The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Thomas Preiss - One of the best experts on this subject based on the ideXlab platform.
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selective translation complex profiling reveals staged Initiation and co translational assembly of Initiation factor complexes
Molecular Cell, 2020Co-Authors: Susan Wagner, Thomas Preiss, Nikolay E. Shirokikh, Anna Herrmannova, Vladislava Hronova, Stanislava Gunisova, Neelam Dabas Sen, Ross D Hannan, Alan G HinnebuschAbstract:Summary Translational control targeting the Initiation Phase is central to the regulation of gene expression. Understanding all of its aspects requires substantial technological advancements. Here we modified yeast translation complex profile sequencing (TCP-seq), related to ribosome profiling, and adapted it for mammalian cells. Human TCP-seq, capable of capturing footprints of 40S subunits (40Ss) in addition to 80S ribosomes (80Ss), revealed that mammalian and yeast 40Ss distribute similarly across 5′TRs, indicating considerable evolutionary conservation. We further developed yeast and human selective TCP-seq (Sel-TCP-seq), enabling selection of 40Ss and 80Ss associated with immuno-targeted factors. Sel-TCP-seq demonstrated that eIF2 and eIF3 travel along 5′ UTRs with scanning 40Ss to successively dissociate upon AUG recognition; notably, a proportion of eIF3 lingers on during the initial elongation cycles. Highlighting Sel-TCP-seq versatility, we also identified four initiating 48S conformational intermediates, provided novel insights into ATF4 and GCN4 mRNA translational control, and demonstrated co-translational assembly of Initiation factor complexes.
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selective translation complex profiling reveals staged Initiation and co translational assembly of Initiation factor complexes
bioRxiv, 2019Co-Authors: Susan Wagner, Thomas Preiss, Nikolay E. Shirokikh, Anna Herrmannova, Vladislava Hronova, Neelam Dabas Sen, Ross D Hannan, Alan G Hinnebusch, Leos Shivaya ValasekAbstract:SUMMARY Translational control targeting mainly the Initiation Phase is central to the regulation of gene expression. Understanding all of its aspects requires substantial technological advancements. Here we modified yeast Translational Complex Profile sequencing (TCP-seq), related to ribosome profiling, and adopted it for mammalian cells. Human TCP-seq, capable of capturing footprints of 40S subunits (40Ses) in addition to 80S ribosomes (80Ses), revealed that mammalian and yeast 40Ses distribute similarly across 5’UTRs indicating considerable evolutionary conservation. We further developed a variation called Selective TCP-seq (Sel-TCP-seq) enabling selection for 40Ses and 80Ses associated with an immuno-targeted factor in yeast and human. Sel-TCP-seq demonstrated that eIF2 and eIF3 travel along 5’UTRs with scanning 40Ses to successively dissociate upon start codon recognition. Manifesting the Sel-TCP-seq versatility for gene expression studies, we also identified four initiating 48S conformational intermediates, provided novel insights into ATF4 and GCN4 mRNA translational control, and demonstrated co-translational assembly of Initiation factor complexes.
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Control of Translation at the Initiation Phase During Glucose Starvation in Yeast
International Journal of Molecular Sciences, 2019Co-Authors: Yoshika Janapala, Thomas Preiss, Nikolay E. ShirokikhAbstract:Glucose is one of the most important sources of carbon across all life. Glucose starvation is a key stress relevant to all eukaryotic cells. Glucose starvation responses have important implications in diseases, such as diabetes and cancer. In yeast, glucose starvation causes rapid and dramatic effects on the synthesis of proteins (mRNA translation). Response to glucose deficiency targets the Initiation Phase of translation by different mechanisms and with diverse dynamics. Concomitantly, translationally repressed mRNAs and components of the protein synthesis machinery may enter a variety of cytoplasmic foci, which also form with variable kinetics and may store or degrade mRNA. Much progress has been made in understanding these processes in the last decade, including with the use of high-throughput/omics methods of RNA and RNA:protein detection. This review dissects the current knowledge of yeast reactions to glucose starvation systematized by the stage of translation Initiation, with the focus on rapid responses. We provide parallels to mechanisms found in higher eukaryotes, such as metazoans, for the most critical responses, and point out major remaining gaps in knowledge and possible future directions of research on translational responses to glucose starvation.
