117 results on '"James DE"'
Search Results
2. SILAC kinase screen identifies potential MASTL substrates.
- Author
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Marzec, KA, Rogers, S, McCloy, R, Parker, BL, James, DE, Watkins, DN, Burgess, A, Marzec, KA, Rogers, S, McCloy, R, Parker, BL, James, DE, Watkins, DN, and Burgess, A
- Abstract
Microtubule-associated serine/threonine kinase-like (MASTL) has emerged as a critical regulator of mitosis and as a potential oncogene in a variety of cancer types. To date, Arpp-19/ENSA are the only known substrates of MASTL. However, with the roles of MASTL expanding and increased interest in development of MASTL inhibitors, it has become critical to determine if there are additional substrates and what the optimal consensus motif for MASTL is. Here we utilized a whole cell lysate in vitro kinase screen combined with stable isotope labelling of amino acids in cell culture (SILAC) to identify potential substrates and the residue preference of MASTL. Using the related AGC kinase family members AKT1/2, the kinase screen identified several known and new substrates highly enriched for the validated consensus motif of AKT. Applying this method to MASTL identified 59 phospho-sites on 67 proteins that increased in the presence of active MASTL. Subsequent in vitro kinase assays suggested that MASTL may phosphorylate hnRNPM, YB1 and TUBA1C under certain in vitro conditions. Taken together, these data suggest that MASTL may phosphorylate several additional substrates, providing insight into the ever-increasing biological functions and roles MASTL plays in driving cancer progression and therapy resistance.
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- 2022
3. Trafficking regulator of GLUT4-1 (TRARG1) is a GSK3 substrate.
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Duan, X, Norris, DM, Humphrey, SJ, Yang, P, Cooke, KC, Bultitude, WP, Parker, BL, Conway, OJ, Burchfield, JG, Krycer, JR, Brodsky, FM, James, DE, Fazakerley, DJ, Duan, X, Norris, DM, Humphrey, SJ, Yang, P, Cooke, KC, Bultitude, WP, Parker, BL, Conway, OJ, Burchfield, JG, Krycer, JR, Brodsky, FM, James, DE, and Fazakerley, DJ
- Abstract
Trafficking regulator of GLUT4-1, TRARG1, positively regulates insulin-stimulated GLUT4 trafficking and insulin sensitivity. However, the mechanism(s) by which this occurs remain(s) unclear. Using biochemical and mass spectrometry analyses we found that TRARG1 is dephosphorylated in response to insulin in a PI3K/Akt-dependent manner and is a novel substrate for GSK3. Priming phosphorylation of murine TRARG1 at serine 84 allows for GSK3-directed phosphorylation at serines 72, 76 and 80. A similar pattern of phosphorylation was observed in human TRARG1, suggesting that our findings are translatable to human TRARG1. Pharmacological inhibition of GSK3 increased cell surface GLUT4 in cells stimulated with a submaximal insulin dose, and this was impaired following Trarg1 knockdown, suggesting that TRARG1 acts as a GSK3-mediated regulator in GLUT4 trafficking. These data place TRARG1 within the insulin signaling network and provide insights into how GSK3 regulates GLUT4 trafficking in adipocytes.
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- 2022
4. Proteome-wide systems genetics identifies UFMylation as a regulator of skeletal muscle function
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Molendijk, J, Blazev, R, Mills, RJ, Ng, Y-K, Watt, K, Chau, D, Gregorevic, P, Crouch, PJ, Hilton, JBW, Lisowski, L, Zhang, P, Reue, K, Lusis, AJ, Hudson, JE, James, DE, Seldin, MM, Parker, BL, Molendijk, J, Blazev, R, Mills, RJ, Ng, Y-K, Watt, K, Chau, D, Gregorevic, P, Crouch, PJ, Hilton, JBW, Lisowski, L, Zhang, P, Reue, K, Lusis, AJ, Hudson, JE, James, DE, Seldin, MM, and Parker, BL
- Abstract
Improving muscle function has great potential to improve the quality of life. To identify novel regulators of skeletal muscle metabolism and function, we performed a proteomic analysis of gastrocnemius muscle from 73 genetically distinct inbred mouse strains, and integrated the data with previously acquired genomics and >300 molecular/phenotypic traits via quantitative trait loci mapping and correlation network analysis. These data identified thousands of associations between protein abundance and phenotypes and can be accessed online (https://muscle.coffeeprot.com/) to identify regulators of muscle function. We used this resource to prioritize targets for a functional genomic screen in human bioengineered skeletal muscle. This identified several negative regulators of muscle function including UFC1, an E2 ligase for protein UFMylation. We show UFMylation is up-regulated in a mouse model of amyotrophic lateral sclerosis, a disease that involves muscle atrophy. Furthermore, in vivo knockdown of UFMylation increased contraction force, implicating its role as a negative regulator of skeletal muscle function.
- Published
- 2022
5. A hierarchical approach to removal of unwanted variation for large-scale metabolomics data
- Author
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Kim, T, Tang, O, Vernon, ST, Kott, KA, Koay, YC, Park, J, James, DE, Grieve, SM, Speed, TP, Yang, P, Figtree, GA, O'Sullivan, JF, Yang, JYH, Kim, T, Tang, O, Vernon, ST, Kott, KA, Koay, YC, Park, J, James, DE, Grieve, SM, Speed, TP, Yang, P, Figtree, GA, O'Sullivan, JF, and Yang, JYH
- Abstract
Liquid chromatography-mass spectrometry-based metabolomics studies are increasingly applied to large population cohorts, which run for several weeks or even years in data acquisition. This inevitably introduces unwanted intra- and inter-batch variations over time that can overshadow true biological signals and thus hinder potential biological discoveries. To date, normalisation approaches have struggled to mitigate the variability introduced by technical factors whilst preserving biological variance, especially for protracted acquisitions. Here, we propose a study design framework with an arrangement for embedding biological sample replicates to quantify variance within and between batches and a workflow that uses these replicates to remove unwanted variation in a hierarchical manner (hRUV). We use this design to produce a dataset of more than 1000 human plasma samples run over an extended period of time. We demonstrate significant improvement of hRUV over existing methods in preserving biological signals whilst removing unwanted variation for large scale metabolomics studies. Our tools not only provide a strategy for large scale data normalisation, but also provides guidance on the design strategy for large omics studies.
- Published
- 2021
6. Core functional nodes and sex-specific pathways in human ischaemic and dilated cardiomyopathy
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Li, M, Parker, BL, Pearson, E, Hunter, B, Cao, J, Koay, YC, Guneratne, O, James, DE, Yang, J, Lal, S, O'Sullivan, JF, Li, M, Parker, BL, Pearson, E, Hunter, B, Cao, J, Koay, YC, Guneratne, O, James, DE, Yang, J, Lal, S, and O'Sullivan, JF
- Abstract
Poor access to human left ventricular myocardium is a significant limitation in the study of heart failure (HF). Here, we utilise a carefully procured large human heart biobank of cryopreserved left ventricular myocardium to obtain direct molecular insights into ischaemic cardiomyopathy (ICM) and dilated cardiomyopathy (DCM), the most common causes of HF worldwide. We perform unbiased, deep proteomic and metabolomic analyses of 51 left ventricular (LV) samples from 44 cryopreserved human ICM and DCM hearts, compared to age-, gender-, and BMI-matched, histopathologically normal, donor controls. We report a dramatic reduction in serum amyloid A1 protein in ICM hearts, perturbed thyroid hormone signalling pathways and significant reductions in oxidoreductase co-factor riboflavin-5-monophosphate and glycolytic intermediate fructose-6-phosphate in both; unveil gender-specific changes in HF, including nitric oxide-related arginine metabolism, mitochondrial substrates, and X chromosome-linked protein and metabolite changes; and provide an interactive online application as a publicly-available resource.
- Published
- 2020
7. Reduced insulin action in muscle of high fat diet rats over the diurnal cycle is not associated with defective insulin signaling
- Author
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Small, L, Brandon, AE, Parker, BL, Deshpande, V, Samsudeen, AF, Kowalski, GM, Reznick, J, Wilks, DL, Preston, E, Bruce, CR, James, DE, Turner, N, Cooney, GJ, Small, L, Brandon, AE, Parker, BL, Deshpande, V, Samsudeen, AF, Kowalski, GM, Reznick, J, Wilks, DL, Preston, E, Bruce, CR, James, DE, Turner, N, and Cooney, GJ
- Abstract
OBJECTIVE: Energy metabolism and insulin action follow a diurnal rhythm. It is therefore important that investigations into dysregulation of these pathways are relevant to the physiology of this diurnal rhythm. METHODS: We examined glucose uptake, markers of insulin action, and the phosphorylation of insulin signaling intermediates in muscle of chow and high fat, high sucrose (HFHS) diet-fed rats over the normal diurnal cycle. RESULTS: HFHS animals displayed hyperinsulinemia but had reduced systemic glucose disposal and lower muscle glucose uptake during the feeding period. Analysis of gene expression, enzyme activity, protein abundance and phosphorylation revealed a clear diurnal regulation of substrate oxidation pathways with no difference in Akt signaling in muscle. Transfection of a constitutively active Akt2 into the muscle of HFHS rats did not rescue diet-induced reductions in insulin-stimulated glucose uptake. CONCLUSIONS: These studies suggest that reduced glucose uptake in muscle during the diurnal cycle induced by short-term HFHS-feeding is not the result of reduced insulin signaling.
- Published
- 2019
8. Phosphoproteomics reveals conserved exercise-stimulated signaling and AMPK regulation of store-operated calcium entry
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Nelson, ME, Parker, BL, Burchfield, JG, Hoffman, NJ, Needham, EJ, Cooke, KC, Naim, T, Sylow, L, Ling, NXY, Francis, D, Norris, DM, Chaudhuri, R, Oakhill, JS, Richter, EA, Lynch, GS, Stockli, J, James, DE, Nelson, ME, Parker, BL, Burchfield, JG, Hoffman, NJ, Needham, EJ, Cooke, KC, Naim, T, Sylow, L, Ling, NXY, Francis, D, Norris, DM, Chaudhuri, R, Oakhill, JS, Richter, EA, Lynch, GS, Stockli, J, and James, DE
- Abstract
Exercise stimulates cellular and physiological adaptations that are associated with widespread health benefits. To uncover conserved protein phosphorylation events underlying this adaptive response, we performed mass spectrometry-based phosphoproteomic analyses of skeletal muscle from two widely used rodent models: treadmill running in mice and in situ muscle contraction in rats. We overlaid these phosphoproteomic signatures with cycling in humans to identify common cross-species phosphosite responses, as well as unique model-specific regulation. We identified > 22,000 phosphosites, revealing orthologous protein phosphorylation and overlapping signaling pathways regulated by exercise. This included two conserved phosphosites on stromal interaction molecule 1 (STIM1), which we validate as AMPK substrates. Furthermore, we demonstrate that AMPK-mediated phosphorylation of STIM1 negatively regulates store-operated calcium entry, and this is beneficial for exercise in Drosophila. This integrated cross-species resource of exercise-regulated signaling in human, mouse, and rat skeletal muscle has uncovered conserved networks and unraveled crosstalk between AMPK and intracellular calcium flux.
