Cyclic Phosphate Research Articles

Copper-Catalyzed Enantioselective Allyl-Allyl Coupling between Allylic Boronates and Phosphates with a Phenol/N-Heterocyclic Carbene Chiral Ligand.

Copper-catalyzed enantioselective allyl-allyl coupling between allylboronates and either Z-acyclic or cyclic allylic phosphates using a new chiral N-heterocyclic carbene ligand, bearing a phenolic hydroxy, is reported. This reaction occurs with exceptional SN 2'-type regioselectivities and high enantioselectivities to deliver chiral 1,5-diene derivatives with a tertiary stereogenic center at the allylic/homoallylic position.

In vitro differentiation of rat spermatogonia into round spermatids in tissue culture.

STUDY QUESTIONDo the organ culture conditions, previously defined for in vitro murine male germ cell differentiation, also result in differentiation of rat spermatogonia into post-meiotic germ cells exhibiting specific markers for haploid germ cells?SUMMARY ANSWERWe demonstrated the differentiation of rat spermatogonia into post-meiotic cells in vitro, with emphasis on exhibiting, protein markers described for round spermatids.WHAT IS KNOWN ALREADYFull spermatogenesis in vitro from immature germ cells using an organ culture technique in mice was first reported 5 years ago. However, no studies reporting the differentiation of rat spermatogonia into post-meiotic germ cells exhibiting the characteristic protein expression profile or into functional sperm have been reported.STUDY DESIGN, SAMPLES/MATERIALS, METHODSOrgan culture of testicular fragments of 5 days postpartum (dpp) neonatal rats was performed for up to 52 days. Evaluation of microscopic morphology, testosterone levels, mRNA and protein expression as measured by RT-qPCR and immunostaining were conducted to monitor germ cell differentiation in vitro. Potential effects of melatonin, Glutamax® medium, retinoic acid and the presence of epidydimal fat tissue on the spermatogenic process were evaluated. A minimum of three biological replicates were performed for all experiments presented in this study. One-way ANOVA, ANOVA on ranks and student's t-test were applied to perform the statistical analysis.MAIN RESULTS AND THE ROLE OF CHANCEMale germ cells, present in testicular tissue pieces grown from 5 dpp rats, exhibited positive protein expression for Acrosin and Crem (cAMP (cyclic adenosine mono phosphate) response element modulator) after 52 days of culture in vitro. Intra-testicular testosterone production could be observed after 3 days of culture, while when epididymal fat tissue was added, spontaneous contractility of cultured seminiferous tubules could be observed after 21 days. However, no supportive effect of the supplementation with any factor or the co-culturing with epididymal fat tissue on germ cell differentiation in vitro or testosterone production was observed.LIMITATIONS, REASONS FOR CAUTIONThe human testis is very different in physiology from the rat testis, further investigations are still needed to optimize the organ culture system for future use in humans.WIDER IMPLICATIONS OF THE FINDINGSThe successful differentiation of undifferentiated spermatogonia using the testis explant culture system might be employed in future to produce sperm from human spermatogonia as a clinical tool for fertility preservation in boys and men suffering infertility.LARGE SCALE DATANone.STUDY FUNDING AND COMPETING INTEREST(S)This work was supported financially by the Frimurare Barnhuset in Stockholm, the Paediatric Research Foundation, Jeanssons Foundation, Sällskåpet Barnåvard in Stockholm, Swedish Research Council/Academy of Finland, Emil and Wera Cornells Foundation, Samariten Foundation, the Swedish Childhood Cancer Foundation as well as through the regional agreement on medical training and clinical research (ALF) between Stockholm County Council and Karolinska Institutet. All authors declare no conflicts of interests.