Kenneth G Mann - One of the best experts on this subject based on the ideXlab platform.
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the function of factor xi in tissue factor initiated thrombin generation
Journal of Thrombosis and Haemostasis, 2003Co-Authors: Saulius Butenas, J D Dee, Kenneth G MannAbstract:Summary. The influence of plasma and platelet factor (F)XI on thrombin generation initiated with 10 pm tissue factor (TF) in a synthetic coagulation model was evaluated in the presence of either 2 × 108 mL−1 platelets or the equivalent (2 µm) phospholipids. In either system, with all proteins present at physiological concentrations, FXI (30 nm) had no effect on thrombin generation. With phospholipids in the absence of FXI, an increase in vitamin K-dependent proteins (VKDP) (up to 500%) significantly prolonged the Initiation Phase of thrombin generation and decreased maximum thrombin levels. The inhibition was principally caused by the elevated prothrombin and FIX concentrations. When 30 nm FXI was added with elevated VKDP and phospholipids, the Initiation Phase was decreased and the maximum thrombin levels generated substantially increased. In experiments with platelets (with and without plasma FXI), an increase in VKDP had little effect on the Initiation Phase of thrombin generation. These data indicate that (i) FXI has no effect on thrombin generation at 10 pm TF and physiological concentrations of VKDP; (ii) platelets and plasma FXI are able to compensate for the inhibitory effects of elevated VKDP.
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thrombin functions during tissue factor induced blood coagulation
Blood, 2002Co-Authors: Kathleen E Brummel, Saulius Butenas, Sara G Paradis, Kenneth G MannAbstract:Tissue factor–induced blood coagulation was studied in 20 individuals, for varying periods of time during 54 months, in contact pathway–inhibited whole blood at 37°C and evaluated in terms of the activation of various substrates. After quenching over time with inhibitors, the soluble Phases were analyzed for thrombin–antithrombin III (TAT) complex formation, prothrombin fragments, platelet activation (osteonectin release), factor Va generation, fibrinopeptide (FP) A and FPB release, and factor XIII activation. TAT complex formation, for 35 experiments, showed an Initiation Phase (up to 4.6 ± 0.6 minutes) in which thrombin was generated at an average rate of 0.93 ± 0.3 nM/min catalyzed by about 1.3 pM prothrombinase yielding approximately 26 nM thrombin. During a subsequent propagation Phase, thrombin was generated at a rate of 83.9 ± 3.8 nM/min by about 120 pM prothrombinase, reaching ultimate levels of 851 ± 53 nM. Clot time, determined subjectively, occurred at 4.7 ± 0.2 minutes and correlated with the inception of the propagation Phase. The thrombin concentrations associated with the transitions to rapid product formation are 510 ± 180 pM for platelet activation (1.9 ± 0.2 minutes), 840 ± 280 pM for factor XIII activation and factor Va generation (2.2 ± 0.6 minutes), 1.3 ± 0.4 nM for FPA release (2.5 ± 0.7 minutes), 1.7 ± 0.5 nM for FPB release and prethrombin 2 (2.8 ± 0.8 minutes), 7.0 ± 2.2 nM for thrombin B chain (3.6 ± 0.2 minutes), and 26 ± 6.2 nM for the propagation Phase of TAT formation (4.6 ± 0.6 minutes). These results illustrate that the initial activation of thrombin substrates occurs during the Initiation Phase at less than 2 nM thrombin (0.2%). Most thrombin (96%) is formed well after clotting occurs.