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- 2019
9. Global redox proteome and phosphoproteome analysis reveals redox switch in Akt.
- Author
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Su, Z, Burchfield, JG, Yang, P, Humphrey, SJ, Yang, G, Francis, D, Yasmin, S, Shin, S-Y, Norris, DM, Kearney, AL, Astore, MA, Scavuzzo, J, Fisher-Wellman, KH, Wang, Q-P, Parker, BL, Neely, GG, Vafaee, F, Chiu, J, Yeo, R, Hogg, PJ, Fazakerley, DJ, Nguyen, LK, Kuyucak, S, James, DE, Su, Z, Burchfield, JG, Yang, P, Humphrey, SJ, Yang, G, Francis, D, Yasmin, S, Shin, S-Y, Norris, DM, Kearney, AL, Astore, MA, Scavuzzo, J, Fisher-Wellman, KH, Wang, Q-P, Parker, BL, Neely, GG, Vafaee, F, Chiu, J, Yeo, R, Hogg, PJ, Fazakerley, DJ, Nguyen, LK, Kuyucak, S, and James, DE
- Abstract
Protein oxidation sits at the intersection of multiple signalling pathways, yet the magnitude and extent of crosstalk between oxidation and other post-translational modifications remains unclear. Here, we delineate global changes in adipocyte signalling networks following acute oxidative stress and reveal considerable crosstalk between cysteine oxidation and phosphorylation-based signalling. Oxidation of key regulatory kinases, including Akt, mTOR and AMPK influences the fidelity rather than their absolute activation state, highlighting an unappreciated interplay between these modifications. Mechanistic analysis of the redox regulation of Akt identified two cysteine residues in the pleckstrin homology domain (C60 and C77) to be reversibly oxidized. Oxidation at these sites affected Akt recruitment to the plasma membrane by stabilizing the PIP3 binding pocket. Our data provide insights into the interplay between oxidative stress-derived redox signalling and protein phosphorylation networks and serve as a resource for understanding the contribution of cellular oxidation to a range of diseases.
- Published
- 2019
10. High dietary fat and sucrose results in an extensive and time-dependent deterioration in health of multiple physiological systems in mice
- Author
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Burchfield, JG, Kebede, MA, Meoli, CC, Stöckli, J, Whitworth, PT, Wright, AL, Hoffman, NJ, Minard, AY, Ma, X, Krycer, JR, Nelson, ME, Tan, SX, Yau, B, Thomas, KC, Wee, NKY, Khor, EC, Enriquez, RF, Vissel, B, Biden, TJ, Baldock, PA, Hoehn, KL, Cantley, J, Cooney, GJ, James, DE, Fazakerley, DJ, Burchfield, JG, Kebede, MA, Meoli, CC, Stöckli, J, Whitworth, PT, Wright, AL, Hoffman, NJ, Minard, AY, Ma, X, Krycer, JR, Nelson, ME, Tan, SX, Yau, B, Thomas, KC, Wee, NKY, Khor, EC, Enriquez, RF, Vissel, B, Biden, TJ, Baldock, PA, Hoehn, KL, Cantley, J, Cooney, GJ, James, DE, and Fazakerley, DJ
- Abstract
© 2018 by The American Society for Biochemistry and Molecular Biology, Inc. Obesity is associated with metabolic dysfunction, including insulin resistance and hyperinsulinemia, and with disorders such as cardiovascular disease, osteoporosis, and neurodegen-eration. Typically, these pathologies are examined in discrete model systems and with limited temporal resolution, and whether these disorders co-occur is therefore unclear. To address this question, here we examined multiple physiological systems in male C57BL/6J mice following prolonged exposure to a high-fat/high-sucrose diet (HFHSD). HFHSD-fed mice rapidly exhibited metabolic alterations, including obesity, hyperleptinemia, physical inactivity, glucose intolerance, peripheral insulin resistance, fasting hyperglycemia, ectopic lipid deposition, and bone deterioration. Prolonged exposure to HFHSD resulted in morbid obesity, ectopic triglyceride deposition in liver and muscle, extensive bone loss, sarcopenia, hyperinsulinemia, and impaired short-term memory. Although many of these defects are typically associated with aging, HFHSD did not alter telomere length in white blood cells, indicating that this diet did not generally promote all aspects of aging. Strikingly, glucose homeostasis was highly dynamic. Glucose intolerance was evident in HFHSD-fed mice after 1 week and was maintained for 24 weeks. Beyond 24 weeks, however, glucose tolerance improved in HFHSD-fed mice, and by 60 weeks, it was indistinguishable from that of chow-fed mice. This improvement coincided with adaptive -cell hyperplasia and hyperinsulinemia, without changes in insulin sensitivity in muscle or adipose tissue. Assessment of insulin secretion in isolated islets revealed that leptin, which inhibited insulin secretion in the chow-fed mice, potentiated glucose-stimulated insulin secretion in the HFHSD-fed mice after 60 weeks. Overall, the excessive calorie intake was accompanied by deteriorating function of numerous physiological systems.
- Published
- 2018
11. Mitochondrial CoQ deficiency is a common driver of mitochondrial oxidants and insulin resistance
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Fazakerley, DJ, Chaudhuri, R, Yang, P, Maghzal, GJ, Thomas, KC, Krycer, JR, Humphrey, SJ, Parker, BL, Fisher-Wellman, KH, Meoli, CC, Hoffman, NJ, Diskin, C, Burchfield, JG, Cowley, MJ, Kaplan, W, Modrusan, Z, Kolumam, G, Yang, JYH, Chen, DL, Samocha-Bonet, D, Greenfield, JR, Hoehn, KL, Stocker, R, James, DE, Fazakerley, DJ, Chaudhuri, R, Yang, P, Maghzal, GJ, Thomas, KC, Krycer, JR, Humphrey, SJ, Parker, BL, Fisher-Wellman, KH, Meoli, CC, Hoffman, NJ, Diskin, C, Burchfield, JG, Cowley, MJ, Kaplan, W, Modrusan, Z, Kolumam, G, Yang, JYH, Chen, DL, Samocha-Bonet, D, Greenfield, JR, Hoehn, KL, Stocker, R, and James, DE
- Abstract
© Fazakerley et al. Insulin resistance in muscle, adipocytes and liver is a gateway to a number of metabolic diseases. Here, we show a selective deficiency in mitochondrial coenzyme Q (CoQ) in insulin-resistant adipose and muscle tissue. This defect was observed in a range of in vitro insulin resistance models and adipose tissue from insulin-resistant humans and was concomitant with lower expression of mevalonate/CoQ biosynthesis pathway proteins in most models. Pharmacologic or genetic manipulations that decreased mitochondrial CoQ triggered mitochondrial oxidants and insulin resistance while CoQ supplementation in either insulin-resistant cell models or mice restored normal insulin sensitivity. Specifically, lowering of mitochondrial CoQ caused insulin resistance in adipocytes as a result of increased superoxide/hydrogen peroxide production via complex II. These data suggest that mitochondrial CoQ is a proximal driver of mitochondrial oxidants and insulin resistance, and that mechanisms that restore mitochondrial CoQ may be effective therapeutic targets for treating insulin resistance.
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- 2018
12. The transcriptional response to oxidative stress is part of, but not sufficient for, insulin resistance in adipocytes
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Chaudhuri, R, Krycer, JR, Fazakerley, DJ, Fisher-Wellman, KH, Su, Z, Hoehn, KL, Yang, JYH, Kuncic, Z, Vafaee, F, James, DE, Chaudhuri, R, Krycer, JR, Fazakerley, DJ, Fisher-Wellman, KH, Su, Z, Hoehn, KL, Yang, JYH, Kuncic, Z, Vafaee, F, and James, DE
- Abstract
Insulin resistance is a major risk factor for metabolic diseases such as Type 2 diabetes. Although the underlying mechanisms of insulin resistance remain elusive, oxidative stress is a unifying driver by which numerous extrinsic signals and cellular stresses trigger insulin resistance. Consequently, we sought to understand the cellular response to oxidative stress and its role in insulin resistance. Using cultured 3T3-L1 adipocytes, we established a model of physiologically-derived oxidative stress by inhibiting the cycling of glutathione and thioredoxin, which induced insulin resistance as measured by impaired insulin-stimulated 2-deoxyglucose uptake. Using time-resolved transcriptomics, we found > 2000 genes differentially-expressed over 24 hours, with specific metabolic and signalling pathways enriched at different times. We explored this coordination using a knowledge-based hierarchical-clustering approach to generate a temporal transcriptional cascade and identify key transcription factors responding to oxidative stress. This response shared many similarities with changes observed in distinct insulin resistance models. However, an anti-oxidant reversed insulin resistance phenotypically but not transcriptionally, implying that the transcriptional response to oxidative stress is insufficient for insulin resistance. This suggests that the primary site by which oxidative stress impairs insulin action occurs post-Transcriptionally, warranting a multi-level 'trans-omic' approach when studying time-resolved responses to cellular perturbations.
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- 2018
13. Quantitative proteomic characterization of cellular pathways associated with altered insulin sensitivity in skeletal muscle following high-fat diet feeding and exercise training.