Abstract 2665: SHOT-RNAs: A novel class of tRNA-derived functional RNAs expressed in hormone-dependent cancers

Abstract Sex hormones and their receptors play critical roles in the genesis and progression of breast and prostate cancers. We here present that sex hormone signaling pathways promote the expression of specific tRNA halves termed Sex HOrmone-dependent TRNA-derived RNAs (SHOT-RNAs). Screenings using 94 cancer cell lines indicated that SHOT-RNAs are specifically and abundantly expressed in estrogen receptor-positive (ER+) breast cancer and androgen receptor-positive (AR+) prostate cancer. None of the ER− breast cancer, AR− prostate cancer, or other examined cancer cell lines from other tissues exhibited abundant expression of SHOT-RNAs. ER-dependent accumulation of SHOT-RNAs is not limited to a cell culture system, but it also occurs in luminal-type breast cancer patient tissues. The expression specificity allowed us to hypothesize that sex hormones and their receptors are the crucial molecular factors for SHOT-RNA generation. Our hypothesis has been confirmed by the following three observations: (1) siRNA-mediated reduction of ER or AR decreased SHOT-RNA levels, (2) hormone-depletion from the culture medium decreased SHOT-RNA levels, and (3) addition of the sex hormone, estrogen or androgen, to the medium increased SHOT-RNA levels. These results indicate the clear dependency of SHOT-RNA expressions on sex hormones and their receptors. SHOT-RNAs are produced from aminoacylated mature tRNAs by angiogenin (ANG)-mediated anticodon cleavage. Resultant 5′- and 3′-SHOT-RNAs, corresponding to 5′- and 3′-tRNA halves, bear 3′-terminal modifications, cyclic phosphate (cP) and amino acid, respectively. Thus, SHOT-RNAs would not be captured by standard RNA-seq methods because the modification inhibits adapter ligation steps. This could be the reason why SHOT-RNAs had not been discovered in spite of their abundance and expression specificity. To identify a comprehensive repertoire of SHOT-RNAs, we developed a method named “cP-RNA-seq” in which 3′-termini of all RNAs, except for those containing a 3′-terminal cP, were cleaved by a periodate treatment, thereby selectively amplifying and sequencing only cP-containing RNAs. The application of cP-RNA-seq to breast cancer cells identified eight cytoplasmic tRNA species as major sources of 5′-SHOT-RNAs. We further investigated the functional significance of SHOT-RNA expression. Intriguingly, siRNA-directed reduction of each one of the three different species of 5′-SHOT-RNA severely impaired cell proliferation, whereas 3′-SHOT-RNA reduction showed no impact. These results strongly suggest that at least 5′-SHOT-RNAs are expressed as functional RNA molecules promoting cell growth. Our results have unveiled a novel tRNA-engaged pathway in tumorigenesis of hormone-dependent cancers and implicate SHOT-RNAs as potential candidates for biomarkers and therapeutic targets. We will discuss our working model and future challenges. Citation Format: Shozo Honda, Megumi Shigematsu, Phillipe Loher, Juan P. Palazzo, Issei Imoto, Isidore Rigoutsos, Yohei Kirino. SHOT-RNAs: A novel class of tRNA-derived functional RNAs expressed in hormone-dependent cancers. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2665.

Structure and pasting properties of alkaline-treated phosphorylated cross-linked waxy maize starches

The objectives of this study were to determine the stability of cross-linked bonds of starch at different pH values and their effects on the pasting property of waxy maize starch cross-linked by 0.05% and 3% sodium trimetaphosphate/sodium tripolyphosphate. The cross-linked waxy maize starch (CLWMS) was slurried (40%, w/w) and subjected to alkali treatments of pH 9, 10, 11, and 12 at 40°C for 4h. The phosphorus in 3% CLWMS decreased with increasing pH and remained unchanged in 0.05% CLWMS for all pH treatments. Decreased settling volumes indicated the reduction of swelling power for the alkali-treated CLWMS at pH 11 and 12. The 31P NMR spectra of 3% CLWMS at pH 12 showed decreased cyclic monostarch phosphate, monostarch monophosphate, and monostarch diphosphate, but significantly increased distarch monophosphate. Alkali treatments of phosphorylated cross-linked starches offer a way to manipulate the rheological properties of cross-linked starch for desired food applications.

