Hydrolase Regulates NAD+ Metabolites and Modulates Cellular Redox

Although the classical redox functions of co-enzyme NAD⁺ are firmly established in metabolism, there are numerous enzymes that catalyze cleavage of NAD⁺ to yield free ADP-ribose (ADPr) or related metabolites, whose functions remain largely unknown. Here we show that the Nudix (nucleoside diphosphate linked to another moiety X) hydrolase Ysa1 from Saccharomyces cerevisiae is a major regulator of cellular ADPr and O-acetyl-ADP-ribose (OAADPr). OAADPr is the direct product of NAD⁺-dependent protein deacetylases (sirtuins) and is readily converted to ADPr. Ysa1 cleaves ADPr/OAADPr into ribose phosphate/acetyl-ribose phosphate and AMP. In cells lacking Ysa1 (Δysa1), ADPr and OAADPr levels increased ∼50%, with a corresponding decrease in AMP. Strikingly, Δysa1 cells display higher resistance to exogenous reactive oxygen species (ROS) and 40% lower basal levels of endogenous ROS, compared with wild type. The biochemical basis for these differences in ROS-related phenotypes was investigated, and the results provide evidence that increased ADPr/OAADPr levels protect cells via the following two pathways: (i) lower ROS production through inhibition of complex I of the mitochondrial electron transport chain, and (ii) generation of higher levels of NADPH to suppress ROS damage. The latter occurs through diverting glucose into the pentose phosphate pathway by ADPr inhibition of glyceraldehyde-3-phosphate dehydrogenase, a central enzyme of glycolysis.NAD⁺ is well known for its role as a hydride-transferring co-enzyme in many oxidation-reduction reactions of metabolism. However, NAD⁺ is also a substrate for NAD⁺ glycohydrolases, ADP-ribose transferases, poly(ADP-ribose) polymerases (PARPs),² cyclic ADP-ribose synthases (1, 2), and sirtuins (3, 4), all of which cleave the glycosidic bond of NAD⁺ to produce nicotinamide and an ADP-ribosyl product. Notably, sirtuins catalyze NAD⁺-dependent lysine deacetylation to generate nicotinamide, deacetylated lysine, and OAADPr (5, 6). OAADPr has been proposed to act as a second messenger, signaling to other processes that NAD⁺-dependent protein deacetylation has occurred (7–9). The biological functions and in vivo metabolism of OAADPr and free ADPr are largely unknown.Through a quantitative microinjection assay of starfish oocytes, both ADPr and OAADPr caused a delay/block in oocyte maturation, suggesting ADPr/OAADPr may have specific biological activity (10). In mammalian cells, intracellular ADPr/OAADPr can activate the TRPM2 (transient receptor melastatin-related ion channel 2) nonselective cationic channel (11–13). TRPM2 contains a conserved intracellular Nudix hydrolase domain (referred to as NudT9H) that directly binds ADPr/OAADPr, but it is incapable of cleaving the ligand because a major catalytic residue is missing (11, 14). Although still disputed, ADPr binding to NudT9H appears to be required for the well known oxidative stress activation of the channel (13, 15). Cell stress via puromycin treatment led to TRPM2-mediated cell death that was dependent on sirtuin deacetylases, presumably from the production of OAADPr (12).Increasing evidence suggests that free ADPr may function as a cellular signal. ADPr can be produced from the coordinate actions of PARPs and poly(ADP-ribose) glycohydrolase (PARG), which cleave ADPr polymers to free ADPr (16, 17). Under massive genotoxic stress, hyper-stimulation of the NAD⁺-dependent PARPs depletes cellular NAD⁺, which is linked to catastrophic ATP loss and cell death (18, 19). The mechanism by which PARP1 hyperactivity in the nucleus impairs ATP production in mitochondria is unclear. The fact that PARP1 and poly(ADP-ribose) are localized in the nucleus adds a perplexing aspect. However, recent data suggest that PARP1-induced loss of ATP requires PARG (20). Under conditions of PARP1 hyperactivation, it has been suggested that the PARG-dependent production of ADPr can exit the nucleus and interfere with ATP production in mitochondria (21, 22). Thus ADPr could be the molecular signal released from the nucleus of cells undergoing massive poly(ADP-ribosyl)ation and rapidly triggers mitochondrial dysfunction.In support for ADPr/OAADPr as potential signaling molecules, the existence of enzymes capable of metabolizing these compounds suggests that their cellular concentrations may be subject to tight regulation (23, 24). To understand the biological roles played by ADPr/OAADPr, it is essential to elucidate the degradation pathways that can modulate their levels. Previously we described the ability of several conserved members of the Nudix hydrolase family to hydrolyze in vitro the diphosphate linkage in ADPr/OAADPr, generating ribose phosphate or acetyl-ribose phosphate and AMP (10, 24). Here we examine the biochemical and cellular functions of the Nudix hydrolase Ysa1 (14) from Saccharomyces cerevisiae. We determined that Ysa1 is the major ADPr Nudix hydrolase and an important regulator of cellular ADPr/OAADPr levels. A Δysa1 strain displays increased resistance to both exogenously and endogenously generated ROS. Basal level of ROS decreased by 40% in the Ysa1 deletion strain. We provide biochemical evidence that increased ADPr/OAADPr levels protect cells via the following two pathways: (i) lower ROS production through the inhibition of complex I of the electron transport chain, and (ii) generation of higher NADPH levels to suppress ROS damage. The latter occurs by diverting glucose into the pentose phosphate pathway by ADPr inhibition of glycolysis.     

