210

RESUMO

Diabetes mellitus (DM) is associated with accelerated cognitive decline. However, the mechanism of diabetic cognitive impairment remains poorly understood. In this study, we found that the expression of Sirtuin 1 (SIRT1), a nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase, was downregulated significantly in the hippocampus of streptozotocin (STZ)-induced diabetic cognitive impairment rats. Viral overexpression of hippocampal SIRT1 ameliorated cognitive impairment in diabetic rats, but viral knockdown of hippocampal SIRT1 mimicked the diabetic effect, eliciting the cognitive decline in normal animals. Further study showed that the decreased level of SIRT1 may result in the increase of acetylated tau protein in the hippocampus, which may mediate the development of diabetic cognitive impairment. These results suggest that SIRT1 may be a key epigenetic regulator that guards against the development of diabetic cognitive impairment by deacetylating the tau protein. SIRT1 activator may serve as a new therapeutic approach for the treatment of diabetic cognitive impairment.


Assuntos
Disfunção Cognitiva , Complicações do Diabetes/metabolismo , Sirtuína 1/metabolismo , Proteínas tau/metabolismo , Acetilação , Animais , Disfunção Cognitiva/etiologia , Disfunção Cognitiva/metabolismo , Diabetes Mellitus Experimental , Regulação para Baixo , Epigênese Genética , Histona Desacetilases do Grupo III/metabolismo , Hipocampo/metabolismo , Processamento de Proteína Pós-Traducional , Ratos
3.
Int J Mol Sci ; 21(10)2020 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-32455951

RESUMO

Chagas disease is an illness caused by the protozoan parasite Trypanosoma cruzi, affecting more than 7 million people in the world. Benznidazole and nifurtimox are the only drugs available for treatment and in addition to causing several side effects, are only satisfactory in the acute phase of the disease. Sirtuins are NAD+-dependent deacetylases involved in several biological processes, which have become drug target candidates in various disease settings. T. cruzi presents two sirtuins, one cytosolic (TcSir2rp1) and the latter mitochondrial (TcSir2rp3). Here, we characterized the effects of human sirtuin inhibitors against T. cruzi sirtuins as an initial approach to develop specific parasite inhibitors. We found that, of 33 compounds tested, two inhibited TcSir2rp1 (15 and 17), while other five inhibited TcSir2rp3 (8, 12, 13, 30, and 32), indicating that specific inhibitors can be devised for each one of the enzymes. Furthermore, all inhibiting compounds prevented parasite proliferation in cultured mammalian cells. When combining the most effective inhibitors with benznidazole at least two compounds, 17 and 32, demonstrated synergistic effects. Altogether, these results support the importance of exploring T. cruzi sirtuins as drug targets and provide key elements to develop specific inhibitors for these enzymes as potential targets for Chagas disease treatment.


Assuntos
Doença de Chagas/tratamento farmacológico , Nitroimidazóis/farmacologia , Sirtuínas/antagonistas & inibidores , Sirtuínas/metabolismo , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Animais , Linhagem Celular , Sinergismo Farmacológico , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/parasitologia , Histona Desacetilases do Grupo III/antagonistas & inibidores , Concentração Inibidora 50 , Macaca mulatta , Simulação de Acoplamento Molecular , Filogenia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Sirtuínas/química , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidade
4.
Gene ; 704: 134-141, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-30981839

RESUMO

To maintain normal function of cartilage tissue normally, the presence of a sufficient amount of type II collagen and aggrecan is essential, and their synthesis is tightly regulated. Therefore, understanding the mechanisms that control the expression of type II collagen and aggrecan would be useful for understanding gene expression changes in diseases such as osteoarthritis. Recently, we have identified two pairs of enhancer elements, termed E1 and E2 in the type II collagen gene and Ea and Eb in the aggrecan gene. However, their different mechanisms of action remained unclear. Thus, the central aim of this study was to clarify the different transcriptional regulation mediated through each enhancer element. To this end, we established different stable reporter cell lines that express a reporter gene under the control of different enhancer elements using a silent reporter system we previously constructed. Using these cell lines, we found that dexamethasone, forskolin, and trichostatin A affect the gene expression of type II collagen and aggrecan via different enhancer elements. Moreover, we clarified that E1 and E2 enhancer activities are regulated through distinct epigenetic modifications by histone deacetylase 10 and sirtuin 6.