Phospholipase D toxins from spiders, bacteria, and fungi generate cyclic phosphate products

Phospholipase D toxins from spiders, bacteria, and fungi generate cyclic phosphate products

Discovery of 1-((2R,4aR,6R,7R,7aR)-2-Isopropoxy-2-oxidodihydro-4H,6H-spiro[furo[3,2-d][1,3,2]dioxaphosphinine-7,2'-oxetan]-6-yl)pyrimidine-2,4(1H,3H)-dione (JNJ-54257099), a 3'-5'-Cyclic Phosphate Ester Prodrug of 2'-Deoxy-2'-Spirooxetane Uridine Triphosphate Useful for HCV Inhibition.

JNJ-54257099 (9) is a novel cyclic phosphate ester derivative that belongs to the class of 2'-deoxy-2'-spirooxetane uridine nucleotide prodrugs which are known as inhibitors of the HCV NS5B RNA-dependent RNA polymerase (RdRp). In the Huh-7 HCV genotype (GT) 1b replicon-containing cell line 9 is devoid of any anti-HCV activity, an observation attributable to inefficient prodrug metabolism which was found to be CYP3A4-dependent. In contrast, in vitro incubation of 9 in primary human hepatocytes as well as pharmacokinetic evaluation thereof in different preclinical species reveals the formation of substantial levels of 2'-deoxy-2'-spirooxetane uridine triphosphate (8), a potent inhibitor of the HCV NS5B polymerase. Overall, it was found that 9 displays a superior profile compared to its phosphoramidate prodrug analogues (e.g., 4) described previously. Of particular interest is the in vivo dose dependent reduction of HCV RNA observed in HCV infected (GT1a and GT3a) human hepatocyte chimeric mice after 7 days of oral administration of 9.

Polyphosphate is involved in cell cycle progression and genomic stability in Saccharomyces cerevisiae.

Polyphosphate (polyP) is a linear chain of up to hundreds of inorganic phosphate residues that is necessary for many physiological functions in all living organisms. In some bacteria, polyP supplies material to molecules such as DNA, thus playing an important role in biosynthetic processes in prokaryotes. In the present study, we set out to gain further insight into the role of polyP in eukaryotic cells. We observed that polyP amounts are cyclically regulated in Saccharomyces cerevisiae, and those mutants that cannot synthesise (vtc4Δ) or hydrolyse polyP (ppn1Δ, ppx1Δ) present impaired cell cycle progression. Further analysis revealed that polyP mutants show delayed nucleotide production and increased genomic instability. Based on these findings, we concluded that polyP not only maintains intracellular phosphate concentrations in response to fluctuations in extracellular phosphate levels, but also muffles internal cyclic phosphate fluctuations, such as those produced by the sudden demand of phosphate to synthetize deoxynucleotides just before and during DNA duplication. We propose that the presence of polyP in eukaryotic cells is required for the timely and accurate duplication of DNA.

Anionic flow polymerizations toward functional polyphosphoesters in microreactors: Polymerization and UV-modification

The polymerization of cyclic phosphates to poly(phosphoester)s, PPEs, is optimized for chip-based microreactors under continuous flow conditions. The anionic ring-opening polymerization of 2-isobutyoxy-2-oxo-1,3,2-dioxaphospholane (iBP) via the use of two organocatalytic systems allowed to polymerize to nearly quantitative monomer conversion within 10 or 3min, respectively at a reaction temperature of 40°C. Further, the optimized polymerization protocol was applied to 2-butenoxy-2-oxo-1,3,2-dioxaphospholane (BP) which yields a polymer that carries an alkene functionality per monomer repeating unit. This material can be postmodified in an UV-induced radical thiol–ene reaction, which was also shown to proceed with very high efficiency under UV-flow conditions. Eventually, both reactions were coupled in a two-stage reactor setup, showing that the thermally-activated polymerization can be coupled with high efficiency to the UV-activated post-polymerization modification reaction. The introduced reactor setup can in the future be used to produce and screen a broad variety of functional PPE materials with various functionalities and physical properties.

Selective amplification and sequencing of cyclic phosphate-containing RNAs by the cP-RNA-seq method.