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thrombin functions during tissue factor induced blood coagulation
Blood, 2002Co-Authors: Kathleen E Brummel, Saulius Butenas, Sara G Paradis, Kenneth G MannAbstract:Tissue factor-induced blood coagulation was studied in 20 individuals, for varying periods of time during 54 months, in contact pathway-inhibited whole blood at 37 degrees C and evaluated in terms of the activation of various substrates. After quenching over time with inhibitors, the soluble Phases were analyzed for thrombin-antithrombin III (TAT) complex formation, prothrombin fragments, platelet activation (osteonectin release), factor Va generation, fibrinopeptide (FP) A and FPB release, and factor XIII activation. TAT complex formation, for 35 experiments, showed an Initiation Phase (up to 4.6 +/- 0.6 minutes) in which thrombin was generated at an average rate of 0.93 +/- 0.3 nM/min catalyzed by about 1.3 pM prothrombinase yielding approximately 26 nM thrombin. During a subsequent propagation Phase, thrombin was generated at a rate of 83.9 +/- 3.8 nM/min by about 120 pM prothrombinase, reaching ultimate levels of 851 +/- 53 nM. Clot time, determined subjectively, occurred at 4.7 +/- 0.2 minutes and correlated with the inception of the propagation Phase. The thrombin concentrations associated with the transitions to rapid product formation are 510 +/- 180 pM for platelet activation (1.9 +/- 0.2 minutes), 840 +/- 280 pM for factor XIII activation and factor Va generation (2.2 +/- 0.6 minutes), 1.3 +/- 0.4 nM for FPA release (2.5 +/- 0.7 minutes), 1.7 +/- 0.5 nM for FPB release and prethrombin 2 (2.8 +/- 0.8 minutes), 7.0 +/- 2.2 nM for thrombin B chain (3.6 +/- 0.2 minutes), and 26 +/- 6.2 nM for the propagation Phase of TAT formation (4.6 +/- 0.6 minutes). These results illustrate that the initial activation of thrombin substrates occurs during the Initiation Phase at less than 2 nM thrombin (0.2%). Most thrombin (96%) is formed well after clotting occurs.
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evaluation of the Initiation Phase of blood coagulation using ultrasensitive assays for serine proteases
Journal of Biological Chemistry, 1997Co-Authors: Saulius Butenas, C Van T Veer, Kenneth G MannAbstract:Abstract The Initiation Phase of enzyme generation in a reconstituted model of the tissue factor (TF) pathway to thrombin was evaluated. At 1.25 pm added TF, no thrombin generation was observed in the absence of factor V. The substitution of factor Va for factor V increased the rate of thrombin generation. Factor X activation during the Initiation Phase was not influenced by the absence of factor VIII or thrombin, leading to the conclusion that initially factor Xa is generated exclusively by the factor VIIa-TF complex. When thrombin was eliminated from the system, no contribution of the factor IXa-factor VIIIa complex to factor X activation was observed during the propagation Phase. Similarly, factor V activation was also not observed in the absence of thrombin, indicating that thrombin is the only enzyme responsible for factor V and factor VIII activation. Only subnanomolar amounts of factor VII were activated when prothrombin activation was almost complete. In the absence of coagulation inhibitors, factor XI did not influence thrombin generation initiated by 1.25 pm factor VIIa-TF complex. The termination of factor XIa generation by added hirudin in the factor XI experiment indicates that factor XI activation occurs exclusively by thrombin.
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regulation of tissue factor initiated thrombin generation by the stoichiometric inhibitors tissue factor pathway inhibitor antithrombin iii and heparin cofactor ii
Journal of Biological Chemistry, 1997Co-Authors: Cornelis Van T Veer, Kenneth G MannAbstract:Abstract The effects of the stoichiometric inhibitors tissue factor pathway inhibitor (TFPI), antithrombin-III (AT-III) and heparin cofactor-II (HC-II) on thrombin generation were evaluated in a reaction system composed of coagulation factors VIIa, X, IX, VIII, and V and prothrombin initiated by tissue factor (TF) and phospholipids. Initiation of the reaction in the absence of inhibitors resulted in explosive thrombin generation for factor VIIa·TF concentrations varying from 100 to 0.25 pM with the lag time or Initiation Phase of thrombin generation increasing from 0 to 180 s with decreasing factor VIIa·TF concentrations. During the propagation Phase, prothrombin