Assuntos
Agrecanas/genética , Colágeno Tipo II/genética , Elementos Facilitadores Genéticos/genética , Epigênese Genética/fisiologia , Agrecanas/metabolismo , Animais , Condrócitos/metabolismo , Condrócitos/patologia , Colágeno Tipo II/metabolismo , Regulação da Expressão Gênica , Histona Desacetilases do Grupo III/metabolismo , Regiões Promotoras Genéticas , Ratos , Sirtuínas/metabolismo , Células Tumorais Cultivadas
5.
Curr Opin Clin Nutr Metab Care ; 21(4): 252-259, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29847446

RESUMO

PURPOSE OF REVIEW: We present a current perspective of epigenetic alterations that can lead to cardiovascular disease (CVD) and the potential of dietary factors to counteract their actions. In addition, we discuss the challenges and opportunities of dietary treatments as epigenetic modifiers for disease prevention and therapy. RECENT FINDINGS: Recent epigenome-wide association studies along with candidate gene approaches and functional studies in cell culture and animal models have delineated mechanisms through which nutrients, food compounds and dietary patterns may affect the epigenome. Several risk factors for CVD, including adiposity, inflammation and oxidative stress, have been associated with changes in histone acetylation, lower global DNA methylation levels and shorter telomere length. A surplus of macronutrients such as in a high-fat diet or deficiencies of specific nutrients such as folate and other B-vitamins can affect the activity of DNA methyltransferases and histone-modifying enzymes, affecting foetal growth, glucose/lipid metabolism, oxidative stress, inflammation and atherosclerosis. Bioactive compounds such as polyphenols (resveratrol, curcumin) or epigallocatechin may activate deacetylases Sirtuins (SIRTs), histone deacetylases or acetyltransferases and in turn the response of inflammatory mediators. Adherence to cardioprotective dietary patterns, such as the Mediterranean diet (MedDiet), has been associated with altered methylation and expression of genes related to inflammation and immuno-competence. SUMMARY: The mechanisms through which nutrients and dietary patterns may alter the cardiovascular epigenome remain elusive. The research challenge is to determine which of these nutriepigenetic effects are reversible, so that novel findings translate into effective dietary interventions to prevent CVD or its progression.


Assuntos
Doenças Cardiovasculares , Metilação de DNA , Dieta , Epigênese Genética , Histonas/metabolismo , Estado Nutricional , Acetilação , Animais , Doenças Cardiovasculares/dietoterapia , Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/genética , Doenças Cardiovasculares/metabolismo , Histona Desacetilases do Grupo III/metabolismo , Humanos , Inflamação/genética , Polifenóis , Processamento de Proteína Pós-Traducional
6.
PLoS One ; 13(3): e0193602, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29543820

RESUMO

The de novo crystal structure of the Leishmania infantum Silent Information Regulator 2 related protein 1 (LiSir2rp1) has been solved at 1.99Å in complex with an acetyl-lysine peptide substrate. The structure is broadly commensurate with Hst2/SIRT2 proteins of yeast and human origin, reproducing many of the structural features common to these sirtuin deacetylases, including the characteristic small zinc-binding domain, and the larger Rossmann-fold domain involved in NAD+-binding interactions. The two domains are linked via a cofactor binding loop ordered in open conformation. The peptide substrate binds to the LiSir2rp1 protein via a cleft formed between the small and large domains, with the acetyl-lysine side chain inserting further into the resultant hydrophobic tunnel. Crystals were obtained only with recombinant LiSir2rp1 possessing an extensive internal deletion of a proteolytically-sensitive region unique to the sirtuins of kinetoplastid origin. Deletion of 51 internal amino acids (P253-E303) from LiSir2rp1 did not appear to alter peptide substrate interactions in deacetylation assays, but was indispensable to obtain crystals. Removal of this potentially flexible region, that otherwise extends from the classical structural elements of the Rossmann-fold, specifically the ß8-ß9 connector, appears to result in lower accumulation of the protein when expressed from episomal vectors in L. infantum SIR2rp1 single knockout promastigotes. The biological function of the large serine-rich insertion in kinetoplastid/trypanosomatid sirtuins, highlighted as a disordered region with strong potential for post-translational modification, remains unknown but may confer additional cellular functions that are distinct from their human counterparts. These unique molecular features, along with the resolution of the first kinetoplastid sirtuin deacetylase structure, present novel opportunities for drug design against a protein target previously established as essential to parasite survival and proliferation.