Sirtuin:依赖NAD+的去乙酰化酶

组蛋白的乙酰化一去乙酰化修饰在基因表达调控中起重要作用。参与去乙酰化的酶除了经典的Ⅰ类和Ⅱ类组蛋白去乙酰化酶(histone deacetylase,HDAC),还有比较特殊的Ⅲ类HDAC——Sirnlin,其活性依赖于NAD^ 。酵母的Sirtuin——Sir2在交配型基因沉默、端粒区基因沉默、rDNA沉默中起重要作用．还可能参与长寿与衰老的调节。在人类,Sirtuin的底物是组蛋白、各种转录因子如p53、FOXO、NF—KB、乙酰化酶如D300和其他的各种功能蛋白质。根据底物特点推测,人类Sirtuin蛋白的生理功能可能一方面是参与调节细胞在应激条件下的存活与死亡的平衡,另一方面是参与代谢的调节。     

地衣次生代谢产物及其生物活性研究进展

地衣是一类独特的生物有机体,其体内合成了多种特殊的次生代谢产物,主要为缩酚酸、缩酚酸环醚、缩酚酮、二苯并呋喃等类化合物。这些次生代谢产物具有多样的生物活性,包括抗氧化、抗辐射、抗菌、抗病毒、抗肿瘤、植物生长抑制和昆虫拒食等活性。本文对近40年来有关地衣化学及生物活性方面的研究进行综述,为地衣资源的进一步研究和开发提供参考。     

内生真菌香柱菌及其次级代谢

两千年以前人们就已经知道一些牧草对牲畜等动物有毒。 1 90 4年Ｆｒｅｅｍａｎ从这些牧草中分离到了一类内生真菌 ,后被称为Ｎｅｏｔｙｐｈｏｄｉｕｍｏｃｃｕｌｔａｎｓ,人们才知晓造成这些动物中毒的真正元凶恰恰是这类微生物[1] 。在 2 0世纪 30年代 ,Ｓａｍｐｓｏｎ对一些植物的内生真菌作了比较性研究[2 ] ,这些内生真菌包括Ｎｅ ｏｔｙｐｈｏｄｉｕｍｌｏｉｉ和香柱菌属 (Ｅｐｉｃｈｌｏｅ)的一些种 ,其中香柱菌可以导致一些草类干死病 (ｃｈｏｋｅ)。而在大多数情况下 ,香柱菌及其相关的内生真菌长期生存于它们的宿主植物中 ,并不使…     

Mitochondrial Matrix Ca2+ Accumulation Regulates Cytosolic NAD+/NADH Metabolism,Protein Acetylation,and Sirtuin Expression

Mitochondrial calcium uptake stimulates bioenergetics and drives energy production in metabolic tissue. It is unknown how a calcium-mediated acceleration in matrix bioenergetics would influence cellular metabolism in glycolytic cells that do not require mitochondria for ATP production. Using primary human endothelial cells (ECs), we discovered that repetitive cytosolic calcium signals (oscillations) chronically loaded into the mitochondrial matrix. Mitochondrial calcium loading in turn stimulated bioenergetics and a persistent elevation in NADH. Rather than serving as an impetus for mitochondrial ATP generation, matrix NADH rapidly transmitted to the cytosol to influence the activity and expression of cytosolic sirtuins, resulting in global changes in protein acetylation. In endothelial cells, the mitochondrion-driven reduction in both the cytosolic and mitochondrial NAD⁺/NADH ratio stimulated a compensatory increase in SIRT1 protein levels that had an anti-inflammatory effect. Our studies reveal the physiologic importance of mitochondrial bioenergetics in the metabolic regulation of sirtuins and cytosolic signaling cascades.     

江西青霉的次生代谢产物研究

江西青霉是药用江西虫草的无性型,本文对江西青霉发酵菌丝体甲醇提取物的正丁醇萃取部位运用反复色谱层析进行了系统的分离纯化,得到了6个化合物。经波谱解析,并结合理化鉴定,确定这6个化合物结构为尿嘧啶(1)2、’-脱氧尿嘧啶核苷(2)、腺嘌呤(3)、腺苷(4)、L-焦谷氨酸甲酯(5)和2’-甲氧基腺苷(6)。其中化合物2、5和6为首次从虫草属中分离获得的化合物。     

Organic Matrix and Secondary Metabolites in Nacre

Marine Biotechnology - Nacre, also called mother-of-pearl, is a naturally occurring biomineral, largely studied by chemists, structural biologists, and physicists to understand its outstanding and...     