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Kleinert, M, Parker, BL, Jensen, TE, Raun, SH, Pham, P, Han, X, James, DE, Richter, EA, Sylow, L, Kleinert, M, Parker, BL, Jensen, TE, Raun, SH, Pham, P, Han, X, James, DE, Richter, EA, and Sylow, L
- Abstract
Regular exercise elicits advantageous metabolic adaptations in skeletal muscle, such as improved insulin sensitivity. However, the underpinning molecular mechanisms and the effect of diet on muscle exercise training benefits are unclear. We therefore characterized the skeletal muscle proteome following exercise training (ET) in mice fed chow or high-fat diet (HFD). ET increased exercise performance, lowered body-weight, decreased fat mass and improved muscle insulin action in chow- and HFD-fed mice. At the molecular level, ET regulated 170 muscle proteins in chow-fed mice, but only 29 proteins in HFD-fed mice. HFD per se altered 56 proteins, most of which were regulated in a similar direction by ET. To identify proteins that might have particular health-related bearing on skeletal muscle metabolism, we filtered for differentially regulated proteins in response to ET and HFD. This yielded 15 proteins, including the major urinary protein 1 (MUP1), which was the protein most decreased after HFD, but increased with ET. The ET-induced Mup1 expression was absent in mouse muscle lacking functional AMPK. MUP1 also potentiated insulin-stimulated GLUT4 translocation in cultured muscle cells. Collectively, we provide a resource of ET-regulated proteins in insulin-sensitive and insulin-resistant skeletal muscle. The identification of MUP1 as a diet-, ET- and AMPK-regulated skeletal muscle protein that improves insulin sensitivity in muscle cells demonstrates the usefulness of these data.
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- 2018
14. Mitochondrial CoQ deficiency is a common driver of mitochondria! oxidants and insulin resistance
- Author
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Fazakerley, DJ, Chaudhuri, R, Yang, P, Maghzal, GJ, Thomas, KC, Krycer, JR, Humphrey, SJ, Parker, BL, Fisher-Wellman, KH, Meoli, CC, Hoffman, NJ, Diskin, C, Burchfield, JG, Cowley, MJ, Kaplan, W, Modrusan, Z, Kolumam, G, Yang, JYH, Chen, DL, Samocha-Bonet, D, Greenfield, JR, Hoehn, KL, Stocker, R, James, DE, Fazakerley, DJ, Chaudhuri, R, Yang, P, Maghzal, GJ, Thomas, KC, Krycer, JR, Humphrey, SJ, Parker, BL, Fisher-Wellman, KH, Meoli, CC, Hoffman, NJ, Diskin, C, Burchfield, JG, Cowley, MJ, Kaplan, W, Modrusan, Z, Kolumam, G, Yang, JYH, Chen, DL, Samocha-Bonet, D, Greenfield, JR, Hoehn, KL, Stocker, R, and James, DE
- Abstract
Insulin resistance in muscle, adipocytes and liver is a gateway to a number of metabolic diseases. Here, we show a selective deficiency in mitochondrial coenzyme Q (CoQ) in insulin-resistant adipose and muscle tissue. This defect was observed in a range of in vitro insulin resistance models and adipose tissue from insulin-resistant humans and was concomitant with lower expression of mevalonate/CoQ biosynthesis pathway proteins in most models. Pharmacologic or genetic manipulations that decreased mitochondrial CoQ triggered mitochondrial oxidants and insulin resistance while CoQ supplementation in either insulin-resistant cell models or mice restored normal insulin sensitivity. Specifically, lowering of mitochondrial CoQ caused insulin resistance in adipocytes as a result of increased superoxide/hydrogen peroxide production via complex II. These data suggest that mitochondrial CoQ is a proximal driver of mitochondrial oxidants and insulin resistance, and that mechanisms that restore mitochondrial CoQ may be effective therapeutic targets for treating insulin resistance.
- Published
- 2018
15. MASTL overexpression promotes chromosome instability and metastasis in breast cancer
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Rogers, S, McCloy, RA, Parker, BL, Gallego-Ortega, D, Law, AMK, Chin, VT, Conway, JRW, Fey, D, Millar, EKA, O'Toole, S, Deng, N, Swarbrick, A, Chastain, PD, Cesare, AJ, Timpson, P, Caldon, CE, Croucher, DR, James, DE, Watkins, DN, Burgess, A, Rogers, S, McCloy, RA, Parker, BL, Gallego-Ortega, D, Law, AMK, Chin, VT, Conway, JRW, Fey, D, Millar, EKA, O'Toole, S, Deng, N, Swarbrick, A, Chastain, PD, Cesare, AJ, Timpson, P, Caldon, CE, Croucher, DR, James, DE, Watkins, DN, and Burgess, A
- Abstract
MASTL kinase is essential for correct progression through mitosis, with loss of MASTL causing chromosome segregation errors, mitotic collapse and failure of cytokinesis. However, in cancer MASTL is most commonly amplified and overexpressed. This correlates with increased chromosome instability in breast cancer and poor patient survival in breast, ovarian and lung cancer. Global phosphoproteomic analysis of immortalised breast MCF10A cells engineered to overexpressed MASTL revealed disruption to desmosomes, actin cytoskeleton, PI3K/AKT/mTOR and p38 stress kinase signalling pathways. Notably, these pathways were also disrupted in patient samples that overexpress MASTL. In MCF10A cells, these alterations corresponded with a loss of contact inhibition and partial epithelial-mesenchymal transition, which disrupted migration and allowed cells to proliferate uncontrollably in 3D culture. Furthermore, MASTL overexpression increased aberrant mitotic divisions resulting in increased micronuclei formation. Mathematical modelling indicated that this delay was due to continued inhibition of PP2A-B55, which delayed timely mitotic exit. This corresponded with an increase in DNA damage and delayed transit through interphase. There were no significant alterations to replication kinetics upon MASTL overexpression, however, inhibition of p38 kinase rescued the interphase delay, suggesting the delay was a G2 DNA damage checkpoint response. Importantly, knockdown of MASTL, reduced cell proliferation, prevented invasion and metastasis of MDA-MB-231 breast cancer cells both in vitro and in vivo, indicating the potential of future therapies that target MASTL. Taken together, these results suggest that MASTL overexpression contributes to chromosome instability and metastasis, thereby decreasing breast cancer patient survival.
- Published
- 2018
16. Exercise-induced molecular mechanisms promoting glycogen supercompensation in human skeletal muscle.
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Hingst, JR, Bruhn, L, Hansen, MB, Rosschou, MF, Birk, JB, Fentz, J, Foretz, M, Viollet, B, Sakamoto, K, Færgeman, NJ, Havelund, JF, Parker, BL, James, DE, Kiens, B, Richter, EA, Jensen, J, Wojtaszewski, JFP, Hingst, JR, Bruhn, L, Hansen, MB, Rosschou, MF, Birk, JB, Fentz, J, Foretz, M, Viollet, B, Sakamoto, K, Færgeman, NJ, Havelund, JF, Parker, BL, James, DE, Kiens, B, Richter, EA, Jensen, J, and Wojtaszewski, JFP
- Abstract
OBJECTIVE: A single bout of exercise followed by intake of carbohydrates leads to glycogen supercompensation in prior exercised muscle. Our objective was to illuminate molecular mechanisms underlying this phenomenon in skeletal muscle of man. METHODS: We studied the temporal regulation of glycogen supercompensation in human skeletal muscle during a 5 day recovery period following a single bout of exercise. Nine healthy men depleted (day 1), normalized (day 2) and supercompensated (day 5) muscle glycogen in one leg while the contralateral leg served as a resting control. Euglycemic hyperinsulinemic clamps in combination with leg balance technique allowed for investigating insulin-stimulated leg glucose uptake under these 3 experimental conditions. Cellular signaling in muscle biopsies was investigated by global proteomic analyses and immunoblotting. We strengthened the validity of proposed molecular effectors by follow-up studies in muscle of transgenic mice. RESULTS: Sustained activation of glycogen synthase (GS) and AMPK in combination with elevated expression of proteins determining glucose uptake capacity were evident in the prior exercised muscle. We hypothesize that these alterations offset the otherwise tight feedback inhibition of glycogen synthesis and glucose uptake by glycogen. In line with key roles of AMPK and GS seen in the human experiments we observed abrogated ability for glycogen supercompensation in muscle with inducible AMPK deletion and in muscle carrying a G6P-insensitive form of GS in muscle. CONCLUSION: Our study demonstrates that both AMPK and GS are key regulators of glycogen supercompensation following a single bout of glycogen-depleting exercise in skeletal muscle of both man and mouse.
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- 2018
17. Caracterización de la práctica pedagógica de una docente de la Institución Educativa la Marina sede San Gabriel, en el municipio de Santuario Risaralda
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Montoya Álzate, Luz Stella, Ramírez Cano, James de Jesús, Montoya Álzate, Luz Stella, and Ramírez Cano, James de Jesús
- Abstract
El presente trabajo investigativo tuvo como objetivo caracterizar la práctica educativa de una docente en su quehacer pedagógico y didáctico, ahora bien, el trabajo ahondó dentro de una mirada a la forma como se desarrolla la vida dentro del aula y su relación entre estudiantes y docente en el componente de escuela nueva, donde asisten 12 estudiantes entre los 7 y 12 años; la investigación fue de enfoque cualitativo, donde durante el trabajo de campo que se ejecutó en la Institución Educativa La Marina sede San Gabriel, en el municipio de Santuario Risaralda, ase utilizó la observación y la video grabación, sin intervenir ni criticar los enfoques pedagógicos caracterizados. En este sentido, dentro del trabajo se resaltó la forma, como se evidencio a través del análisis, que la docente no se enmarca en ningún modelo pedagógico, confirmando los estudios realizados por la UNESCO y la OEI, quienes establecen que el proceso de aprendizaje de niños y jóvenes tiene que ver con lo que creen, pueden y están dispuestos a hacer los docentes; es decir, la situación de los profesores en América Latina se marca por falencias de tipo personal, debilidades en la formación y condiciones de trabajo, más que por un déficit en la disponibilidad de profesores. Iniciaremos un recorrido estructurado, que nos permitirá ir avanzando conforme a los objetivos propuestos, dentro de la investigación, se buscó ser lo más asertivo y claro posible en la caracterización de la práctica educativa, utilizando un lenguaje sencillo que estuviese al alcance del mas desprevenido lector, para su fácil entendimiento y discernimiento, en este sentido se acudió a reconocidos teóricos tradicionales y no tradicionales, así como a reconocidas organizaciones cuyo objeto es la educación en el ámbito nacional e internacional, para tener mayores elementos de juicio que le permitieran tener fundamentos más amplios, objetividad, razonabilidad y coherencia al presente trabajo. Esperando haber contribuido en parte, para