Assuntos
Histona Desacetilases do Grupo III/química , Histona Desacetilases do Grupo III/metabolismo , Leishmania infantum/metabolismo , Sítios de Ligação , Cristalografia por Raios X , Desenho de Fármacos , Humanos , Modelos Moleculares , Peptídeos/metabolismo , Ligação Proteica , Estrutura Secundária de Proteína , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo
7.
J Biol Chem ; 293(14): 5281-5294, 2018 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-29440391

RESUMO

Heart failure is an aging-associated disease that is the leading cause of death worldwide. Sirtuin family members have been largely studied in the context of aging and aging-associated diseases. Sirtuin 2 (SIRT2) is a cytoplasmic protein in the family of sirtuins that are NAD+-dependent class III histone deacetylases. In this work, we studied the role of SIRT2 in regulating nuclear factor of activated T-cells (NFAT) transcription factor and the development of cardiac hypertrophy. Confocal microscopy analysis indicated that SIRT2 is localized in the cytoplasm of cardiomyocytes and SIRT2 levels are reduced during pathological hypertrophy of the heart. SIRT2-deficient mice develop spontaneous pathological cardiac hypertrophy, remodeling, fibrosis, and dysfunction in an age-dependent manner. Moreover, young SIRT2-deficient mice develop exacerbated agonist-induced hypertrophy. In contrast, SIRT2 overexpression attenuated agonist-induced cardiac hypertrophy in cardiomyocytes in a cell-autonomous manner. Mechanistically, SIRT2 binds to and deacetylates NFATc2 transcription factor. SIRT2 deficiency stabilizes NFATc2 and enhances nuclear localization of NFATc2, resulting in increased transcription activity. Our results suggest that inhibition of NFAT rescues the cardiac dysfunction in SIRT2-deficient mice. Thus, our study establishes SIRT2 as a novel endogenous negative regulator of NFAT transcription factor.


Assuntos
Cardiomegalia/metabolismo , Fatores de Transcrição NFATC/metabolismo , Sirtuína 2/metabolismo , Acetilação , Animais , Regulação da Expressão Gênica/genética , Histona Desacetilases do Grupo III/metabolismo , Insuficiência Cardíaca/metabolismo , Homeostase , Camundongos , Camundongos Knockout , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/fisiologia , Sirtuína 2/fisiologia
8.
J Neuromuscul Dis ; 5(1): 59-73, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29278895

RESUMO

BACKGROUND: Mutations in the LAMA2 gene encoding laminin-α2 cause congenital muscular dystrophy Type 1A (MDC1A), a severe recessive disease with no effective treatment. Previous studies have shown that aberrant activation of caspases and cell death through a pathway regulated by BAX and KU70 is a significant contributor to pathogenesis in laminin-α2-deficiency. OBJECTIVES: To identify mechanisms of pathogenesis in MDC1A. METHODS: We used immunocytochemical and molecular studies of human myogenic cells and mouse muscles-comparing laminin-α2-deficient vs. healthy controls-to identify mechanisms that regulate pathological activation of caspase in laminin-α2-deficiency. RESULTS: In cultures of myogenic cells from MDC1A donors, p53 accumulated in a subset of nuclei and aberrant caspase activation was inhibited by the p53 inhibitor pifithrin-alpha. Also, the p53 target BBC3 (PUMA) was upregulated in both MDC1A myogenic cells and Lama2-/- mouse muscles. In addition, studies with sirtuin inhibitors and SIRT1 overexpression showed that caspase activation in MDC1A myotubes was inversely related to sirtuin deacetylase activity. Caspase activation in laminin-α2-deficiency was, however, not associated with increased phosphorylation of p38 MAPK. CONCLUSIONS: Aberrant caspase activation in MDC1A cells was mediated both by sirtuin deacetylase activity and by p53. Interventions that inhibit aberrant caspase activation by targeting sirtuin or p53 function could potentially be useful in ameliorating MDC1A.