Mitochondrial NAD+ Controls Nuclear ARTD1-Induced ADP-Ribosylation

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Fungus-Specific Sirtuin HstD Coordinates Secondary Metabolism and Development through Control of LaeA

The sirtuins are members of the NAD⁺-dependent histone deacetylase family that contribute to various cellular functions that affect aging, disease, and cancer development in metazoans. However, the physiological roles of the fungus-specific sirtuin family are still poorly understood. Here, we determined a novel function of the fungus-specific sirtuin HstD/Aspergillus oryzae Hst4 (AoHst4), which is a homolog of Hst4 in A. oryzae yeast. The deletion of all histone deacetylases in A. oryzae demonstrated that the fungus-specific sirtuin HstD/AoHst4 is required for the coordination of fungal development and secondary metabolite production. We also show that the expression of the laeA gene, which is the most studied fungus-specific coordinator for the regulation of secondary metabolism and fungal development, was induced in a ΔhstD strain. Genetic interaction analysis of hstD/Aohst4 and laeA clearly indicated that HstD/AoHst4 works upstream of LaeA to coordinate secondary metabolism and fungal development. The hstD/Aohst4 and laeA genes are fungus specific but conserved in the vast family of filamentous fungi. Thus, we conclude that the fungus-specific sirtuin HstD/AoHst4 coordinates fungal development and secondary metabolism via the regulation of LaeA in filamentous fungi.     

放线菌次级代谢产物研究进展

随着耐药细菌和新型病毒的不断出现，癌症的发病率和死亡率持续上升，迫使人们不断寻找新的化合物来治疗疾病。放线菌次级代谢产物结构新颖，作用独特，具有抗菌、杀虫、抗肿瘤、免疫抑制等活性，广泛应用于医疗、农业、食品等领域，深入挖掘放线菌资源来开发新型抗生素潜力巨大。然而从自然界分离的放线菌生产目标化合物的能力较弱，这直接影响其工业应用，增加其生产成本，因此构建目标化合物高产菌株显得尤为重要。本文以此为出发点，从放线菌新药资源挖掘和放线菌产抗能力提高两个方面对近年来的研究情况进行概述，为放线菌资源开发提供参考。     

Secondary Metabolites and Biological Properties of Gesneriaceae Species

The family Gesneriaceae comprises ca. 150 genera and 3000 species, distributed in the tropics around the world. It is constituted of herbs, lianas, or shrubs, frequently with ornamental potential, due to the beauty of their flowers. Some species have been used in traditional medicine, mainly against fever, cough, colds, snakebite, pains, and infectious and inflammatory diseases. Although Gesneriaceae are a large family, only few species were chemically investigated, and this took place mainly in the last decade. In the present work, chemical and pharmacological studies on Gesneriaceae are reviewed based on original articles published. Altogether 300 compounds have been reported in Gesneriaceae species, including flavonoids, terpenes and steroids, phenolic glucosides, simple phenolics, quinones, lignans, xanthones, and compounds with unusual skeletons. Several species had been used in folk medicine, and some constituents have shown biological activities, such as antimicrobial, anti‐inflamatory, antioxidant, and antitumor properties.     

次生代谢产物与植物抗病防御反应

次生代谢产物是由植物次生代谢产生的许多结构不同的小分子有机化合物,它们广泛参与植物的生长、发育、防御等生理过程。次生代谢产物在植物的抗病防御反应中发挥着重要作用,可以作为生化壁垒防御病原物侵染,还可以作为信号物质参与植物的抗病反应;在植物与病原物互作中,植物合成新的抗菌物质植保素,原有的抗菌物质也会增加。植物次生代谢产物的积累受到病原物、发育,环境等多种因素的调节。本文重点介绍次生代谢产物在植物抗病防御中的相关作用以及影响其合成的各种因素。     

海洋沉积环境来源真菌次生代谢产物的研究

海洋沉积环境复杂多样,微生物往往进化形成与其环境相适应的代谢系统.真菌作为海洋沉积环境中的重要微生物,它们的次生代谢产物结构出新率高、生物活性显著,是开发海洋先导药物的新兴资源.本文综述了1995-2011年间发表的200个海洋沉积环境真菌次生代谢产物,其来源菌株广泛分布在真菌16个属,主要集中在Penicillium属(30％)、Aspergillus属(19％)、Spicaria属(18％)和Trichoderma属(13％);其结构类型包括生物碱类(86个)、萜类(42个)、聚酮类(42个)、肽类(15个)、甾体类(5个)、脑苷脂类(4个)和其他类(6个),活性研究主要集中在细胞毒性和抗菌等方面.