is quantitatively activated to 1.4 μM α-thrombin. At normal plasma concentration (2.5 nM) full-length recombinant TFPI prolonged the Initiation Phase of thrombin generation 2-fold, and the rate of thrombin generation in the propagation Phase of the reaction was 25-50% that of the uninhibited reaction when the reaction was initiated with 1.25-20 pM factor VIIa·TF. Inhibition of the reaction by TFPI is associated with a delay in factor V activation. In the presence of TFPI no explosive thrombin generation was observed when factor VIII was omitted from reactions initiated by factor VIIa·TF concentrations ≤20 pM. This indicates that in the presence of TFPI the factor IXa·factor VIIIa pathway becomes essential at low factor VIIa·TF concentrations. In the reconstituted system, AT-III (3.4 μM) did not prolong the Initiation Phase of thrombin generation when the reaction was initiated with 1.25 pM factor VIIa·TF, nor did AT-III delay factor V activation. The rate of thrombin formation in the presence of AT-III was reduced to 30% that of the uninhibited reaction, and the α-thrombin formed was rapidly inhibited subsequent to its generation. The addition of HC-II alone at its physiological concentration (1.38 μM) to the procoagulant mixture did not alter the rate or extent of thrombin generation. Subsequently, the thrombin formed was slowly inhibited by HC-II. The slow inactivation of thrombin by HC-II does not contribute to thrombin inhibition in the presence of AT-III. In contrast, the combination of physiological levels of AT-III and TFPI inhibited explosive thrombin generation initiated by 1.25 pM factor VIIa·TF completely. The absence of prothrombin consumption indicated that the combination of TFPI and AT-III is able to prevent the formation of prothrombinase activity at low factor VIIa·TF concentrations. The data indicate that TFPI potentiates the action of AT-III by decreasing the rate of formation and thus the amount of catalyst formed in the reaction, enabling AT-III to effectively scavenge the limited traces of factor IXa and factor Xa formed in the presence of TFPI. The Initiation of thrombin generation by increasing factor VIIa·TF concentrations in the presence of physiological concentrations of TFPI and AT-III showed dramatic changes in the maximal rates of thrombin generation over small changes in initiator concentration. These data demonstrate that significant thrombin generation becomes a “threshold-limited” event with regard to the initiating factor VIIa·TF concentration in the presence of TFPI and AT-III.
Karam Elbayoumy - One of the best experts on this subject based on the ideXlab platform.
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effects of 1 4 phenylenebis methylene selenocyanate and selenomethionine on 4 methylnitrosamino 1 3 pyridyl 1 butanone induced tumorigenesis in a j mouse lung
Carcinogenesis, 1997Co-Authors: Bogdan Prokopczyk, Pramod Upadhyaya, Shantu Amin, Dhimant Desai, Christine Kurtzke, Karam ElbayoumyAbstract:We reported earlier that continuous feeding of 1,4-phenylenebis(methylene)selenocyanate (p-XSC) inhibited lung tumor induction by the tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in the A/J mouse (El-Bayoumy et al., Carcinogenesis, 14, 1111-1113, 1993). The present investigation was designed to determine whether p-XSC inhibits pulmonary neoplasia induced by NNK in female A/J mice during the Initiation Phase of carcinogenesis or during the post-Initiation Phase. The naturally occurring selenomethionine was also included in this study. Doses higher than 4 p.p.m. of selenomethionine can induce toxic effects, therefore, dietary supplementation of this compound was selected at a dose level of 3.75 p.p.m. However, we were able to give p-XSC at selenium levels of 7.5 and 15 p.p.m., as mice can tolerate such doses in this form without any adverse effects. NNK was given by a single i.p. injection at dose of 10 micromol in 0.1 ml of saline. Selenomethionine did not show chemopreventive activity when administered in either Phase of tumorigenesis. In contrast, p-XSC significantly reduced lung tumor multiplicity regardless of whether it was given during the Initiation Phase of tumorigenesis (P = 0.0009 at both levels of selenium) or post-Initiation (P = 0.0009 at 15 p.p.m. and P = 0.036 for 7.5 p.p.m.). This is the first report describing that the synthetic organoselenium compound, p-XSC, can effectively block and suppress chemically (NNK)-induced lung tumor development in mice.