Assuntos
Caspases/metabolismo , Laminina/genética , Fibras Musculares Esqueléticas/metabolismo , Distrofias Musculares/metabolismo , Sirtuínas/metabolismo , Células-Tronco/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Benzotiazóis/farmacologia , Histona Desacetilases do Grupo III/metabolismo , Humanos , Laminina/metabolismo , Camundongos , Camundongos Knockout , Desenvolvimento Muscular , Fibras Musculares Esqueléticas/efeitos dos fármacos , Distrofias Musculares/genética , Fosforilação , Proteínas Proto-Oncogênicas/metabolismo , Sirtuína 1/metabolismo , Células-Tronco/efeitos dos fármacos , Tolueno/análogos & derivados , Tolueno/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
9.
Mol Microbiol ; 107(4): 577-594, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29266439

RESUMO

Protein acetylation is a rapid mechanism for control of protein function. Acetyl-CoA synthetase (AMP-forming, Acs) is the paradigm for the control of metabolic enzymes by lysine acetylation. In many bacteria, type I or II protein acetyltransferases acetylate Acs, however, in actinomycetes type III protein acetyltransferases control the activity of Acs. We measured changes in the activity of the Streptomyces lividans Acs (SlAcs) enzyme upon acetylation by PatB using in vitro and in vivo analyses. In addition to the acetylation of residue K610, residue S608 within the acetylation motif of SlAcs was also acetylated (PKTRSGK610 ). S608 acetylation rendered SlAcs inactive and non-acetylatable by PatB. It is unclear whether acetylation of S608 is enzymatic, but it was clear that this modification occurred in vivo in Streptomyces. In S. lividans, an NAD+ -dependent sirtuin deacetylase from Streptomyces, SrtA (a homologue of the human SIRT4 protein) was needed to maintain SlAcs function in vivo. We have characterized a sirtuin-dependent reversible lysine acetylation system in Streptomyces lividans that targets and controls the Acs enzyme of this bacterium. These studies raise questions about acetyltransferase specificity, and describe the first Acs enzyme in any organism whose activity is modulated by O-Ser and Nɛ -Lys acetylation.


Assuntos
Acetato-CoA Ligase/metabolismo , Acetilcoenzima A/metabolismo , Lisina/metabolismo , Serina/metabolismo , Streptomyces lividans/enzimologia , Acetato-CoA Ligase/genética , Acetilação , Acetiltransferases/genética , Acetiltransferases/metabolismo , Aminoaciltransferases/genética , Aminoaciltransferases/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , DNA Bacteriano/genética , Deleção de Genes , Histona Desacetilases do Grupo III/genética , Histona Desacetilases do Grupo III/metabolismo , NAD/metabolismo , Streptomyces lividans/genética
10.
Med Res Rev ; 38(1): 147-200, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-28094444

RESUMO

Sirtuins are NAD+ -dependent protein deacylases that cleave off acetyl, as well as other acyl groups, from the ε-amino group of lysines in histones and other substrate proteins. Seven sirtuin isotypes (Sirt1-7) have been identified in mammalian cells. As sirtuins are involved in the regulation of various physiological processes such as cell survival, cell cycle progression, apoptosis, DNA repair, cell metabolism, and caloric restriction, a dysregulation of their enzymatic activity has been associated with the pathogenesis of neoplastic, metabolic, infectious, and neurodegenerative diseases. Thus, sirtuins are promising targets for pharmaceutical intervention. Growing interest in a modulation of sirtuin activity has prompted the discovery of several small molecules, able to inhibit or activate certain sirtuin isotypes. Herein, we give an update to our previous review on the topic in this journal (Schemies, 2010), focusing on recent developments in sirtuin biology, sirtuin modulators, and their potential as novel therapeutic agents.


Assuntos
Histona Desacetilases do Grupo III/antagonistas & inibidores , Histona Desacetilases do Grupo III/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Animais , Humanos , Terapia de Alvo Molecular
12.
Stem Cell Reports ; 9(6): 1839-1852, 2017 12 12.
Artigo em Inglês | MEDLINE | ID: mdl-29129681

RESUMO

Mitochondrial changes have long been implicated in the pathogenesis of Parkinson's disease (PD). The glycine to serine mutation (G2019S) in leucine-rich repeat kinase 2 (LRRK2) is the most common genetic cause for PD and has been shown to impair mitochondrial function and morphology in multiple model systems. We analyzed mitochondrial function in LRRK2 G2019S induced pluripotent stem cell (iPSC)-derived neurons to determine whether the G2019S mutation elicits similar mitochondrial deficits among central and peripheral nervous system neuron subtypes. LRRK2 G2019S iPSC-derived dopaminergic neuron cultures displayed unique abnormalities in mitochondrial distribution and trafficking, which corresponded to reduced sirtuin deacetylase activity and nicotinamide adenine dinucleotide levels despite increased sirtuin levels. These data indicate that mitochondrial deficits in the context of LRRK2 G2019S are not a global phenomenon and point to distinct sirtuin and bioenergetic deficiencies intrinsic to dopaminergic neurons, which may underlie dopaminergic neuron loss in PD.