- Published
- 2018
18. MASTL overexpression promotes chromosome instability and metastasis in breast cancer.
- Author
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Rogers, S, McCloy, RA, Parker, BL, Gallego-Ortega, D, Law, AMK, Chin, VT, Conway, JRW, Fey, D, Millar, EKA, O'Toole, S, Deng, N, Swarbrick, A, Chastain, PD, Cesare, AJ, Timpson, P, Caldon, CE, Croucher, DR, James, DE, Watkins, DN, Burgess, A, Rogers, S, McCloy, RA, Parker, BL, Gallego-Ortega, D, Law, AMK, Chin, VT, Conway, JRW, Fey, D, Millar, EKA, O'Toole, S, Deng, N, Swarbrick, A, Chastain, PD, Cesare, AJ, Timpson, P, Caldon, CE, Croucher, DR, James, DE, Watkins, DN, and Burgess, A
- Abstract
MASTL kinase is essential for correct progression through mitosis, with loss of MASTL causing chromosome segregation errors, mitotic collapse and failure of cytokinesis. However, in cancer MASTL is most commonly amplified and overexpressed. This correlates with increased chromosome instability in breast cancer and poor patient survival in breast, ovarian and lung cancer. Global phosphoproteomic analysis of immortalised breast MCF10A cells engineered to overexpressed MASTL revealed disruption to desmosomes, actin cytoskeleton, PI3K/AKT/mTOR and p38 stress kinase signalling pathways. Notably, these pathways were also disrupted in patient samples that overexpress MASTL. In MCF10A cells, these alterations corresponded with a loss of contact inhibition and partial epithelial-mesenchymal transition, which disrupted migration and allowed cells to proliferate uncontrollably in 3D culture. Furthermore, MASTL overexpression increased aberrant mitotic divisions resulting in increased micronuclei formation. Mathematical modelling indicated that this delay was due to continued inhibition of PP2A-B55, which delayed timely mitotic exit. This corresponded with an increase in DNA damage and delayed transit through interphase. There were no significant alterations to replication kinetics upon MASTL overexpression, however, inhibition of p38 kinase rescued the interphase delay, suggesting the delay was a G2 DNA damage checkpoint response. Importantly, knockdown of MASTL, reduced cell proliferation, prevented invasion and metastasis of MDA-MB-231 breast cancer cells both in vitro and in vivo, indicating the potential of future therapies that target MASTL. Taken together, these results suggest that MASTL overexpression contributes to chromosome instability and metastasis, thereby decreasing breast cancer patient survival.
- Published
- 2018
19. High dietary fat and sucrose results in an extensive and time-dependent deterioration in health of multiple physiological systems in mice
- Author
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Burchfield, JG, Kebede, MA, Meoli, CC, Stöckli, J, Whitworth, PT, Wright, AL, Hoffman, NJ, Minard, AY, Ma, X, Krycer, JR, Nelson, ME, Tan, SX, Yau, B, Thomas, KC, Wee, NKY, Khor, EC, Enriquez, RF, Vissel, B, Biden, TJ, Baldock, PA, Hoehn, KL, Cantley, J, Cooney, GJ, James, DE, Fazakerley, DJ, Burchfield, JG, Kebede, MA, Meoli, CC, Stöckli, J, Whitworth, PT, Wright, AL, Hoffman, NJ, Minard, AY, Ma, X, Krycer, JR, Nelson, ME, Tan, SX, Yau, B, Thomas, KC, Wee, NKY, Khor, EC, Enriquez, RF, Vissel, B, Biden, TJ, Baldock, PA, Hoehn, KL, Cantley, J, Cooney, GJ, James, DE, and Fazakerley, DJ
- Abstract
© 2018 by The American Society for Biochemistry and Molecular Biology, Inc. Obesity is associated with metabolic dysfunction, including insulin resistance and hyperinsulinemia, and with disorders such as cardiovascular disease, osteoporosis, and neurodegen-eration. Typically, these pathologies are examined in discrete model systems and with limited temporal resolution, and whether these disorders co-occur is therefore unclear. To address this question, here we examined multiple physiological systems in male C57BL/6J mice following prolonged exposure to a high-fat/high-sucrose diet (HFHSD). HFHSD-fed mice rapidly exhibited metabolic alterations, including obesity, hyperleptinemia, physical inactivity, glucose intolerance, peripheral insulin resistance, fasting hyperglycemia, ectopic lipid deposition, and bone deterioration. Prolonged exposure to HFHSD resulted in morbid obesity, ectopic triglyceride deposition in liver and muscle, extensive bone loss, sarcopenia, hyperinsulinemia, and impaired short-term memory. Although many of these defects are typically associated with aging, HFHSD did not alter telomere length in white blood cells, indicating that this diet did not generally promote all aspects of aging. Strikingly, glucose homeostasis was highly dynamic. Glucose intolerance was evident in HFHSD-fed mice after 1 week and was maintained for 24 weeks. Beyond 24 weeks, however, glucose tolerance improved in HFHSD-fed mice, and by 60 weeks, it was indistinguishable from that of chow-fed mice. This improvement coincided with adaptive -cell hyperplasia and hyperinsulinemia, without changes in insulin sensitivity in muscle or adipose tissue. Assessment of insulin secretion in isolated islets revealed that leptin, which inhibited insulin secretion in the chow-fed mice, potentiated glucose-stimulated insulin secretion in the HFHSD-fed mice after 60 weeks. Overall, the excessive calorie intake was accompanied by deteriorating function of numerous physiological systems.
- Published
- 2018
20. High dietary fat and sucrose results in an extensive and time-dependent deterioration in health of multiple physiological systems in mice
- Author
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Burchfield, JG, Kebede, MA, Meoli, CC, Stöckli, J, Whitworth, PT, Wright, AL, Hoffman, NJ, Minard, AY, Ma, X, Krycer, JR, Nelson, ME, Tan, SX, Yau, B, Thomas, KC, Wee, NKY, Khor, EC, Enriquez, RF, Vissel, B, Biden, TJ, Baldock, PA, Hoehn, KL, Cantley, J, Cooney, GJ, James, DE, Fazakerley, DJ, Burchfield, JG, Kebede, MA, Meoli, CC, Stöckli, J, Whitworth, PT, Wright, AL, Hoffman, NJ, Minard, AY, Ma, X, Krycer, JR, Nelson, ME, Tan, SX, Yau, B, Thomas, KC, Wee, NKY, Khor, EC, Enriquez, RF, Vissel, B, Biden, TJ, Baldock, PA, Hoehn, KL, Cantley, J, Cooney, GJ, James, DE, and Fazakerley, DJ
- Abstract
© 2018 by The American Society for Biochemistry and Molecular Biology, Inc. Obesity is associated with metabolic dysfunction, including insulin resistance and hyperinsulinemia, and with disorders such as cardiovascular disease, osteoporosis, and neurodegen-eration. Typically, these pathologies are examined in discrete model systems and with limited temporal resolution, and whether these disorders co-occur is therefore unclear. To address this question, here we examined multiple physiological systems in male C57BL/6J mice following prolonged exposure to a high-fat/high-sucrose diet (HFHSD). HFHSD-fed mice rapidly exhibited metabolic alterations, including obesity, hyperleptinemia, physical inactivity, glucose intolerance, peripheral insulin resistance, fasting hyperglycemia, ectopic lipid deposition, and bone deterioration. Prolonged exposure to HFHSD resulted in morbid obesity, ectopic triglyceride deposition in liver and muscle, extensive bone loss, sarcopenia, hyperinsulinemia, and impaired short-term memory. Although many of these defects are typically associated with aging, HFHSD did not alter telomere length in white blood cells, indicating that this diet did not generally promote all aspects of aging. Strikingly, glucose homeostasis was highly dynamic. Glucose intolerance was evident in HFHSD-fed mice after 1 week and was maintained for 24 weeks. Beyond 24 weeks, however, glucose tolerance improved in HFHSD-fed mice, and by 60 weeks, it was indistinguishable from that of chow-fed mice. This improvement coincided with adaptive -cell hyperplasia and hyperinsulinemia, without changes in insulin sensitivity in muscle or adipose tissue. Assessment of insulin secretion in isolated islets revealed that leptin, which inhibited insulin secretion in the chow-fed mice, potentiated glucose-stimulated insulin secretion in the HFHSD-fed mice after 60 weeks. Overall, the excessive calorie intake was accompanied by deteriorating function of numerous physiological systems.