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chemoprevention of colon carcinogenesis by the synthetic organoselenium compound 1 4 phenylenebis methylene selenocyanate
Cancer Research, 1992Co-Authors: Bandaru S Reddy, Nalini Kulkarni, A Rivenson, Pramod Upadhyaya, Karam ElbayoumyAbstract:The chemopreventive effect of 40% and 80% maximum tolerated dose (MTD) levels of 1,4-phenylenebis(methylene)selenocyanate ( p -XSC) administered in the diet during the Initiation Phase (2 weeks before, during, and up to 3 days after carcinogen administration) and the post-Initiation Phase (3 days after carcinogen treatment until termination) of azoxymethane (AOM)-induced colon carcinogenesis was studied in male F344 rats. The MTD of p -XSC was determined in male F344 rats and found to be 50 ppm. Beginning at 5 weeks of age, all animals were divided into various experimental groups (42 rats/group) and fed the high-fat semipurified diet or diets containing 20 (40% MTD) and 40 (80% MTD) ppm p -XSC. At 7 weeks of age, all animals (30 rats/group) except the vehicle-treated groups (12 rats/group) were administered s.c. injections of AOM (15 mg/kg body weight/week for 2 weeks). Three days after the second injection of AOM or vehicle (normal saline), groups of animals fed the p -XSC diets and control diet were transferred, respectively, to control diet and p -XSC diets and continued on these diets until the termination of the study. All animals were necropsied during the 36th week after AOM treatment. Colonic mucosal prostaglandin E2 and selenium-dependent glutathione peroxidase were measured in animals fed the control and p -XSC diets at the termination of the study. The results indicate that 40 ppm p -XSC administered during the Initiation Phase significantly inhibited the colon tumor incidence (percentage of animals with tumors). Dietary p -XSC administered at 20 and 40 ppm levels during the Initiation Phase significantly inhibited colon tumor multiplicity (tumors/animal and tumors/tumor-bearing animal). Colon tumor incidence and multiplicity were significantly reduced in groups fed 20 and 40 ppm p -XSC diets at the postInitiation Phase of carcinogenesis. Colonic mucosal selenium-dependent glutathione peroxidase activity was increased, and prostaglandin E2 was reduced in animals fed the p -XSC diet compared to animals fed the control diet. Whereas the precise mechanisms of p -XSC-induced inhibition of colon carcinogenesis remain to be elucidated, it is likely that the effect during the Initiation and postInitiation Phases may be due to alteration in carcinogen metabolism and to modulation of prostaglandin synthesis and selenium-dependent glutathione peroxidase activity.
Shadia Rifai Habbal - One of the best experts on this subject based on the ideXlab platform.
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bridging euv and white light observations to inspect the Initiation Phase of a two stage solar eruptive event
Solar Physics, 2014Co-Authors: Jason P Byrne, Huw Morgan, Daniel B Seaton, H M Bain, Shadia Rifai HabbalAbstract:The Initiation Phase of coronal mass ejections (CMEs) is a very important aspect of solar physics, as these phenomena ultimately drive space weather in the heliosphere. This Phase is known to occur between the photosphere and low corona, where many models introduce an instability and/or magnetic reconnection that triggers a CME, often with associated flaring activity. To this end, it is important to obtain a variety of observations of the low corona to build as clear a picture as possible of the dynamics that occur therein. Here, we combine the EUV imagery of the Sun Watcher using Active Pixel System Detector and Image Processing (SWAP) instrument onboard the Project for Onboard Autonomy (PROBA2) with the white-light imagery of the ground-based Mark-IV K-coronameter (Mk4) at Mauna Loa Solar Observatory (MLSO) to bridge the observational gap that exists between the disk imagery of the Atmospheric Imaging Assembly (AIA) onboard the Solar Dynamics Observatory (SDO) and the coronal imagery of the Large Angle Spectrometric Coronagraph (LASCO) onboard the Solar and Heliospheric Observatory (SOHO). Methods of multiscale image analysis were applied to the observations to better reveal the coronal signal while suppressing noise and other features. This allowed an investigation into the Initiation Phase of a CME that was driven by a rising flux-rope structure from a “two-stage” flaring event underlying an extended helmet streamer. It was found that the initial outward motion of the erupting loop system in the EUV observations coincided with the first X-ray flare peak and led to a plasma pile-up of the white-light CME core material. The characterized CME core then underwent a strong jerk in its motion, as the early acceleration increased abruptly, simultaneously with the second X-ray flare peak. The overall system expanded into the helmet streamer to become the larger CME structure observed in the LASCO coronagraph images, which later became concave-outward in shape. Theoretical models for the event are discussed in light of these unique observations, and it is concluded that the formation of either a kink-unstable or torus-unstable flux rope may be the likeliest scenario.