Assuntos
Neurônios Dopaminérgicos/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Mitocôndrias/patologia , Doença de Parkinson/genética , Animais , Modelos Animais de Doenças , Neurônios Dopaminérgicos/citologia , Histona Desacetilases do Grupo III/genética , Humanos , Células-Tronco Pluripotentes Induzidas/patologia , Mitocôndrias/genética , Mutação , Neuritos/metabolismo , Doença de Parkinson/patologia , Doença de Parkinson/terapia
13.
Free Radic Biol Med ; 108: 929-939, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28506746

RESUMO

Silent information regulator 2 (Sir2) enzymes which catalyze NAD+-dependent protein/histone deacetylation. The mammalian sirtuin family SIRT1, SIRT2, SIRT3 and SIRT6 can regulate oxidative stress. The probiotics (Bifidobacterium longum(B.longum) and Lactobacillus acidophilus(L. acidophilus)) have Sir2 gene family and have antioxidant activity in human body. it remains unknown whether probiotics Sir2 has a direct role in regulating oxidative stress. To this end, we knockout BL-sir2(sir2 B. longum) and LA-sir2(sir2 L.acidophilus) in low oxygen level. The antioxidant activities of two sir2 deficient strains was decreased, while when reintroduction of BL-sir2 and LA-sir2, the antioxidant activities were recoveried. In order to understand the regulation mechanism of probiotics Sir2 oxidation response. Then, we screened 65 acetylated protein, and found that SigH (σH) was a substrate of BL-Sir2. In addition, the acetylation level of σH decreased with the increase of BL-Sir2 level in B. longum. Thus, BL-Sir2 deacetylated σH in response to oxidative stress. Next, we transfected BL-Sir2 into H2O2-induced oxidative damage of 293T cells, BL-Sir2 increased the activity of manganese superoxide dismutase (MnSOD/SOD2) and catalase (CAT) and reduced reactive oxygen species(ROS). Then, we analyzed the differential gene by RNA sequencing and Gene ontology (GO) and found that BL-Sir2 regulated forkhead transcription factor (FOXO3a) mediated antioxidant genes in overexpressed BL-Sir2 HEK293T cells. Our study is the first to link probiotics Sir2 with oxidative stress and uncover the antioxidant mechanism of BL-Sir2 in B. longum itself and human body.


Assuntos
Proteínas de Bactérias/metabolismo , Bifidobacterium longum/fisiologia , Histona Desacetilases do Grupo III/metabolismo , Lactobacillus acidophilus/fisiologia , Probióticos , Fator sigma/metabolismo , Acetilação , Proteínas de Bactérias/genética , Células HEK293 , Humanos , Peróxido de Hidrogênio/metabolismo , Microrganismos Geneticamente Modificados , NAD/metabolismo , Oxirredução , Estresse Oxidativo , RNA Interferente Pequeno/genética , Espécies Reativas de Oxigênio/metabolismo , Fator sigma/genética , Superóxido Dismutase/metabolismo , Ativação Transcricional
14.
Genetics ; 204(2): 569-579, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27527516

RESUMO

Nicotinamide is both a reaction product and an inhibitor of the conserved sirtuin family of deacetylases, which have been implicated in a broad range of cellular functions in eukaryotes from yeast to humans. Phenotypes observed following treatment with nicotinamide are most often assumed to stem from inhibition of one or more of these enzymes. Here, we used this small molecule to inhibit multiple sirtuins at once during treatment with DNA damaging agents in the Saccharomyces cerevisiae model system. Since sirtuins have been previously implicated in the DNA damage response, we were surprised to observe that nicotinamide actually increased the survival of yeast cells exposed to the DNA damage agent MMS. Remarkably, we found that enhanced resistance to MMS in the presence of nicotinamide was independent of all five yeast sirtuins. Enhanced resistance was also independent of the nicotinamide salvage pathway, which uses nicotinamide as a substrate to generate NAD+, and of a DNA damage-induced increase in the salvage enzyme Pnc1 Our data suggest a novel and unexpected function for nicotinamide that has broad implications for its use in the study of sirtuin biology across model systems.