- Published
- 2018
21. Caracterización de la práctica pedagógica de una docente de la Institución Educativa la Marina sede San Gabriel, en el municipio de Santuario Risaralda
- Author
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Montoya Álzate, Luz Stella, Ramírez Cano, James de Jesús, Montoya Álzate, Luz Stella, and Ramírez Cano, James de Jesús
- Abstract
El presente trabajo investigativo tuvo como objetivo caracterizar la práctica educativa de una docente en su quehacer pedagógico y didáctico, ahora bien, el trabajo ahondó dentro de una mirada a la forma como se desarrolla la vida dentro del aula y su relación entre estudiantes y docente en el componente de escuela nueva, donde asisten 12 estudiantes entre los 7 y 12 años; la investigación fue de enfoque cualitativo, donde durante el trabajo de campo que se ejecutó en la Institución Educativa La Marina sede San Gabriel, en el municipio de Santuario Risaralda, ase utilizó la observación y la video grabación, sin intervenir ni criticar los enfoques pedagógicos caracterizados. En este sentido, dentro del trabajo se resaltó la forma, como se evidencio a través del análisis, que la docente no se enmarca en ningún modelo pedagógico, confirmando los estudios realizados por la UNESCO y la OEI, quienes establecen que el proceso de aprendizaje de niños y jóvenes tiene que ver con lo que creen, pueden y están dispuestos a hacer los docentes; es decir, la situación de los profesores en América Latina se marca por falencias de tipo personal, debilidades en la formación y condiciones de trabajo, más que por un déficit en la disponibilidad de profesores. Iniciaremos un recorrido estructurado, que nos permitirá ir avanzando conforme a los objetivos propuestos, dentro de la investigación, se buscó ser lo más asertivo y claro posible en la caracterización de la práctica educativa, utilizando un lenguaje sencillo que estuviese al alcance del mas desprevenido lector, para su fácil entendimiento y discernimiento, en este sentido se acudió a reconocidos teóricos tradicionales y no tradicionales, así como a reconocidas organizaciones cuyo objeto es la educación en el ámbito nacional e internacional, para tener mayores elementos de juicio que le permitieran tener fundamentos más amplios, objetividad, razonabilidad y coherencia al presente trabajo. Esperando haber contribuido en parte, para
- Published
- 2018
22. Caracterización de la práctica pedagógica de una docente de la Institución Educativa la Marina sede San Gabriel, en el municipio de Santuario Risaralda
- Author
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Montoya Álzate, Luz Stella, Ramírez Cano, James de Jesús, Montoya Álzate, Luz Stella, and Ramírez Cano, James de Jesús
- Abstract
El presente trabajo investigativo tuvo como objetivo caracterizar la práctica educativa de una docente en su quehacer pedagógico y didáctico, ahora bien, el trabajo ahondó dentro de una mirada a la forma como se desarrolla la vida dentro del aula y su relación entre estudiantes y docente en el componente de escuela nueva, donde asisten 12 estudiantes entre los 7 y 12 años; la investigación fue de enfoque cualitativo, donde durante el trabajo de campo que se ejecutó en la Institución Educativa La Marina sede San Gabriel, en el municipio de Santuario Risaralda, ase utilizó la observación y la video grabación, sin intervenir ni criticar los enfoques pedagógicos caracterizados. En este sentido, dentro del trabajo se resaltó la forma, como se evidencio a través del análisis, que la docente no se enmarca en ningún modelo pedagógico, confirmando los estudios realizados por la UNESCO y la OEI, quienes establecen que el proceso de aprendizaje de niños y jóvenes tiene que ver con lo que creen, pueden y están dispuestos a hacer los docentes; es decir, la situación de los profesores en América Latina se marca por falencias de tipo personal, debilidades en la formación y condiciones de trabajo, más que por un déficit en la disponibilidad de profesores. Iniciaremos un recorrido estructurado, que nos permitirá ir avanzando conforme a los objetivos propuestos, dentro de la investigación, se buscó ser lo más asertivo y claro posible en la caracterización de la práctica educativa, utilizando un lenguaje sencillo que estuviese al alcance del mas desprevenido lector, para su fácil entendimiento y discernimiento, en este sentido se acudió a reconocidos teóricos tradicionales y no tradicionales, así como a reconocidas organizaciones cuyo objeto es la educación en el ámbito nacional e internacional, para tener mayores elementos de juicio que le permitieran tener fundamentos más amplios, objetividad, razonabilidad y coherencia al presente trabajo. Esperando haber contribuido en parte, para
- Published
- 2018
23. Inhibition of hepatic lipogenesis enhances liver tumorigenesis by increasing antioxidant defence and promoting cell survival
- Author
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Nelson, ME, Lahiri, S, Chow, JDY, Byrne, FL, Hargett, SR, Breen, DS, Olzomer, EM, Wu, LE, Cooney, GJ, Turner, N, James, DE, Slack-Davis, JK, Lackner, C, Caldwell, SH, Hoehn, KL, Nelson, ME, Lahiri, S, Chow, JDY, Byrne, FL, Hargett, SR, Breen, DS, Olzomer, EM, Wu, LE, Cooney, GJ, Turner, N, James, DE, Slack-Davis, JK, Lackner, C, Caldwell, SH, and Hoehn, KL
- Abstract
The metabolic pathway of de novo lipogenesis is frequently upregulated in human liver tumours, and its upregulation is associated with poor prognosis. Blocking lipogenesis in cultured liver cancer cells is sufficient to decrease cell viability; however, it is not known whether blocking lipogenesis in vivo can prevent liver tumorigenesis. Herein, we inhibit hepatic lipogenesis in mice by liver-specific knockout of acetyl-CoA carboxylase (ACC) genes and treat the mice with the hepatocellular carcinogen diethylnitrosamine (DEN). Unexpectedly, mice lacking hepatic lipogenesis have a twofold increase in tumour incidence and multiplicity compared to controls. Metabolomics analysis of ACC-deficient liver identifies a marked increase in antioxidants including NADPH and reduced glutathione. Importantly, supplementing primary wild-type hepatocytes with glutathione precursors improves cell survival following DEN treatment to a level indistinguishable from ACC-deficient primary hepatocytes. This study shows that lipogenesis is dispensable for liver tumorigenesis in mice treated with DEN, and identifies an important role for ACC enzymes in redox regulation and cell survival.
- Published
- 2017
24. Dynamic Metabolomics Reveals that Insulin Primes the Adipocyte for Glucose Metabolism
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Krycer, JR, Yugi, K, Hirayama, A, Fazakerley, DJ, Quek, LE, Scalzo, R, Ohno, S, Hodson, MP, Ikeda, S, Shoji, F, Suzuki, K, Domanova, W, Parker, BL, Nelson, ME, Humphrey, SJ, Turner, N, Hoehn, KL, Cooney, GJ, Soga, T, Kuroda, S, James, DE, Krycer, JR, Yugi, K, Hirayama, A, Fazakerley, DJ, Quek, LE, Scalzo, R, Ohno, S, Hodson, MP, Ikeda, S, Shoji, F, Suzuki, K, Domanova, W, Parker, BL, Nelson, ME, Humphrey, SJ, Turner, N, Hoehn, KL, Cooney, GJ, Soga, T, Kuroda, S, and James, DE
- Abstract
Insulin triggers an extensive signaling cascade to coordinate adipocyte glucose metabolism. It is considered that the major role of insulin is to provide anabolic substrates by activating GLUT4-dependent glucose uptake. However, insulin stimulates phosphorylation of many metabolic proteins. To examine the implications of this on glucose metabolism, we performed dynamic tracer metabolomics in cultured adipocytes treated with insulin. Temporal analysis of metabolite concentrations and tracer labeling revealed rapid and distinct changes in glucose metabolism, favoring specific glycolytic branch points and pyruvate anaplerosis. Integrating dynamic metabolomics and phosphoproteomics data revealed that insulin-dependent phosphorylation of anabolic enzymes occurred prior to substrate accumulation. Indeed, glycogen synthesis was activated independently of glucose supply. We refer to this phenomenon as metabolic priming, whereby insulin signaling creates a demand-driven system to “pull” glucose into specific anabolic pathways. This complements the supply-driven regulation of anabolism by substrate accumulation and highlights an additional role for insulin action in adipocyte glucose metabolism. Krycer et al. explore how insulin regulates adipocyte metabolism. It is widely held that energy storage (anabolism) occurs as a substrate accumulates. However, using dynamic tracer metabolomics and overlaying phosphoproteomics data, they find that insulin signaling triggers anabolism before substrates accumulate, creating a “demand-driven” system to prime adipocytes for glucose metabolism.
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- 2017
25. ORTI: An open-access Repository of transcriptional interactions for interrogating mammalian gene expression data
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Vafaee, F, Krycer, JR, Ma, X, Burykin, T, James, DE, Kuncic, Z, Vafaee, F, Krycer, JR, Ma, X, Burykin, T, James, DE, and Kuncic, Z
- Abstract
© 2016 Vafaee et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Transcription factors (TFs) play a fundamental role in coordinating biological processes in response to stimuli. Consequently, we often seek to determine the key TFs and their regulated target genes (TGs) amidst gene expression data. This requires a knowledge-base of TF-TG interactions, which would enable us to determine the topology of the transcriptional network and predict novel regulatory interactions. To address this, we generated an Openaccess Repository of Transcriptional Interactions, ORTI, by integrating available TF-TG interaction databases. These databases rely on different types of experimental evidence, including low-throughput assays, high-throughput screens, and bioinformatics predictions. We have subsequently categorised TF-TG interactions in ORTI according to the quality of this evidence. To demonstrate its capabilities, we applied ORTI to gene expression data and identified modulated TFs using an enrichment analysis. Combining this with pairwise TF-TG interactions enabled us to visualise temporal regulation of a transcriptional network. Additionally, ORTI enables the prediction of novel TF-TG interactions, based on how well candidate genes co-express with known TGs of the target TF. By filtering out known TF-TG interactions that are unlikely to occur within the experimental context, this analysis predicts context-specific TF-TG interactions. We show that this can be applied to experimental designs of varying complexities. In conclusion, ORTI is a rich and publicly available database of experimentally validated mammalian transcriptional interactions which is accompanied with tools that can identify and predict transcriptional interactions, serving as a useful resource for unravelling the topology
- Published
- 2016
26. Skeletal muscle and plasma lipidomic signatures of insulin resistance and overweight/obesity in humans
- Author
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Tonks, KT, Coster, AC, Christopher, MJ, Chaudhuri, R, Xu, A, Gagnon-Bartsch, J, Chisholm, DJ, James, DE, Meikle, PJ, Greenfield, JR, Samocha-Bonet, D, Tonks, KT, Coster, AC, Christopher, MJ, Chaudhuri, R, Xu, A, Gagnon-Bartsch, J, Chisholm, DJ, James, DE, Meikle, PJ, Greenfield, JR, and Samocha-Bonet, D
- Abstract
Objective Alterations in lipids in muscle and plasma have been documented in insulin-resistant people with obesity. Whether these lipid alterations are a reflection of insulin resistance or obesity remains unclear. Methods Nondiabetic sedentary individuals not treated with lipid-lowering medications were studied (n = 51). Subjects with body mass index (BMI) > 25 kg/m2 (n = 28) were stratified based on median glucose infusion rate during a hyperinsulinemic-euglycemic clamp into insulin-sensitive and insulin-resistant groups (above and below median, obesity/insulin-sensitive and obesity/insulin-resistant, respectively). Lean individuals (n = 23) served as a reference group. Lipidomics was performed in muscle and plasma by liquid chromatography electrospray ionization-tandem mass spectrometry. Pathway analysis of gene array in muscle was performed in a subset (n = 35). Results In muscle, insulin resistance was characterized by higher levels of C18:0 sphingolipids, while in plasma, higher levels of diacylglycerol and cholesterol ester, and lower levels of lysophosphatidylcholine and lysoalkylphosphatidylcholine, indicated insulin resistance, irrespective of overweight/obesity. The sphingolipid metabolism gene pathway was upregulated in muscle in insulin resistance independent of obesity. An overweight/obesity lipidomic signature was only apparent in plasma, predominated by higher triacylglycerol and lower plasmalogen species. Conclusions Muscle C18:0 sphingolipids may play a role in insulin resistance independent of excess adiposity.
