Discovery of genes for ginsenoside biosynthesis by analysis of ginseng expressed sequence tags

Expressed sequence tags (ESTs) provide a valuable tool that can be used to identify genes in secondary metabolite biosynthesis. Ginseng (Panax ginseng C.A Meyer) is a medicinal plant that accumulates ginsenosides in roots. We sequenced 11,636 ESTs from five ginseng libraries in order to create a gene resource for biosynthesis of ginsenosides, which are thought to be the major active component in roots. Only 59% of the ginseng ESTs exhibited significant homology to previously known polypeptide sequences. Stress- and pathogen-response proteins were most abundant in 4-year-old ginseng roots. ESTs involved in ginsenoside biosynthesis were identified by a keyword search of BLASTX results and a domain search of ginseng ESTs. We identified 4 oxidosqualene cyclase candidates involved in the cyclization reaction of 2,3-oxidosqualene, 9 nine cytochrome P450 and 12 glycosyltransferse candidates, which may be involved in modification of the triterpene backbone.Abbreviations cDNA Complementary DNA - ESTs Expressed sequence tagsCommunicated by I.S. Chung     

Supercomputer analysis of purine and pyrimidine metabolism leading to DNA synthesis

A model-system is established to analyze purine and pyrimidine metabolism leading to DNA synthesis. The principal aim is to explore the flow and regulation of terminal deoxynucleoside triophosphates (dNTPs) in various input and parametric conditions. A series of flow equations are established, which are subsequently converted to differential equations. These are programmed (Fortran) and analyzed on a Cray X-MP/48 supercomputer. The pool concentrations are presented as a function of time in conditions in which various pertinent parameters of the system are modified. The system is formulated by 100 differential equations.     

Deglutarylation of glutaryl-CoA dehydrogenase by deacylating enzyme SIRT5 promotes lysine oxidation in mice

A wide range of protein acyl modifications has been identified on enzymes across various metabolic processes; however, the impact of these modifications remains poorly understood. Protein glutarylation is a recently identified modification that can be nonenzymatically driven by glutaryl-CoA. In mammalian systems, this unique metabolite is only produced in the lysine and tryptophan oxidative pathways. To better understand the biology of protein glutarylation, we studied the relationship between enzymes within the lysine/tryptophan catabolic pathways, protein glutarylation, and regulation by the deglutarylating enzyme sirtuin 5 (SIRT5). Here, we identify glutarylation on the lysine oxidation pathway enzyme glutaryl-CoA dehydrogenase (GCDH) and show increased GCDH glutarylation when glutaryl-CoA production is stimulated by lysine catabolism. Our data reveal that glutarylation of GCDH impacts its function, ultimately decreasing lysine oxidation. We also demonstrate the ability of SIRT5 to deglutarylate GCDH, restoring its enzymatic activity. Finally, metabolomic and bioinformatic analyses indicate an expanded role for SIRT5 in regulating amino acid metabolism. Together, these data support a feedback loop model within the lysine/tryptophan oxidation pathway in which glutaryl-CoA is produced, in turn inhibiting GCDH function via glutaryl modification of GCDH lysine residues and can be relieved by SIRT5 deacylation activity.     

Changes in the lipid and fatty acid composition of hemolymph and ovaries during the reproductive cycle of Labidura riparia

Lipid metabolism was investigated during the reproductive cycle of Labidura riparia (Pallas). The lipid classes and their constitutive fatty acids present in hemolymph and ovaries were measured using thin‐layer chromatography and gas‐liquid chromatography. In the hemolymph, total lipids increase steadily from the previtellogenic period to vitellogenic arrest. These lipids are predominantly diacylglycerols and phospholipids. In the ovaries, total lipids increase during vitellogenesis then decrease during the vitellogenesis arrest period. The major lipids are triacylglycerols, followed by phospholipids. In both hemolymph and ovaries, all lipid classes contained variable proportions of seven main fatty acids: the saturated fatty acids myristic acid (14:0), palmetic acid (16:0), and stearic acid (18:0); the monounsaturated fatty acids palmitoleic acid (16:1) and oleic acid (18:1); and the polyunsaturated fatty acids linoleic acid (18:2) and linolenic acid (18:3). Unsaturated fatty acids predominate throughout the reproductive cycle. The percentage compositions of total and triacylglycerol fatty acids do not change markedly during the reproductive cycle in hemolymph nor in ovaries, with 18:2, 18:1 and 16:0 fatty acids being the major components. However, for diacylglycerols and phospholipids, the proportions of fatty acids vary systematically. For phospholipids during the vitellogenesis period, 18:2 increases considerably whereas other fatty acids decrease; for diacylglycerols, these fatty acids vary in the reverse way.     