RNA digestions catalyzed by many ribonucleases generate RNA fragments that contain a 2',3'-cyclic phosphate (cP) at their 3' termini. However, standard RNA-seq methods are unable to accurately capture cP-containing RNAs because the cP inhibits the adapter ligation reaction. We recently developed a method named cP-RNA-seq that is able to selectively amplify and sequence cP-containing RNAs. Here we describe the cP-RNA-seq protocol in which the 3' termini of all RNAs, except those containing a cP, are cleaved through a periodate treatment after phosphatase treatment; hence, subsequent adapter ligation and cDNA amplification steps are exclusively applied to cP-containing RNAs. cP-RNA-seq takes ∼6 d, excluding the time required for sequencing and bioinformatics analyses, which are not covered in detail in this protocol. Biochemical validation of the existence of cP in the identified RNAs takes ∼3 d. Even though the cP-RNA-seq method was developed to identify angiogenin-generating 5'-tRNA halves as a proof of principle, the method should be applicable to global identification of cP-containing RNA repertoires in various transcriptomes.

Distinct Contributions of Enzymic Functional Groups to the 2',3'-Cyclic Phosphodiesterase, 3'-Phosphate Guanylylation, and 3'-ppG/5'-OH Ligation Steps of the Escherichia coli RtcB Nucleic Acid Splicing Pathway.

Escherichia coli RtcB is a founding member of a family of manganese-dependent RNA repair enzymes that join RNA 2′,3′-cyclic phosphate (RNA>p) or RNA 3′-phosphate (RNAp) ends to 5′-OH RNA (HORNA) ends in a multistep pathway whereby RtcB (i) hydrolyzes RNA>p to RNAp, (ii) transfers GMP from GTP to RNAp to form to RNAppG, and (iii) directs the attack of 5′-OH on RNAppG to form a 3′-5′ phosphodiester splice junction. The crystal structure of the homologous archaeal RtcB enzyme revealed an active site with two closely spaced manganese ions, Mn1 and Mn2, that interact with the GTP phosphates. By studying the reactions of wild-type E. coli RtcB and RtcB alanine mutants with 3′-phosphate-, 2′,3′-cyclic phosphate-, and 3′-ppG-terminated substrates, we found that enzymic constituents of the two metal coordination complexes (Cys78, His185, and His281 for Mn1 and Asp75, Cys78, and His168 for Mn2 in E. coli RtcB) play distinct catalytic roles. For example, whereas the C78A mutation abolished all steps assayed, the D75A mutation allowed cyclic phosphodiester hydrolysis but crippled 3′-phosphate guanylylation, and the H281A mutant was impaired in overall HORNAp and HORNA>p ligation but was able to seal a preguanylylated substrate. The archaeal counterpart of E. coli RtcB Arg189 coordinates a sulfate anion construed to mimic the position of an RNA phosphate. We propose that Arg189 coordinates a phosphodiester at the 5′-OH end, based on our findings that the R189A mutation slowed the step of RNAppG/HORNA sealing by a factor of 200 compared to that with wild-type RtcB while decreasing the rate of RNAppG formation by only 3-fold. RtcB enzymes comprise a widely distributed family of manganese- and GTP-dependent RNA repair enzymes that ligate 2′,3′-cyclic phosphate ends to 5′-OH ends via RNA 3′-phosphate and RNA(3′)pp(5′)G intermediates. The RtcB active site includes two adjacent manganese ions that engage the GTP phosphates. Alanine scanning of Escherichia coli RtcB reveals distinct contributions of metal-binding residues Cys78, Asp75, and His281 at different steps of the RtcB pathway. The RNA contacts of RtcB are uncharted. Mutagenesis implicates Arg189 in engaging the 5′-OH RNA end.

Does Bimodal Agonism in Cyclic Nucleotide-Gated (CNG) Channels Preserve the Classical Binding Site and Pore Structure?

