Prolonged activation of S6K1 does not suppress IRS or PI-3 kinase signaling during muscle cell differentiation

Background: Myogenesis in C2C12 cells requires the activation of the PI3K/mTOR signaling pathways. Since mTOR signaling can feedback through S6K1 to inhibit the activation of PI3K, the aim of this work was to assess whether feedback from S6K1 played a role in myogenesis and determine whether siRNA mediated knockdown of S6K1 would lead to an increased rate of myotube formation. Results: S6K1 activity increased in a linear fashion following plating and was more than 3-fold higher after Day 3 of differentiation (subconfluent = 11.09 ± 3.05, Day 3 = 29.34 ± 3.58). IRS-1 levels tended to increase upon serum withdrawal but decreased approximately 2-fold (subconfluent = 0.88 ± 0.10, Day 3 = 0.42 ± 0.06) 3 days following differentiation whereas IRS-2 protein remained stable. IRS-1 associated p85 was significantly reduced upon serum withdrawal (subconfluent = 0.86 ± 0.07, Day 0 = 0.31 ± 0.05), remaining low through day 1. IRS-2 associated p85 decreased following serum withdrawal (subconfluent = 0.96 ± 0.05, Day 1 = 0.56 ± 0.08) and remained suppressed up to Day 3 following differentiation (0.56 ± 0.05). Phospho-tyrosine associated p85 increased significantly from subconfluent to Day 0 and remained elevated throughout differentiation. siRNA directed against S6K1 and S6K2 did not result in changes in IRS-1 levels after either 48 or 96 hrs. Furthermore, neither 48 nor 96 hrs of S6K1 knockdown caused a change in myotube formation. Conclusions: Even though S6K1 activity increases throughout muscle cell differentiation and IRS-1 levels decrease over this period, siRNA suggests that S6K1 is not mediating the decrease in IRS-1. The decrease in IRS-1/2 associated p85 together with the increase in phospho-tyrosine associated p85 suggests that PI3K associates primarily with scaffolds other than IRS-1/2 during muscle cell differentiation

Evaluating the role of a galanin enhancer genotype on a range of metabolic, depressive and addictive phenotypes

Funded by •ERC. Grant Number: 284167 •NIH. Grant Number: 1RO1DK0921127-01 •NWO. Grant Numbers: 463-06-001, 451-04-034Peer reviewedPublisher PD

Molecular brakes regulating mTORC1 activation in skeletal muscle following synergist ablation

The goal of the current work was to profile positive (mTORC1 activation, autocrine/paracrine growth factors) and negative [AMPK, unfolded protein response (UPR)] pathways that might regulate overload-induced mTORC1 (mTOR complex 1) activation with the hypothesis that a number of negative regulators of mTORC1 will be engaged during a supraphysiological model of hypertrophy. To achieve this, mTORC1- IRS-1/2 signaling, BiP/CHOP/IRE1, and AMPK activation were determined in rat plantaris muscle following synergist ablation (SA). SA resulted in significant increases in muscle mass of 4% per day throughout the 21 days of the experiment. The expression of the insulin-like growth factors (IGF) were high throughout the 21st day of overload. However, IGF signaling was limited, since IRS-1 and -2 were undetectable in the overloaded muscle from day 3 to day 9. The decreases in IRS-1/2 protein were paralleled by increases in GRB10 Ser501/503 and S6K1 Thr389 phosphorylation, two mTORC1 targets that can destabilize IRS proteins. PKB Ser473 phosphorylation was higher from 3– 6 days, and this was associated with increased TSC2 Thr939 phosphorylation. The phosphorylation of TSC2 Thr1345 (an AMPK site) was also elevated, whereas phosphorylation at the other PKB site, Thr1462, was unchanged at 6 days. In agreement with the phosphorylation of Thr1345, SA led to activation of AMPK1 during the initial growth phase, lasting the first 9 days before returning to baseline by day 12. The UPR markers CHOP and BiP were elevated over the first 12 days following ablation, whereas IRE1 levels decreased. These data suggest that during supraphysiological muscle loading at least three potential molecular brakes engage to downregulate mTORC1. m

Glycogen Content Regulates Peroxisome Proliferator Activated Receptor-∂ (PPAR-∂) Activity in Rat Skeletal Muscle

Performing exercise in a glycogen depleted state increases skeletal muscle lipid utilization and the transcription of genes regulating mitochondrial β-oxidation. Potential candidates for glycogen-mediated metabolic adaptation are the peroxisome proliferator activated receptor (PPAR) coactivator-1α (PGC-1α) and the transcription factor/nuclear receptor PPAR-∂. It was therefore the aim of the present study to examine whether acute exercise with or without glycogen manipulation affects PGC-1α and PPAR-∂ function in rodent skeletal muscle. Twenty female Wistar rats were randomly assigned to 5 experimental groups (n = 4): control [CON]; normal glycogen control [NG-C]; normal glycogen exercise [NG-E]; low glycogen control [LG-C]; and low glycogen exercise [LG-E]). Gastrocnemius (GTN) muscles were collected immediately following exercise and analyzed for glycogen content, PPAR-∂ activity via chromatin immunoprecipitation (ChIP) assays, AMPK α1/α2 kinase activity, and the localization of AMPK and PGC-1α. Exercise reduced muscle glycogen by 47 and 75% relative to CON in the NG-E and LG-E groups, respectively. Exercise that started with low glycogen (LG-E) finished with higher AMPK-α2 activity (147%, p<0.05), nuclear AMPK-α2 and PGC-1α, but no difference in AMPK-α1 activity compared to CON. In addition, PPAR-∂ binding to the CPT1 promoter was significantly increased only in the LG-E group. Finally, cell reporter studies in contracting C2C12 myotubes indicated that PPAR-∂ activity following contraction is sensitive to glucose availability, providing mechanistic insight into the association between PPAR-∂ and glycogen content/substrate availability. The present study is the first to examine PPAR-∂ activity in skeletal muscle in response to an acute bout of endurance exercise. Our data would suggest that a factor associated with muscle contraction and/or glycogen depletion activates PPAR-∂ and initiates AMPK translocation in skeletal muscle in response to exercise