Assuntos
Dano ao DNA/genética , Histona Desacetilases do Grupo III/genética , Nicotinamidase/genética , Proteínas de Saccharomyces cerevisiae/genética , Sirtuínas/genética , Dano ao DNA/efeitos dos fármacos , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Inativação Gênica , Histona Desacetilases do Grupo III/biossíntese , Metanossulfonato de Metila/toxicidade , Niacinamida/farmacologia , Fenótipo , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Sirtuínas/biossíntese
15.
G3 (Bethesda) ; 6(10): 3297-3305, 2016 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-27543294

RESUMO

Gene duplication promotes the diversification of protein functions in several ways. Ancestral functions can be partitioned between the paralogs, or a new function can arise in one paralog. These processes are generally viewed as unidirectional. However, paralogous proteins often retain related functions and can substitute for one another. Moreover, in the event of gene loss, the remaining paralog might regain ancestral functions that had been shed. To explore this possibility, we focused on the sirtuin deacetylase SIR2 and its homolog HST1 in the CTG clade of yeasts. HST1 has been consistently retained throughout the clade, whereas SIR2 is only present in a subset of species. These NAD+-dependent deacetylases generate condensed chromatin that represses transcription and stabilizes tandemly repeated sequences. By analyzing phylogenetic trees and gene order, we found that a single duplication of the SIR2/HST1 gene occurred, likely prior to the emergence of the CTG clade. This ancient duplication was followed by at least two independent losses of SIR2 Functional characterization of Sir2 and Hst1 in three species revealed that these proteins have not maintained consistent functions since the duplication. In particular, the rDNA locus is deacetylated by Sir2 in Candida albicans, by Hst1 in C. lusitaniae, and by neither paralog in C. parapsilosis In addition, the subtelomeres in C. albicans are deacetylated by Sir2 rather than by Hst1, which is orthologous to the sirtuin associated with Saccharomyces cerevisiae subtelomeres. These differences in function support the model that sirtuin deacetylases can regain ancestral functions to compensate for gene loss.


Assuntos
Candida/genética , Candida/metabolismo , Deleção de Genes , Duplicação Gênica , Histona Desacetilases do Grupo III/metabolismo , Acetilação , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Loci Gênicos , Genótipo , Histonas/metabolismo , Filogenia , Sirtuína 2/genética , Sirtuína 2/metabolismo , Telômero/genética , Telômero/metabolismo
16.
Exp Parasitol ; 169: 28-33, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27423969

RESUMO

Sir2 family proteins are highly conserved and catalyze Nicotinamide Adenine Dinucleotide (NAD(+))-dependent protein deacetylation reaction that regulates multiple cellular processes. Little is known about Sir2 family proteins in Giardia. In this research, Sir2 homologs of Giardia were Phylogenetically analyzed. GL50803_10707 (GlSIR2.2) showed strong homology to SIRT1 and was the only parasite SIRT1 homolog being reported to date. Recombinant GlSIR2.2 (rGlSIR2.2) was expressed and purified. The renaturied recombinant protein showed a typical NAD-dependent protein deacetylase activity that could be inhibited by nicotinamide, with IC50 of 4.47 mM rGlSIR2.2 displayed deacetylase activity under varied NAD(+), with Km, kcat and kcat/Km values of 31.71 µM, 1.4 × 10(-3) s(-1), and 4.42 × 10(-5) µM(-1) s(-1). Similarly, the steady-state kinetic parameters with varied ZMAL, yielded Km, kcat and kcat/Km values of 96.89 µM, 4.7 × 10(-3) s(-1), and 4.85 × 10(-5) µM(-1) s(-1). Anti-rGlSIR2.2 serum was used to probe subcellular localization of GlSIR2.2 and strong staining was found predominantly in the nucleus. So we demonstrated that GlSIR2.2 was a SIRT1-like, nuclear-located, NAD(+)-dependent deacetylase. This is the first report of deacetylase activity of Sir2 family protein in Giardia.


Assuntos
Núcleo Celular/enzimologia , Giardia lamblia/enzimologia , Histona Desacetilases do Grupo III/metabolismo , Sirtuínas/metabolismo , Sequência de Aminoácidos , Benzamidas/farmacologia , Técnica Indireta de Fluorescência para Anticorpo , Giardia lamblia/classificação , Giardia lamblia/ultraestrutura , Histona Desacetilases do Grupo III/antagonistas & inibidores , Histona Desacetilases do Grupo III/isolamento & purificação , Humanos , Concentração Inibidora 50 , Naftalenos/farmacologia , Naftóis/farmacologia , Niacinamida/farmacologia , Filogenia , Pironas/farmacologia , Alinhamento de Sequência , Sirtuínas/antagonistas & inibidores , Sirtuínas/isolamento & purificação
17.
Plant Sci ; 248: 28-36, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27181944