Binding of cGMP or cAMP to cyclic nucleotide-gated (CNG) channels typically increases open probability, and negative agonism (ligand-dependent suppression of opening) is rarely reported. The catfish CNGA2 subtype exhibits an unusual “bimodal agonism”: steady-state open probability increases with initial cGMP binding events (“pro” action) but decreases with subsequent cGMP binding events (“con” action). Con action is specific for cGMP and does not occur for cAMP. We investigated whether con action could be explained without postulating a non-canonical secondary binding site or an atypical non-conductive pore structure. We formulated a simple allosteric model postulating a switch in binding site affinity and coupling behaviour triggered by a critical ligand occupancy, and applied this model to three derivatives of catfish CNGA2 differing only in CNB fold sequence and exhibiting different maximum open probabilities. Dose-response relations of all three channels for cGMP (bimodal) and cAMP (pro action only) were well-explained by global model-fitting with shared closed-state dissociation constants, consistent with conservation of state-independent cyclic phosphate contacts. The effect of a mutation (V457S) outside the canonical binding site was well explained as enhanced pro action coupling and weakened con action coupling; an alternative model where V457S left con action coupling unchanged but affected only binding (a non-canonical site) produced inferior fits to data. The closed state induced by con action exhibited strong binding affinity for the pore-blocker tetracaine, resembling the classical closed channel formed during pro action at low open probabilities. Our findings support a model where V457 indirectly influences the orientation of ligand contacts in the canonical binding site; con action is produced when destabilization of cGMP contacts unique to the high-occupancy open state reverses the pore conformational change associated with pro action gate-opening.

Reversible chair ↔ skew conformational interconversion of 1,3,2-dioxaphosphorinane ring in the solid state.

β-NH4[(MeO)2cDHAP] (where cDHAP is cyclic dihydroxyacetone phosphate) has been investigated by X-ray crystallography in the temperature range 350-100 K. Three reversible single-crystal-to-single-crystal phase transitions have been examined and four phases (high-, room-, medium- and low-temperature phase, HTP, RTP, MTP and LTP, respectively) have been structurally determined: HTP (at 350 K, P2(1)/a, Z = 24), RTP (290 K, P1, Z = 12), MTP (205 K, P2(1)/a, Z = 4) and LTP (100 K, P1, Z = 24). Various puckering modes of the 1,3,2-dioxaphosphorinane ring of the organic cyclic phosphate anion have been revealed in the crystal: chair and skew. The chair ↔ skew ring flipping, which has been shown to take place during the phase transitions, results in various populations of skew and chair conformers in different phases. The flexibility in the coordination geometry of ammonium cations is considered to assist in the conformational flexibility of the dioxaphosphorinane ring.

Micro-intumescent flame retardant polyamide 6 based on cyclic phosphate grafting phenol formaldehyde

In this paper, thermoplastic phenol formaldehyde (PF) grafted cyclic neopentyl phosphate (PFCP) was synthesized by using PF and 2,2-dimethyl-1,3-propanediol phosphoryl chloride. It was characterized by Fourier transform infrared spectroscopy (FTIR), 1H and 31P nuclear magnetic resonance (NMR). Compared to PF, PFCP shows improved thermal and thermoxidative stability and allows itself to be used in polyamide 6 (PA6). A micro-intumescent flame retardant system was constructed by using cyclic neopentyl phosphate as acid source, PF as charring agent and PA6 whose decomposition products work as blowing agent. The results showed that PA6/PFCP composite is classified the UL-94V-0 rating and get a LOI value of 35.5% at 25% loading of PFCP. SEM results showed that the outside of char residues is continuous and dense, but the inside is micro-intumescent and porous. XPS analysis of char revealed that most of phosphorus remained in the char layer. All the results suggest that the mode of flame retardant's action for PA6/PFCP composites is shifted from melting away to charring protection with the content of PFCP increasing. The coherent char generated by the decomposition of PFCP contributes most to flame retardancy of PA6. Copyright (c) 2016 John Wiley & Sons, Ltd.

3':5'-Cyclic nucleotides: two sodium salts of cdTMP.