Deoxyribonuclease 1-like 3 Inhibits Hepatocellular Carcinoma Progression by Inducing Apoptosis and Reprogramming Glucose Metabolism

HCC has remained one of the challenging cancers to treat, owing to the paucity of drugs targeting the critical survival pathways. Considering the cancer cells are deficient in DNase activity, the increase of an autonomous apoptisis endonuclease should be a reasonable choice for cancer treatment. In this study, we investigated whether DNASE1L3, an endonuclease implicated in apoptosis, could inhibit the progress of HCC. We found DNASE1L3 was down-regulated in HCC tissues, whereas its high expression was positively associated with the favorable prognosis of patients with HCC. Besides, serum DNASE1L3 levels were lower in HCC patients than in healthy individuals. Functionally, we found that DNASE1L3 inhibited the proliferation of tumor cells by inducing G0/G1 cell cycle arrest and cell apoptosis in vitro. Additionally, DNASE1L3 overexpression suppressed tumor growth in vivo. Furthermore, we found that DNASE1L3 overexpression weakened glycolysis in HCC cells and tissues via inactivating the rate-limiting enzymes involved in PTPN2-HK2 and CEBPβ-p53-PFK1 pathways. Finally, we identified the HBx to inhibit DNASE1L3 expression by up-regulating the expression of ZNF384. Collectively, our findings demonstrated that DNASE1L3 could inhibit the HCC progression through inducing cell apoptosis and weakening glycolysis. We believe DNASE1L3 could be considered as a promising prognostic biomarker and therapeutic target for HCC.     

Lipids and lipid metabolism in the brown alga, Fucus serratus

The lipids of the brown alga Fucus serratus were isolated, identified and quantified. The major acyl lipids were the three glycosylglycerides, diacylgalactosylglycerol, diacyldigalactosylglycerol and diacylsulphoquinovosylglycerol. These represent over 70% of the total acyl lipids. The fatty acid compositions of the major lipids were examined and most showed rather distinctive fatty acid contents. For example, diacylgalactosylglycerol was enriched in n-3 polyunsaturated fatty acids while phosphatidylcholine and phosphatidylethanolamine had very high levels of arachidonate. Phosphatidylglycerol contained the unusual trans-Δ3-hexadecenoic acid. The labelling of lipids and fatty acids from [¹⁴C]acetate was examined and the distribution of label between individual components as a function of the incubation period and in algae collected at different times of the year is reported. Algae collected in the winter incorporated much more radioactivity into non-esterified fatty acids when compared to algae collected in the summer. All algae could label myristate, palmitate, stearate and oleate at high rates. Longer incubation times allowed the labelling of polyunsaturated fatty acids such as linoleic acid.     

Activation of Toll-like Receptor 4 (TLR4) Attenuates Adaptive Thermogenesis via Endoplasmic Reticulum Stress

Adaptive thermogenesis is the cellular process transforming chemical energy into heat in response to cold. A decrease in adaptive thermogenesis is a contributing factor to obesity. However, the molecular mechanisms responsible for the compromised adaptive thermogenesis in obese subjects have not yet been elucidated. In this study we hypothesized that Toll-like receptor 4 (TLR4) activation and subsequent inflammatory responses are key regulators to suppress adaptive thermogenesis. To test this hypothesis, C57BL/6 mice were either fed a palmitate-enriched high fat diet or administered with chronic low-dose LPS before cold acclimation. TLR4 stimulation by a high fat diet or LPS were both associated with reduced core body temperature and heat release. Impairment of thermogenic activation was correlated with diminished expression of brown-specific markers and mitochondrial dysfunction in subcutaneous white adipose tissue (sWAT). Defective sWAT browning was concomitant with elevated levels of endoplasmic reticulum (ER) stress and autophagy. Consistently, TLR4 activation by LPS abolished cAMP-induced up-regulation of uncoupling protein 1 (UCP1) in primary human adipocytes, which was reversed by silencing of C/EBP homologous protein (CHOP). Moreover, the inactivation of ER stress by genetic deletion of CHOP or chemical chaperone conferred a resistance to the LPS-induced suppression of adaptive thermogenesis. Collectively, our data indicate the existence of a novel signaling network that links TLR4 activation, ER stress, and mitochondrial dysfunction, thereby antagonizing thermogenic activation of sWAT. Our results also suggest that TLR4/ER stress axis activation may be a responsible mechanism for obesity-mediated defective brown adipose tissue activation.     

A 31P-NMR study of the intact liver fluke Fasciola hepatica

³¹P-NMR techniques offer a useful method of studying changes in the metabolism of intact parasitic worms. The liver flukes, Fasciola hepatica, provide good quality ³¹P high resolution NMR spectra for at least 6 h under anaerobic conditions. The levels of ATP remain constant throughout this period. There is no signal for phosphocreatine or phosphoarginine. In contrast to the findings in mammalian tissues, there is a distinct peak for the terminal phosphate of ADP. A number of signals are observed in the phosphodiester region of the spectrum the largest of which is identified as l-α-glycerophosphoryl choline. Serotonin (5-hydroxytryptamine) causes an appreciable increase in the levels of sugar phosphates when the flukes are incubated in the absence of glucose. The addition of glucose also causes a marked increase in the signals for the hexose phosphate.