RESUMO

OsSRT1 is a NAD(+)-dependent histone deacetylase, closely related to the human SIRT6 that plays key roles in genome stability and metabolic homeostasis. In this work, we investigated the role of OsSRT1 in rice seed development. Down-regulation of OsSRT1 induced higher expression of Rice Starch Regulator1 (RSR1) and amylases genes in developing seeds, which resulted in a decrease of starch synthesis and an increase of starch degradation, leading to abnormal seed development. ChIP assay showed that OsSRT1 was required to reduce histone H3K9 acetylation on starch metabolism genes and transposons in developing seeds. In addition, OsSRT1 was detected to directly bind to starch metabolism genes such as OsAmy3B, OsAmy3E, OsBmy4, and OsBmy9. Our results suggested that OsSRT1-mediated histone deacetylation is involved in starch accumulation and transposon repression to regulate normal seed development.


Assuntos
Regulação da Expressão Gênica de Plantas/genética , Genes de Plantas/fisiologia , Histona Desacetilases do Grupo III/fisiologia , Oryza/crescimento & desenvolvimento , Proteínas de Plantas/fisiologia , Sementes/crescimento & desenvolvimento , Amido/metabolismo , Endosperma/crescimento & desenvolvimento , Regulação da Expressão Gênica de Plantas/fisiologia , Técnicas de Silenciamento de Genes , Genes de Plantas/genética , Histona Desacetilases do Grupo III/genética , Microscopia Eletrônica de Varredura , Análise de Sequência com Séries de Oligonucleotídeos , Oryza/metabolismo , Proteínas de Plantas/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sementes/metabolismo , Sementes/ultraestrutura
18.
J Neurochem ; 137(3): 371-83, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26896748

RESUMO

Aberrant epigenetic modifications are implicated in maternal diabetes-induced neural tube defects (NTDs). Because cellular stress plays a causal role in diabetic embryopathy, we investigated the possible role of the stress-resistant sirtuin (SIRT) family histone deacetylases. Among the seven sirtuins (SIRT1-7), pre-gestational maternal diabetes in vivo or high glucose in vitro significantly reduced the expression of SIRT 2 and SIRT6 in the embryo or neural stem cells, respectively. The down-regulation of SIRT2 and SIRT6 was reversed by superoxide dismutase 1 (SOD1) over-expression in the in vivo mouse model of diabetic embryopathy and the SOD mimetic, tempol and cell permeable SOD, PEGSOD in neural stem cell cultures. 2,3-dimethoxy-1,4-naphthoquinone (DMNQ), a superoxide generating agent, mimicked high glucose-suppressed SIRT2 and SIRT6 expression. The acetylation of histone 3 at lysine residues 56 (H3K56), H3K14, H3K9, and H3K27, putative substrates of SIRT2 and SIRT6, was increased by maternal diabetes in vivo or high glucose in vitro, and these increases were blocked by SOD1 over-expression or tempol treatment. SIRT2 or SIRT6 over-expression abrogated high glucose-suppressed SIRT2 or SIRT6 expression, and prevented the increase in acetylation of their histone substrates. The potent sirtuin activator (SRT1720) blocked high glucose-increased histone acetylation and NTD formation, whereas the combination of a pharmacological SIRT2 inhibitor and a pan SIRT inhibitor mimicked the effect of high glucose on increased histone acetylation and NTD induction. Thus, diabetes in vivo or high glucose in vitro suppresses SIRT2 and SIRT6 expression through oxidative stress, and sirtuin down-regulation-induced histone acetylation may be involved in diabetes-induced NTDs. The mechanism underlying pre-gestational diabetes-induced neural tube defects (NTDs) is still elusive. Our study unravels a new epigenetic mechanism in which maternal diabetes-induced oxidative stress represses sirtuin deacetylase 2 (SIRT2) and 6 (SIRT6) expression leading to histone acetylation and gene expression. SIRT down-regulation mediates the teratogenicity of diabetes leading to (NTD) formation. The study provides a mechanistic basis for the development of natural antioxidants and SIRT activators as therapeutics for diabetic embryopathy.