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- 2016
27. Skeletal muscle and plasma lipidomic signatures of insulin resistance and overweight/obesity in humans
- Author
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Tonks, KT, Coster, ACF, Christopher, MJ, Chaudhuri, R, Xu, A, Gagnon-Bartsch, J, Chisholm, DJ, James, DE, Meikle, PJ, Greenfield, JR, Samocha-Bonet, D, Tonks, KT, Coster, ACF, Christopher, MJ, Chaudhuri, R, Xu, A, Gagnon-Bartsch, J, Chisholm, DJ, James, DE, Meikle, PJ, Greenfield, JR, and Samocha-Bonet, D
- Abstract
OBJECTIVE: Alterations in lipids in muscle and plasma have been documented in insulin-resistant people with obesity. Whether these lipid alterations are a reflection of insulin resistance or obesity remains unclear. METHODS: Nondiabetic sedentary individuals not treated with lipid-lowering medications were studied (n = 51). Subjects with body mass index (BMI) > 25 kg/m(2) (n = 28) were stratified based on median glucose infusion rate during a hyperinsulinemic-euglycemic clamp into insulin-sensitive and insulin-resistant groups (above and below median, obesity/insulin-sensitive and obesity/insulin-resistant, respectively). Lean individuals (n = 23) served as a reference group. Lipidomics was performed in muscle and plasma by liquid chromatography electrospray ionization-tandem mass spectrometry. Pathway analysis of gene array in muscle was performed in a subset (n = 35). RESULTS: In muscle, insulin resistance was characterized by higher levels of C18:0 sphingolipids, while in plasma, higher levels of diacylglycerol and cholesterol ester, and lower levels of lysophosphatidylcholine and lysoalkylphosphatidylcholine, indicated insulin resistance, irrespective of overweight/obesity. The sphingolipid metabolism gene pathway was upregulated in muscle in insulin resistance independent of obesity. An overweight/obesity lipidomic signature was only apparent in plasma, predominated by higher triacylglycerol and lower plasmalogen species. CONCLUSIONS: Muscle C18:0 sphingolipids may play a role in insulin resistance independent of excess adiposity.
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- 2016
28. mTORC2 and AMPK differentially regulate muscle triglyceride content via Perilipin 3
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Kleinert, M, Parker, BL, Chaudhuri, R, Fazakerley, DJ, Serup, A, Thomas, KC, Krycer, JR, Sylow, L, Fritzen, AM, Hoffman, NJ, Jeppesen, J, Schjerling, P, Ruegg, MA, Kiens, B, James, DE, Richter, EA, Kleinert, M, Parker, BL, Chaudhuri, R, Fazakerley, DJ, Serup, A, Thomas, KC, Krycer, JR, Sylow, L, Fritzen, AM, Hoffman, NJ, Jeppesen, J, Schjerling, P, Ruegg, MA, Kiens, B, James, DE, and Richter, EA
- Abstract
OBJECTIVE: We have recently shown that acute inhibition of both mTOR complexes (mTORC1 and mTORC2) increases whole-body lipid utilization, while mTORC1 inhibition had no effect. Therefore, we tested the hypothesis that mTORC2 regulates lipid metabolism in skeletal muscle. METHODS: Body composition, substrate utilization and muscle lipid storage were measured in mice lacking mTORC2 activity in skeletal muscle (specific knockout of RICTOR (Ric mKO)). We further examined the RICTOR/mTORC2-controlled muscle metabolome and proteome; and performed follow-up studies in other genetic mouse models and in cell culture. RESULTS: Ric mKO mice exhibited a greater reliance on fat as an energy substrate, a re-partitioning of lean to fat mass and an increase in intramyocellular triglyceride (IMTG) content, along with increases in several lipid metabolites in muscle. Unbiased proteomics revealed an increase in the expression of the lipid droplet binding protein Perilipin 3 (PLIN3) in muscle from Ric mKO mice. This was associated with increased AMPK activity in Ric mKO muscle. Reducing AMPK kinase activity decreased muscle PLIN3 expression and IMTG content. AMPK agonism, in turn, increased PLIN3 expression in a FoxO1 dependent manner. PLIN3 overexpression was sufficient to increase triglyceride content in muscle cells. CONCLUSIONS: We identified a novel link between mTORC2 and PLIN3, which regulates lipid storage in muscle. While mTORC2 is a negative regulator, we further identified AMPK as a positive regulator of PLIN3, which impacts whole-body substrate utilization and nutrient partitioning.
- Published
- 2016
29. Guide pratique de l'export du vin
- Author
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Roany, James de, Resnick, Evelyne, Roany, James de, Roany, James de, Resnick, Evelyne, and Roany, James de
- Abstract
À l’heure où les vignobles français voient le marché intérieur se réduire, les regards se tournent vers les marchés internationaux : États-Unis, Chine, Russie, Inde et bien d’autres. Toutefois, se lancer à l’export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffisante des marchés, mauvais choix des partenaires, financement insuffisant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de : • comprendre les raisons et les objectifs de l’export, • faire votre diagnostic export, • construire votre stratégie export, • gérer la logistique, • évaluer les marchés matures, émergents et prometteurs. Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu’aux étudiants en commercialisation du vin à l’international.
- Published
- 2012
30. Cross-species gene expression analysis identifies a novel set of genes implicated in human insulin sensitivity
- Author
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Chaudhuri, R, Khoo, PS, Tonks, K, Junutula, JR, Kolumam, G, Modrusan, Z, Samocha-Bonet, D, Meoli, CC, Hocking, S, Fazakerley, DJ, Stöckli, J, Hoehn, KL, Greenfield, JR, Yang, JYH, James, DE, Chaudhuri, R, Khoo, PS, Tonks, K, Junutula, JR, Kolumam, G, Modrusan, Z, Samocha-Bonet, D, Meoli, CC, Hocking, S, Fazakerley, DJ, Stöckli, J, Hoehn, KL, Greenfield, JR, Yang, JYH, and James, DE
- Abstract
OBJECTIVE: Insulin resistance (IR) is one of the earliest predictors of type 2 diabetes. However, diagnosis of IR is limited. High fat fed mouse models provide key insights into IR. We hypothesized that early features of IR are associated with persistent changes in gene expression (GE) and endeavored to (a) develop novel methods for improving signal:noise in analysis of human GE using mouse models; (b) identify a GE motif that accurately diagnoses IR in humans; and (c) identify novel biology associated with IR in humans. METHODS: We integrated human muscle GE data with longitudinal mouse GE data and developed an unbiased three-level crossspecies analysis platform (single gene, gene set, and networks) to generate a gene expression motif (GEM) indicative of IR. A logistic regression classification model validated GEM in three independent human data sets (n = 115). RESULTS: This GEM of 93 genes substantially improved diagnosis of IR compared with routine clinical measures across multiple independent data sets. Individuals misclassified by GEM possessed other metabolic features raising the possibility that they represent a separate metabolic subclass. The GEM was enriched in pathways previously implicated in insulin action and revealed novel associations between β-catenin and Jak1 and IR. Functional analyses using small molecule inhibitors showed an important role for these proteins in insulin action. CONCLUSIONS: This study shows that systems approaches for identifying molecular signatures provides a powerful way to stratify individuals into discrete metabolic groups. Moreover, we speculate that the β-catenin pathway may represent a novel biomarker for IR in humans that warrant future investigation.
- Published
- 2015
31. Acetyl CoA carboxylase 2 is dispensable for CD8+ T cell responses
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Lee, JE, Walsh, MC, Hoehn, KL, James, DE, Wherry, EJ, Choi, Y, Lee, JE, Walsh, MC, Hoehn, KL, James, DE, Wherry, EJ, and Choi, Y
- Abstract
Differentiation of T cells is closely associated with dynamic changes in nutrient and energy metabolism. However, the extent to which specific metabolic pathways and molecular components are determinative of CD8+ T cell fate remains unclear. It has been previously established in various tissues that acetyl CoA carboxylase 2 (ACC2) regulates fatty acid oxidation (FAO) by inhibiting carnitine palmitoyltransferase 1 (CPT1), a rate-limiting enzyme of FAO in mitochondria. Here, we explore the cell-intrinsic role of ACC2 in T cell immunity in response to infections. We report here that ACC2 deficiency results in a marginal increase of cellular FAO in CD8+ T cells, but does not appear to influence antigen-specific effector and memory CD8+ T cell responses during infection with listeria or lymphocytic choriomeningitis virus. These results suggest that ACC2 is dispensable for CD8+ T cell responses.
- Published
- 2015
32. An Actin Filament Population Defined by the Tropomyosin Tpm3.1 Regulates Glucose Uptake
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Kee, AJ, Yang, L, Lucas, CA, Greenberg, MJ, Martel, N, Leong, GM, Hughes, WE, Cooney, GJ, James, DE, Ostap, EM, Han, W, Gunning, PW, Hardeman, EC, Kee, AJ, Yang, L, Lucas, CA, Greenberg, MJ, Martel, N, Leong, GM, Hughes, WE, Cooney, GJ, James, DE, Ostap, EM, Han, W, Gunning, PW, and Hardeman, EC
- Abstract
Actin has an ill-defined role in the trafficking of GLUT4 glucose transporter vesicles to the plasma membrane (PM). We have identified novel actin filaments defined by the tropomyosin Tpm3.1 at glucose uptake sites in white adipose tissue (WAT) and skeletal muscle. In Tpm 3.1-overexpressing mice, insulin-stimulated glucose uptake was increased; while Tpm3.1-null mice they were more sensitive to the impact of high-fat diet on glucose uptake. Inhibition of Tpm3.1 function in 3T3-L1 adipocytes abrogates insulin-stimulated GLUT4 translocation and glucose uptake. In WAT, the amount of filamentous actin is determined by Tpm3.1 levels and is paralleled by changes in exocyst component (sec8) and Myo1c levels. In adipocytes, Tpm3.1 localizes with MyoIIA, but not Myo1c, and it inhibits Myo1c binding to actin. We propose that Tpm3.1 determines the amount of cortical actin that can engage MyoIIA and generate contractile force, and in parallel limits the interaction of Myo1c with actin filaments. The balance between these actin filament populations may determine the efficiency of movement and/or fusion of GLUT4 vesicles with the PM. Actin's role in GLUT4 glucose transporter trafficking is unclear. We have identified novel actin filaments defined by the tropomyosin Tpm3.1 that regulates GLUT4 trafficking and glucose uptake. MyoIIA and Myo1c are known regulators of GLUT4 trafficking. Tpm3.1 has been shown to recruit MyoIIA to actin filaments and here we show that it inhibits Myo1c binding. The balance between Tpm3.1/MyoIIA and Tpm3.1-free/Myo1c actin filament populations may determine the efficiency of movement and/or fusion of GLUT4 vesicles with the plasma membrane.