3':5'-Cyclic nucleotides play an outstanding role in signal transduction at the cellular level but, in spite of comprehensive knowledge of the biological role of cyclic nucleotides, their structures are not established fully. Two hydrated sodium salts of thymidine 3':5'-cyclic phosphate (cdTMP, C10H12N2O7P), namely sodium thymidine 3':5'-cyclic phosphate heptahydrate, Na(+)·C10H12N2O7P(-)·7H2O or Na(cdTMP)·7H2O, (I), and sodium thymidine 3':5'-cyclic phosphate 3.7-hydrate, Na(+)·C10H12N2O7P(-)·3.7H2O or Na(cdTMP)·3.7H2O, (II), have been obtained in crystalline form and structurally characterized, revealing one nucleotide in the asymmetric unit of (I) and eight different nucleotides in (II). All the cyclic nucleotide anions adopt a similar conformation with regard to nucleobase orientation, sugar conformation and 1,3,2-dioxaphosphorinane ring puckering. In (I), no direct inter-nucleotide hydrogen bonds are present, and adjacent nucleotide anions interact via water-mediated and Na(+)-mediated contacts. In contrast, in (II), direct thymine-phosphate N-H...O inter-nucleotide hydrogen bonds occur and these are assisted by numerous inter-nucleotide C-H...O contacts, giving rise to the self-assembly of cdTMP(-) anions into three different ribbons. Two of these three ribbons run in the same direction, while the third is antiparallel.

The Role of Metals in the Reaction Catalyzed by Metal-Ion-Independent Bacillary RNase.

Extracellular enzymes of intestinal microbiota are the key agents that affect functional activity of the body as they directly interact with epithelial and immune cells. Several species of the Bacillus genus, like Bacillus pumilus, a common producer of extracellular RNase binase, can populate the intestinal microbiome as a colonizing organism. Without involving metal ions as cofactors, binase depolymerizes RNA by cleaving the 3′,5′-phosphodiester bond and generates 2′,3′-cyclic guanosine phosphates in the first stage of a catalytic reaction. Maintained in the reaction mixture for more than one hour, such messengers can affect the human intestinal microflora and the human body. In the present study, we found that the rate of 2′,3′-cGMP was growing in the presence of transition metals that stabilized the RNA structure. At the same time, transition metal ions only marginally reduced the amount of 2′,3′-cGMP, blocking binase recognition sites of guanine at N7 of nucleophilic purine bases.