Assuntos
Glucose/toxicidade , Histona Desacetilases do Grupo III/biossíntese , Histona Desacetilases do Grupo III/genética , Histonas/metabolismo , Defeitos do Tubo Neural/induzido quimicamente , Defeitos do Tubo Neural/genética , Estresse Oxidativo/efeitos dos fármacos , Acetilação , Animais , Diabetes Gestacional/patologia , Epigênese Genética , Feminino , Histona Desacetilases do Grupo III/antagonistas & inibidores , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Camundongos , Naftoquinonas/farmacologia , Células-Tronco Neurais/metabolismo , Defeitos do Tubo Neural/prevenção & controle , Gravidez , Sirtuína 2/antagonistas & inibidores , Sirtuína 2/biossíntese , Sirtuína 2/genética , Sirtuínas/antagonistas & inibidores , Sirtuínas/biossíntese , Sirtuínas/genética , Superóxido Dismutase/biossíntese , Superóxido Dismutase/genética , Superóxido Dismutase-1
19.
Cell Cycle ; 13(18): 2821-6, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25486469

RESUMO

Melanoma continues to cause more deaths than any other skin cancer, necessitating the development of new avenues of treatment. One promising new opportunity comes in the form of mechanism-based therapeutic targets. We recently reported the overexpression and delocalization of the class III histone deacetylase SIRT1 in melanoma, and demonstrated that its small molecule inhibition via Tenovin-1 decreased cell growth and viability of melanoma cells, possibly by a p53 mediated induction of p21. Here, we support our data using additional SIRT inhibitors, viz. Sirtinol and Ex-527, which suggests possible benefits of concomitantly inhibiting more than one Sirtuin for an effective cancer management strategy. This "Extra View" paper also includes a discussion of our results in the context of similar recent and concurrent studies. Furthermore, we expand upon our findings in an analysis of new research that may link the cellular localization and growth effects of SIRT1 with the PI3K signaling pathway.


Assuntos
Benzamidas/uso terapêutico , Carbazóis/uso terapêutico , Histona Desacetilases do Grupo III/antagonistas & inibidores , Melanoma/tratamento farmacológico , Terapia de Alvo Molecular , Naftóis/uso terapêutico , Apoptose/efeitos dos fármacos , Benzamidas/farmacologia , Carbazóis/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Clonais , Humanos , Melanoma/patologia , Modelos Biológicos , Naftóis/farmacologia , Transdução de Sinais/efeitos dos fármacos
20.
Nucleic Acids Res ; 42(20): 12600-13, 2014 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-25300492

RESUMO

The African trypanosome, Trypanosoma brucei, is a parasitic protozoan that achieves antigenic variation through DNA-repair processes involving Variant Surface Glycoprotein (VSG) gene rearrangements at subtelomeres. Subtelomeric suppression of DNA repair operates in eukaryotes but little is known about these controls in trypanosomes. Here, we identify a trypanosome histone acetyltransferase (HAT3) and a deacetylase (SIR2rp1) required for efficient RAD51-dependent homologous recombination. HAT3 and SIR2rp1 were required for RAD51-focus assembly and disassembly, respectively, at a chromosome-internal locus and a synthetic defect indicated distinct contributions to DNA repair. Although HAT3 promoted chromosome-internal recombination, it suppressed subtelomeric VSG recombination, and these locus-specific effects were mediated through differential production of ssDNA by DNA resection; HAT3 promoted chromosome-internal resection but suppressed subtelomeric resection. Consistent with the resection defect, HAT3 was specifically required for the G2-checkpoint response at a chromosome-internal locus. HAT3 also promoted resection at a second chromosome-internal locus comprising tandem-duplicated genes. We conclude that HAT3 and SIR2rp1 can facilitate temporally distinct steps in DNA repair. HAT3 promotes ssDNA formation and recombination at chromosome-internal sites but has the opposite effect at a subtelomeric VSG. These locus-specific controls reveal compartmentalization of the T. brucei genome in terms of the DNA-damage response and suppression of antigenic variation by HAT3.


Assuntos
Variação Antigênica/genética , Histona Desacetilases do Grupo III/fisiologia , Histona Acetiltransferases/fisiologia , Proteínas de Protozoários/fisiologia , Reparo de DNA por Recombinação , Trypanosoma brucei brucei/genética , Glicoproteínas Variantes de Superfície de Trypanosoma/genética , Quebras de DNA de Cadeia Dupla , DNA de Cadeia Simples/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Loci Gênicos , Histona Desacetilases do Grupo III/genética , Histona Acetiltransferases/genética , Proteínas de Protozoários/genética , Rad51 Recombinase/metabolismo , Telômero , Trypanosoma brucei brucei/enzimologia , Trypanosoma brucei brucei/imunologia
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