- Published
- 2015
33. Cross-species gene expression analysis identifies a novel set of genes implicated in human insulin sensitivity
- Author
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Chaudhuri, R, Khoo, PS, Tonks, K, Junutula, JR, Kolumam, G, Modrusan, Z, Samocha-Bonet, D, Meoli, CC, Hocking, S, Fazakerley, DJ, Stöckli, J, Hoehn, KL, Greenfield, JR, Yang, JYH, James, DE, Chaudhuri, R, Khoo, PS, Tonks, K, Junutula, JR, Kolumam, G, Modrusan, Z, Samocha-Bonet, D, Meoli, CC, Hocking, S, Fazakerley, DJ, Stöckli, J, Hoehn, KL, Greenfield, JR, Yang, JYH, and James, DE
- Abstract
OBJECTIVE: Insulin resistance (IR) is one of the earliest predictors of type 2 diabetes. However, diagnosis of IR is limited. High fat fed mouse models provide key insights into IR. We hypothesized that early features of IR are associated with persistent changes in gene expression (GE) and endeavored to (a) develop novel methods for improving signal:noise in analysis of human GE using mouse models; (b) identify a GE motif that accurately diagnoses IR in humans; and (c) identify novel biology associated with IR in humans. METHODS: We integrated human muscle GE data with longitudinal mouse GE data and developed an unbiased three-level crossspecies analysis platform (single gene, gene set, and networks) to generate a gene expression motif (GEM) indicative of IR. A logistic regression classification model validated GEM in three independent human data sets (n = 115). RESULTS: This GEM of 93 genes substantially improved diagnosis of IR compared with routine clinical measures across multiple independent data sets. Individuals misclassified by GEM possessed other metabolic features raising the possibility that they represent a separate metabolic subclass. The GEM was enriched in pathways previously implicated in insulin action and revealed novel associations between β-catenin and Jak1 and IR. Functional analyses using small molecule inhibitors showed an important role for these proteins in insulin action. CONCLUSIONS: This study shows that systems approaches for identifying molecular signatures provides a powerful way to stratify individuals into discrete metabolic groups. Moreover, we speculate that the β-catenin pathway may represent a novel biomarker for IR in humans that warrant future investigation.
- Published
- 2015
34. Selective insulin resistance in adipocytes
- Author
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Tan, SX, Fisher-Wellman, KH, Fazakerley, DJ, Ng, Y, Pant, H, Li, J, Meoli, CC, Coster, ACF, Stöckli, J, James, DE, Tan, SX, Fisher-Wellman, KH, Fazakerley, DJ, Ng, Y, Pant, H, Li, J, Meoli, CC, Coster, ACF, Stöckli, J, and James, DE
- Abstract
Aside from glucose metabolism, insulin regulates a variety of pathways in peripheral tissues. Under insulin-resistant conditions, it is well known that insulin-stimulated glucose uptake is impaired, and many studies attribute this to a defect in Akt signaling. Here we make use of several insulin resistance models, including insulin-resistant 3T3-L1 adipocytes and fat explants prepared from high fat-fed C57BL/6J and ob/ob mice, to comprehensively distinguish defective from unaffected aspects of insulin signaling and its downstream consequences in adipocytes. Defective regulation of glucose uptake was observed in all models of insulin resistance, whereas other major actions of insulin such as protein synthesis and anti-lipolysis were normal. This defect corresponded to a reduction in the maximum response to insulin. The pattern of change observed for phosphorylation in the Akt pathway was inconsistent with a simple defect at the level of Akt. The only Akt substrate that showed consistently reduced phosphorylation was the RabGAP AS160 that regulates GLUT4 translocation. We conclude that insulin resistance in adipose tissue is highly selective for glucose metabolism and likely involves a defect in one of the components regulating GLUT4 translocation to the cell surface in response to insulin.
- Published
- 2015
35. Dataset from the global phosphoproteomic mapping of early mitotic exit in human cells
- Author
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Rogers, S, McCloy, RA, Parker, BL, Chaudhuri, R, Gayevskiy, V, Hoffman, NJ, Watkins, DN, Daly, RJ, James, DE, Burgess, A, Rogers, S, McCloy, RA, Parker, BL, Chaudhuri, R, Gayevskiy, V, Hoffman, NJ, Watkins, DN, Daly, RJ, James, DE, and Burgess, A
- Abstract
The presence or absence of a phosphorylation on a substrate at any particular point in time is a functional readout of the balance in activity between the regulatory kinase and the counteracting phosphatase. Understanding how stable or short-lived a phosphorylation site is required for fully appreciating the biological consequences of the phosphorylation. Our current understanding of kinases and their substrates is well established; however, the role phosphatases play is less understood. Therefore, we utilized a phosphatase dependent model of mitotic exit to identify potential substrates that are preferentially dephosphorylated. Using this method, we identified >16,000 phosphosites on >3300 unique proteins, and quantified the temporal phosphorylation changes that occur during early mitotic exit (McCloy et al., 2015 [1]). Furthermore, we annotated the majority of these phosphorylation sites with a high confidence upstream kinase using published, motif and prediction based methods. The results from this study have been deposited into the ProteomeXchange repository with identifier PXD001559. Here we provide additional analysis of this dataset; for each of the major mitotic kinases we identified motifs that correlated strongly with phosphorylation status. These motifs could be used to predict the stability of phosphorylated residues in proteins of interest, and help infer potential functional roles for uncharacterized phosphorylations. In addition, we provide validation at the single cell level that serine residues phosphorylated by Cdk are stable during phosphatase dependent mitotic exit. In summary, this unique dataset contains information on the temporal mitotic stability of thousands of phosphorylation sites regulated by dozens of kinases, and information on the potential preference that phosphatases have at both the protein and individual phosphosite level. The compellation of this data provides an invaluable resource for the wider research community.
- Published
- 2015
36. PhosphOrtholog: a web-based tool for cross-species mapping of orthologous protein post-translational modifications
- Author
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Chaudhuri, R, Sadrieh, A, Hoffman, NJ, Parker, BL, Humphrey, SJ, Stoeckli, J, Hill, AP, James, DE, Yang, JYH, Chaudhuri, R, Sadrieh, A, Hoffman, NJ, Parker, BL, Humphrey, SJ, Stoeckli, J, Hill, AP, James, DE, and Yang, JYH
- Abstract
BACKGROUND: Most biological processes are influenced by protein post-translational modifications (PTMs). Identifying novel PTM sites in different organisms, including humans and model organisms, has expedited our understanding of key signal transduction mechanisms. However, with increasing availability of deep, quantitative datasets in diverse species, there is a growing need for tools to facilitate cross-species comparison of PTM data. This is particularly important because functionally important modification sites are more likely to be evolutionarily conserved; yet cross-species comparison of PTMs is difficult since they often lie in structurally disordered protein domains. Current tools that address this can only map known PTMs between species based on known orthologous phosphosites, and do not enable the cross-species mapping of newly identified modification sites. Here, we addressed this by developing a web-based software tool, PhosphOrtholog ( www.phosphortholog.com ) that accurately maps protein modification sites between different species. This facilitates the comparison of datasets derived from multiple species, and should be a valuable tool for the proteomics community. RESULTS: Here we describe PhosphOrtholog, a web-based application for mapping known and novel orthologous PTM sites from experimental data obtained from different species. PhosphOrtholog is the only generic and automated tool that enables cross-species comparison of large-scale PTM datasets without relying on existing PTM databases. This is achieved through pairwise sequence alignment of orthologous protein residues. To demonstrate its utility we apply it to two sets of human and rat muscle phosphoproteomes generated following insulin and exercise stimulation, respectively, and one publicly available mouse phosphoproteome following cellular stress revealing high mapping and coverage efficiency. Although coverage statistics are dataset dependent, PhosphOrtholog increased the number of cross
- Published
- 2015
37. Guide pratique de l'export du vin Ed. 2
- Author
-
Resnick, Evelyne, Roany, James de, Durieux, Bruno, Resnick, Evelyne, Resnick, Evelyne, Roany, James de, Durieux, Bruno, and Resnick, Evelyne
- Abstract
Si les marchés internationaux (États-Unis, Chine, Russie, Inde…) sont de plus en plus prometteurs, se lancer à l'export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffi sante des marchés, mauvais choix des partenaires, financement insuffi sant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de :• comprendre les raisons et les objectifs de l'export,• faire votre diagnostic export,• construire votre stratégie export,• gérer la logistique,• évaluer les marchés matures, émergents et prometteurs.Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu'aux étudiants en commercialisation du vin à l'international. Cette seconde édition entièrement actualisée détaille en autres les cas des Tigres d'Asie (Vietnam, Laos, Thaïlande) et de l'Afrique.
- Published
- 2014
38. Guide pratique de l'export du vin Ed. 2
- Author
-
Roany, James de, Durieux, Bruno, Resnick, Evelyne, Roany, James de, Durieux, Bruno, and Resnick, Evelyne
- Abstract
Si les marchés internationaux (États-Unis, Chine, Russie, Inde…) sont de plus en plus prometteurs, se lancer à l'export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffi sante des marchés, mauvais choix des partenaires, financement insuffi sant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de :• comprendre les raisons et les objectifs de l'export,• faire votre diagnostic export,• construire votre stratégie export,• gérer la logistique,• évaluer les marchés matures, émergents et prometteurs.Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu'aux étudiants en commercialisation du vin à l'international. Cette seconde édition entièrement actualisée détaille en autres les cas des Tigres d'Asie (Vietnam, Laos, Thaïlande) et de l'Afrique.