Encapsulation of a hydrophobic drug in polyphosphate-based nanocarriers

Event Abstract Back to Event Encapsulation of a hydrophobic drug in polyphosphate-based nanocarriers Stéphanie Vanslambrouck1, Benoît Clément1, Raphaël Riva1, Bernard Ucakar2, Véronique Préat2, Mick Gagliardi3, Daniel G. Molin3, Guy Broze1, Leo H. Koole4, Philippe Lecomte1 and Christine Jérôme1 1 University of Liege, Center for Education and Research on Macromolecules, Belgium 2 Université catholique de Louvain, Louvain Drug Research Institute, Pharmaceutics and Drug Delivery, Belgium 3 Maastricht University, Faculty of Health, Medicine and Life Science, Department of Physiology, Netherlands 4 Maastricht University, Faculty of Health, Medicine and Life Science, Department of Biomedical Engineering/Biomaterials Sci., Netherlands Thanks to their biocompatibility and degradability properties, polyphosphates are appealing polymers for biomedical applications, especially for drug delivery systems. In contrast to aliphatic polyesters, such as poly(ε-caprolactone) and poly(lactide), the pentavalency of phosphorus atom allows the easy modification of the polyphosphate properties by simply adjusting the nature, the length and the functionality of the polyphosphate pendant groups. Poly(ethylene glycol) (PEG)-b-polyphosphate block copolymers made of hydrophilic PEG and hydrophobic polyphosphates are amphiphilic copolymers prone to self-assemble in aqueous medium into micelles. In order to enhance the encapsulation of hydrophobic drugs, a series of amphiphilic PEG-b-polyphosphate copolymers were synthesized by organocatalyzed ring-opening polymerization of cyclic phosphates and, in some case, followed by thiol-ene click post-polymerization reactions between the pendant vinyl group of a preformed polyphosphate block and some thiols (i.e. dodecyl thiol and tocopherol thiol). Our work aims at changing the nature of the pendant groups of the polyphosphate block to investigate the influence of this structural modification (i) on the size of the micelles, (ii) on the critical aggregation concentration (CAC), (iii) on the loading of an hydrophobic model drug (i.e. ketoconazole) and, finally, (iv) on the release of this drug. As biomedical applications are foreseen, the cytotoxicity of these PEG-b-polyphosphate copolymers were also evaluated by live/dead cell viability assays. BioMIMedics, Interreg EMR IV-A Consortium Keywords: Drug delivery, polymer, Biodegradable material Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: Poster Topic: Self-assembling micellar systems Citation: Vanslambrouck S, Clément B, Riva R, Ucakar B, Préat V, Gagliardi M, Molin DG, Broze G, Koole LH, Lecomte P and Jérôme C (2016). Encapsulation of a hydrophobic drug in polyphosphate-based nanocarriers. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.00879 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Mar 2016; Published Online: 30 Mar 2016. Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Stéphanie Vanslambrouck Benoît Clément Raphaël Riva Bernard Ucakar Véronique Préat Mick Gagliardi Daniel G Molin Guy Broze Leo H Koole Philippe Lecomte Christine Jérôme Google Stéphanie Vanslambrouck Benoît Clément Raphaël Riva Bernard Ucakar Véronique Préat Mick Gagliardi Daniel G Molin Guy Broze Leo H Koole Philippe Lecomte Christine Jérôme Google Scholar Stéphanie Vanslambrouck Benoît Clément Raphaël Riva Bernard Ucakar Véronique Préat Mick Gagliardi Daniel G Molin Guy Broze Leo H Koole Philippe Lecomte Christine Jérôme PubMed Stéphanie Vanslambrouck Benoît Clément Raphaël Riva Bernard Ucakar Véronique Préat Mick Gagliardi Daniel G Molin Guy Broze Leo H Koole Philippe Lecomte Christine Jérôme Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Observation on the efficacy of simvastatin combined with cyclic adenosine phosphate in the treatment of chronic heart failure in patients with coronary heart disease

Objective To explore the clinical effect of simvastatin combined with cyclic adenosine phosphate in the treatment of chronic heart failure in patients with coronary heart disease. Methods 100 patients with chronic heart failure who received treatment during July 2013 and June 2015 were randomly divided into control group (50 cases) and observation group (50 cases).The control group was treated with conventional therapy, and the observation group was treated with simvastatin combined with cyclic adenosine. Results The total effective rate of the observation group was 81.0%, which of the control group was 70.0%, the total effective rate of the two groups had no obvious difference(χ2=3.060 6, P>0.05).The LVEDD, LVESD in the two groups significantly decreased compared with before treatment, LVEF significantly increased after treatment(P<0.05).After treatment, the LVESD, LVEDD of observation group were (36.1±3.9)mm, (45.0±10.4)mm, which were significantly lower than (41.3±4.2)mm, (58.0±10.6)mm of the control group.The LVEF of observation group was (55.0±9.2)%, which was significantly higher than (38.0±7.5)% of the control group (t=6.123 7, 7.114 9, 6.325 7, all P<0.05). Conclusion Simvastatin combined with adenosine cyclophosphate for the treatment of chronic heart failure of coronary heart disease has good treatment effect, can effectively improve the prognosis of patients with cardiac function to improve patients quality of life. Key words: Simvastatin; Adenosine cyclophosphate; Coronary artery disease; Heart failure

Coordinated Ribosomal ITS2 RNA Processing by the Las1 Complex Integrating Endonuclease, Polynucleotide Kinase, and Exonuclease Activities

The rapidly evolving internal transcribed spacer 2 (ITS2) in the pre-ribosomal RNA is one of the most commonly applied phylogenetic markers at species and genus level. Yet, during ribosome biogenesis ITS2 is removed in all eukaryotes by a common, but still unknown, mechanism. Here we describe the existence of an RNA processome, assembled from four conserved subunits, Las1-Grc3-Rat1-Rai1, that carries all the necessary RNA processing enzymes to mediate coordinated ITS2 rRNA removal. Las1 is the long-sought-after endonuclease cleaving 27SB pre-rRNA at site C2 to yield a 5'-OH end at the 26S pre-rRNA and 2',3' cyclic phosphate at the 3' end of 7S pre-rRNA. Subsequently, polynucleotide kinase Grc3 catalyzes ATP-dependent 5'-OH phosphorylation of 26S pre-rRNA, which in turn enables Rat1-Rai1 exonuclease to generate 25S' pre-rRNA. ITS2 processing is reminiscent of tRNA splicing, but instead of subsequent tRNA ligation, the Las1 complex carries along an exonuclease tool to degrade the ITS2 rRNA.