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bridging euv and white light observations to inspect the Initiation Phase of a two stage solar eruptive event
arXiv: Solar and Stellar Astrophysics, 2014Co-Authors: Jason P Byrne, Huw Morgan, Daniel B Seaton, H M Bain, Shadia Rifai HabbalAbstract:The Initiation Phase of CMEs is a very important aspect of solar physics, as these phenomena ultimately drive space weather in the heliosphere. This Phase is known to occur between the photosphere and low corona, where many models introduce an instability and/or magnetic reconnection that triggers a CME, often with associated flaring activity. To this end, it is important to obtain a variety of observations of the low corona in order to build as clear a picture as possible of the dynamics that occur therein. Here, we combine the EUV imagery of the SWAP instrument on board PROBA2 with the white-light imagery of the ground-based Mk4 coronameter at MLSO in order to bridge the observational gap that exists between the disk imagery of AIA on board SDO and the coronal imagery of LASCO on board SOHO. Methods of multiscale image analysis were applied to the observations to better reveal the coronal signal while suppressing noise and other features. This allowed an investigation into the Initiation Phase of a CME that was driven by a rising flux rope structure from a "two-stage" flaring active region underlying an extended helmet streamer. It was found that the initial outward motion of the erupting loop system in the EUV observations coincided with the first X-ray flare peak, and led to a plasma pile-up of the white-light CME core material. The characterized CME core then underwent a strong jerk in its motion, as the early acceleration increased abruptly, simultaneous with the second X-ray flare peak. The overall system expanded into the helmet streamer to become the larger CME structure observed in the LASCO coronagraph images, which later became concave-outward in shape. Theoretical models for the event are discussed in light of these unique observations, and it is concluded that the formation of either a kink-unstable or torus-unstable flux rope may be the likeliest scenario.
Raghu Sinha - One of the best experts on this subject based on the ideXlab platform.
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Mechanisms of mammary cancer chemoprevention by organoselenium compounds.
Mutation research, 2004Co-Authors: Karam El-bayoumy, Raghu SinhaAbstract:Searching for optimal diets and for naturally occurring agents in routinely consumed foods that may inhibit cancer development, although challenging, constitutes a valuable and plausible approach to finding ways to control and prevent cancer. To date, the use of the micronutrient selenium in human clinical trials is limited but the outcome of these investigations indicates that selenium is one of the most promising agents. Data presented in this mini-review indicate that the dose and the form (structure) in which selenium is used are the most critical determinants of success in future clinical trials. The focus of this mini-review is on the mechanisms of mammary cancer chemoprevention by organoselenium compounds. Among the naturally occurring organoselenium compounds, Se-Methylselenocysteine is more efficacious than the most extensively studied forms, such as selenomethionine. However, we showed that synthetic organoselenium compounds can be tailored to achieve greater chemopreventive efficacy with minimal side effects by structural modifications; it is evident that synthetic agents are superior to the inorganic selenite, naturally occurring selenium compounds and their sulfur-containing analogs. We have demonstrated that 1,4-phenylenebis (methylene) selenocyanate (p-XSC) and its putative metabolite glutathione conjugate (p-XSeSG) are highly promising agents in the chemoprevention of mammary carcinogenesis in the 7,12-dimethylbenz[a]anthracene (DMBA)-rat mammary tumor model system. Both compounds inhibit the Initiation Phase of carcinogenesis by inhibiting DMBA-DNA adduct formation in the target organ in vivo. cDNA microarray analysis indicates that both selenium compounds alter genes in a manner that leads to inhibition of cell proliferation and induction of apoptosis; modulation of apoptosis and cell proliferation can account for chemoprevention during the post-Initiation Phase of mammary carcinogenesis. Using a rat mammary cancer cell line, we compared p-XSC and p-XSeSG as inhibitors of cell proliferation; depending on the selenium dose and time point selected, p-XSC was comparable to or better than p-XSeSG. Collectively, the results described here, suggest that the molecular targets modulated by organoselenium compounds are highly useful indicators of success in clinical cancer chemoprevention trials.