- Published
- 2014
39. Genetic inhibition of hepatic acetyl-CoA carboxylase activity increases liver fat and alters global protein acetylation
- Author
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Chow, JDY, Lawrence, RT, Healy, ME, Dominy, JE, Liao, JA, Breen, DS, Byrne, FL, Kenwood, BM, Lackner, C, Okutsu, S, Mas, VR, Caldwell, SH, Tomsig, JL, Cooney, GJ, Puigserver, PB, Turner, N, James, DE, Villén, J, Hoehn, KL, Chow, JDY, Lawrence, RT, Healy, ME, Dominy, JE, Liao, JA, Breen, DS, Byrne, FL, Kenwood, BM, Lackner, C, Okutsu, S, Mas, VR, Caldwell, SH, Tomsig, JL, Cooney, GJ, Puigserver, PB, Turner, N, James, DE, Villén, J, and Hoehn, KL
- Abstract
Lipid deposition in the liver is associated with metabolic disorders including fatty liver disease, type II diabetes, and hepatocellular cancer. The enzymes acetyl-CoA carboxylase 1 (ACC1) and ACC2 are powerful regulators of hepatic fat storage; therefore, their inhibition is expected to prevent the development of fatty liver. In this study we generated liver-specific ACC1 and ACC2 double knockout (LDKO) mice to determine how the loss of ACC activity affects liver fat metabolism and whole-body physiology. Characterization of LDKO mice revealed unexpected phenotypes of increased hepatic triglyceride and decreased fat oxidation. We also observed that chronic ACC inhibition led to hyper-acetylation of proteins in the extra-mitochondrial space. In sum, these data reveal the existence of a compensatory pathway that protects hepatic fat stores when ACC enzymes are inhibited. Furthermore, we identified an important role for ACC enzymes in the regulation of protein acetylation in the extra-mitochondrial space. © 2014 The Authors.
- Published
- 2014
40. Guide pratique de l'export du vin Ed. 2
- Author
-
Resnick, Evelyne, Roany, James de, Durieux, Bruno, Resnick, Evelyne, Resnick, Evelyne, Roany, James de, Durieux, Bruno, and Resnick, Evelyne
- Abstract
Si les marchés internationaux (États-Unis, Chine, Russie, Inde…) sont de plus en plus prometteurs, se lancer à l'export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffi sante des marchés, mauvais choix des partenaires, financement insuffi sant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de :• comprendre les raisons et les objectifs de l'export,• faire votre diagnostic export,• construire votre stratégie export,• gérer la logistique,• évaluer les marchés matures, émergents et prometteurs.Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu'aux étudiants en commercialisation du vin à l'international. Cette seconde édition entièrement actualisée détaille en autres les cas des Tigres d'Asie (Vietnam, Laos, Thaïlande) et de l'Afrique.
- Published
- 2014
41. Guide pratique de l'export du vin Ed. 2
- Author
-
Resnick, Evelyne, Roany, James de, Durieux, Bruno, Resnick, Evelyne, Resnick, Evelyne, Roany, James de, Durieux, Bruno, and Resnick, Evelyne
- Abstract
Si les marchés internationaux (États-Unis, Chine, Russie, Inde…) sont de plus en plus prometteurs, se lancer à l'export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffi sante des marchés, mauvais choix des partenaires, financement insuffi sant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de :• comprendre les raisons et les objectifs de l'export,• faire votre diagnostic export,• construire votre stratégie export,• gérer la logistique,• évaluer les marchés matures, émergents et prometteurs.Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu'aux étudiants en commercialisation du vin à l'international. Cette seconde édition entièrement actualisée détaille en autres les cas des Tigres d'Asie (Vietnam, Laos, Thaïlande) et de l'Afrique.
- Published
- 2014
42. Guide pratique de l'export du vin Ed. 2
- Author
-
Resnick, Evelyne, Roany, James de, Durieux, Bruno, Resnick, Evelyne, Resnick, Evelyne, Roany, James de, Durieux, Bruno, and Resnick, Evelyne
- Abstract
Si les marchés internationaux (États-Unis, Chine, Russie, Inde…) sont de plus en plus prometteurs, se lancer à l'export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffi sante des marchés, mauvais choix des partenaires, financement insuffi sant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de :• comprendre les raisons et les objectifs de l'export,• faire votre diagnostic export,• construire votre stratégie export,• gérer la logistique,• évaluer les marchés matures, émergents et prometteurs.Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu'aux étudiants en commercialisation du vin à l'international. Cette seconde édition entièrement actualisée détaille en autres les cas des Tigres d'Asie (Vietnam, Laos, Thaïlande) et de l'Afrique.
- Published
- 2014
43. Detection of Chern numbers and entanglement in topological two-species systems through subsystem winding numbers
- Author
-
Lisle, James de, De, Suvabrata, Alba, Emilio, Bullivant, Alex, García-Ripoll, Juan José, Lahtinen, Ville, Pachos, Jiannis K., Lisle, James de, De, Suvabrata, Alba, Emilio, Bullivant, Alex, García-Ripoll, Juan José, Lahtinen, Ville, and Pachos, Jiannis K.
- Abstract
Topological invariants, such as the Chern number, characterize topological phases of matter. Here we provide a method to detect Chern numbers in systems with two distinct species of fermion, such as spins, orbitals or several atomic states. We analytically show that the Chern number can be decomposed as a sum of component specific winding numbers, which are themselves physically observable. We apply this method to two systems, the quantum spin Hall insulator and a staggered topological superconductor, and show that (spin) Chern numbers are accurately reproduced. The measurements required for constructing the component winding numbers also enable one to probe the entanglement spectrum with respect to component partitions. Our method is particularly suited to experiments with cold atoms in optical lattices where time-of-flight images can give direct access to the relevant observables. © 2014 IOP Publishing Ltd and Deutsche Physikalische Gesellschaft.
- Published
- 2014
44. Impaired Akt phosphorylation in insulin-resistant human muscle is accompanied by selective and heterogeneous downstream defects
- Author
-
Tonks, KT, Ng, Y, Miller, S, Coster, ACF, Samocha-Bonet, D, Iseli, TJ, Xu, A, Patrick, E, Yang, JYH, Junutula, JR, Modrusan, Z, Kolumam, G, Stöckli, J, Chisholm, DJ, James, DE, Greenfield, JR, Tonks, KT, Ng, Y, Miller, S, Coster, ACF, Samocha-Bonet, D, Iseli, TJ, Xu, A, Patrick, E, Yang, JYH, Junutula, JR, Modrusan, Z, Kolumam, G, Stöckli, J, Chisholm, DJ, James, DE, and Greenfield, JR
- Published
- 2013
45. Guide pratique de l'export du vin
- Author
-
Resnick, Evelyne, Roany, James de, Resnick, Evelyne, and Roany, James de
- Abstract
À l’heure où les vignobles français voient le marché intérieur se réduire, les regards se tournent vers les marchés internationaux : États-Unis, Chine, Russie, Inde et bien d’autres. Toutefois, se lancer à l’export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffisante des marchés, mauvais choix des partenaires, financement insuffisant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de : • comprendre les raisons et les objectifs de l’export, • faire votre diagnostic export, • construire votre stratégie export, • gérer la logistique, • évaluer les marchés matures, émergents et prometteurs. Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu’aux étudiants en commercialisation du vin à l’international.
- Published
- 2012
46. Guide pratique de l'export du vin
- Author
-
Resnick, Evelyne, Roany, James de, Resnick, Evelyne, and Roany, James de
- Abstract
À l’heure où les vignobles français voient le marché intérieur se réduire, les regards se tournent vers les marchés internationaux : États-Unis, Chine, Russie, Inde et bien d’autres. Toutefois, se lancer à l’export sans aucune expérience peut présenter des dangers : mauvaise évaluation des capacités de son entreprise à exporter, analyse insuffisante des marchés, mauvais choix des partenaires, financement insuffisant, etc. Ce guide pratique répondra à toutes vos questions, en vous permettant de : • comprendre les raisons et les objectifs de l’export, • faire votre diagnostic export, • construire votre stratégie export, • gérer la logistique, • évaluer les marchés matures, émergents et prometteurs. Il est destiné aux producteurs, aux caves coopératives, aux négociants et à tous les opérateurs du monde vitivinicole ainsi qu’aux étudiants en commercialisation du vin à l’international.
- Published
- 2012
47. Use of a GLUT4 translocation assay strategy to identify new insulin sensitisers with efficacy in vitro and in vivo
- Author
-
Molero, JC, Konstantopoulos, N, Hohnen-Behrens, C, Ye, Jing, Segal, D, Collier, G, Walder, Ken, James, DE, Molero, JC, Konstantopoulos, N, Hohnen-Behrens, C, Ye, Jing, Segal, D, Collier, G, Walder, Ken, and James, DE
- Published
- 2008
48. Use of a GLUT4 translocation assay strategy to identify new insulin sensitisers with efficacy in vitro and in vivo
- Author
-
Molero, JC, Konstantopoulos, N, Hohnen-Behrens, C, Ye, Jing, Segal, D, Collier, G, Walder, Ken, James, DE, Molero, JC, Konstantopoulos, N, Hohnen-Behrens, C, Ye, Jing, Segal, D, Collier, G, Walder, Ken, and James, DE
- Published
- 2008
49. Erratum: c-Cbl-deficient mice have reduced adiposity, higher energy expenditure, and improved peripheral insulin action (Journal of Clinical Investigation (2004) 114 (1326-1333) (doi:10.1172/JCI200421480))
- Author
-
Molero, JC, Jensen, TE, Withers, PC, Couzens, M, Herzog, H, Thien, CBF, Langdon, WY, Walder, Ken, Murphy, MA, Bowtell, DDL, Hardeman, E, Ghoddusi, M, James, DE, Cooney, GJ, Molero, JC, Jensen, TE, Withers, PC, Couzens, M, Herzog, H, Thien, CBF, Langdon, WY, Walder, Ken, Murphy, MA, Bowtell, DDL, Hardeman, E, Ghoddusi, M, James, DE, and Cooney, GJ
- Published
- 2005
50. Theology News and Notes - Vol. 43, No. 5
- Author
-
Morrison, James; De Pree, Max; Mouw, Richard J.; Grounds, Vernon C.; Spittler, Russell P.; McCullough, Donald W.; Martin, Ralph P.; Hestenes, Roberta; Gillespie, Thomas W.; Cooley, Robert E.; Smith, Inez T., Ryder, Janice E., Morrison, James; De Pree, Max; Mouw, Richard J.; Grounds, Vernon C.; Spittler, Russell P.; McCullough, Donald W.; Martin, Ralph P.; Hestenes, Roberta; Gillespie, Thomas W.; Cooley, Robert E.; Smith, Inez T., and Ryder, Janice E.
- Subjects
- Christian leadership., Leadership chrétien.
- Abstract
Theology News & Notes was a theological journal published for Fuller Theological Seminary alumni/ae from 1954 through 2014. The digital scans of Theology News & Notes were made possible by Fuller Studio and the Fuller Seminary Archives.
- Published
- 1997
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