Mechanistic Investigation of cPMP Synthase in Molybdenum Cofactor Biosynthesis Using an Uncleavable Substrate Analogue.

Molybdenum cofactor (Moco) is essential for all kingdoms of life, plays central roles in various biological processes, and must be biosynthesized de novo. During its biosynthesis, the characteristic pyranopterin ring is constructed by a complex rearrangement of guanosine 5'-triphosphate (GTP) into cyclic pyranopterin monophosphate (cPMP) through the action of two enzymes, MoaA and MoaC. Recent studies revealed that MoaC catalyzes the majority of the transformation and produces cPMP from a unique cyclic nucleotide, 3',8-cyclo-7,8-dihydro-GTP (3',8-cH2GTP). However, the mechanism by which MoaC catalyzes this complex rearrangement is largely unexplored. Here, we report the mechanistic characterization of MoaC using an uncleavable substrate analogue, 3',8-cH2GMP[CH2]PP, as a probe to investigate the timing of cyclic phosphate formation. Using partially active MoaC variants, 3',8-cH2GMP[CH2]PP was found to be accepted by MoaC as a substrate and was converted to an analogue of the previously described MoaC reaction intermediate, suggesting that the early stage of catalysis proceeds without cyclic phosphate formation. In contrast, when it was incubated with wt-MoaC, 3',8-cH2GMP[CH2]PP caused mechanism-based inhibition. Detailed characterization of the inhibited MoaC suggested that 3',8-cH2GMP[CH2]PP is mainly converted to a molecule (compound Y) with an acid-labile triaminopyrimidinone base without an established pyranopterin structure. MS analysis of MoaC treated with 3',8-cH2GMP[CH2]PP provided strong evidence that compound Y forms a tight complex with MoaC likely through a covalent linkage. These observations are consistent with a mechanism in which cyclic phosphate ring formation proceeds in concert with the pterin ring formation. This mechanism would provide a thermodynamic driving force to complete the formation of the unique tetracyclic structure of cPMP.

Irisin relaxes mouse mesenteric arteries through endothelium-dependent and endothelium-independent mechanisms

Irisin, a newly discovered myokine, has been shown to produce modest weight loss and improve glucose intolerance in mice. The purpose of this study was to investigate the effects of irisin on vascular activity and the mechanisms involved. Experiments were performed on mouse mesenteric arteries. We demonstrated that irisin induced relaxation in mesenteric arteries with or without endothelium in a concentration-dependent manner. It was further demonstrated that the irisin-induced vasorelaxation effects on endothelium-intact mesenteric arteries were reduced by pretreatment with Nω-nitro-l-arginine methyl ester (l-NAME) or 1H-[1, 2, 4] oxadizolo [4, 3-a] quinoxalin-1-one (ODQ). However, pretreatment with indomethacin (INDO), a nonselective cyclooxygenase inhibitor did not modulate irisin-induced relaxation. In addition, the contraction due to extracellular Ca2+ influx and intracellular Ca2+ release was also inhibited by irisin. In summary, these results suggested that the endothelium-dependent relaxation of irisin is mediated by the nitric oxide (NO)-guanosine 3′, 5′-cyclic phosphate (cGMP)-dependent pathway but not the prostaglandin I2 (PGI2)-cyclic adenosine monophosphate (cAMP)-dependent mechanism. Endothelium-independent relaxation may be depend on inhibiting Ca2+ influx through blocking VDCCs and intracellular Ca2+ release through both IP3R and RyR channels.