A Limited Role for PI(3,4,5)P-3 Regulation in Controlling Skeletal Muscle Mass in Response to Resistance Exercise

Background: Since activation of the PI3K/(protein kinase B; PKB/akt) pathway has been shown to alter muscle mass and growth, the aim of this study was to determine whether resistance exercise increased insulin like growth factor (IGF) I/ phosphoinositide 3-kinase (PI3K) signalling and whether altering PI(3,4,5)P3 metabolism genetically would increase load induced muscle growth. Methodology/Principal Findings: Acute and chronic resistance exercise in wild type and muscle specific PTEN knockout mice were used to address the role of PI(3,4,5)P3 regulation in the development of skeletal muscle hypertrophy. Acute resistance exercise did not increase either IGF-1 receptor phosphorylation or IRS1/2 associated p85. Since insulin/IGF signalling to PI3K was unchanged, we next sought to determine whether inactivation of PTEN played a role in load-induced muscle growth. Muscle specific knockout of PTEN resulted in small but significant increases in heart (PTEN+/+ = 5.00&plusmn;0.02 mg/g, PTEN-/- = 5.50&plusmn;0.09 mg/g), and TA (PTEN+/+ = 1.74&plusmn;0.04 mg/g, PTEN-/- = 1.89 &plusmn;0.03) muscle mass, while the GTN, SOL, EDL and PLN remain unchanged. Following ablation, hypertrophy of the PLN, SOL or EDL muscles was similar between PTEN-/- and PTEN+/+ animals. Even though there were some changes in overload-induced PKB and S6K1 phosphorylation, 1 hr following acute resistance exercise there was no difference in the phosphorylation state of S6K1 Thr389 between genotypes. Conclusions/Significance: These data suggest that physiological loading does not lead to the enhanced activation of the PI3K/PKB/mTORC1 axis and that neither PI3K activation nor PTEN, and by extension PI(3,4,5)P3 levels, play a significant role in adult skeletal muscle growth

More about orbitally excited hadrons from lattice QCD

This is a second paper describing the calculation of spectroscopy for orbitally excited states from lattice simulations of Quantum Chromodynamics. New features include higher statistics for P-wave systems and first results for the spectroscopy of D-wave mesons and baryons, for relatively heavy quark masses. We parameterize the Coulomb gauge wave functions for P-wave and D-wave systems and compare them to those of their corresponding S-wave states.Comment: 21 pages plus 14 figs, 3 include

Immunostimulatory Membrane Proteins Potentiate \u3cem\u3eH. pylori-\u3c/em\u3eInduced Carcinogenesis by Enabling CagA Translocation

Infection with Helicobacter pylori is the single greatest risk factor for developing gastric adenocarcinoma. In prospective, population-based studies, seropositivity to the uncharacterized H. pylori proteins Hp0305 and Hp1564 was significantly associated with cancer risk in East Asia. However, the mechanism underlying this observation has not been elucidated. Here, we show that Hp0305 and Hp1564 act in concert with previously ascribed H. pylori virulence mechanisms to orchestrate cellular alterations that promote gastric carcinogenesis. In samples from 546 patients exhibiting premalignant gastric lesions, seropositivity to Hp0305 and Hp1564 was significantly associated with increased gastric atrophy across all stomach conditions. In vitro, depletion of Hp0305 and Hp1564 significantly reduced levels of gastric cell-associated bacteria and markedly impaired the ability of H. pylori to stimulate pro-inflammatory cytokine production. Remarkably, our studies revealed that Hp1564 is required for translocation of the oncoprotein CagA into gastric epithelial cells. Our data provide experimental insight into the molecular mechanisms governing novel H. pylori pathogenicity factors that are strongly associated with gastric disease and highlight the potential of Hp0305 and Hp1564 as robust molecular tools that can improve identification of individuals that are highly susceptible to gastric cancer. We demonstrate that Hp0305 and Hp1564 augment H. pylori-mediated inflammation and gastric cancer risk by promoting key bacteria-gastric cell interactions that facilitate delivery of oncogenic microbial cargo to target cells. Thus, therapeutically targeting microbial interactions driven by Hp0305/Hp1564 may enable focused H. pylori eradication strategies to prevent development of gastric malignancies in high-risk populations

Constituting monetary conservatives via the 'savings habit': New Labour and the British housing market bubble

The ongoing world credit crunch might well kill off the most recent bubble dynamics in the British housing market by driving prices systematically downwards from their 2007 peak. Nonetheless, the experience of that bubble still warrants analytical attention. The Labour Government might not have been responsible for consciously creating it, but it has certainly grasped the opportunities the bubble has provided in an attempt to enforce a process of agential change at the heart of the British economy. The key issue in this respect is the way in which the Government has challenged the legitimacy of passive welfare receipts in favour of establishing a welfare system based on incorporating the individual into an active asset-holding society. The housing market has taken on new political significance as a means for individuals first to acquire assets and then to accumulate wealth on the back of asset ownership. The ensuing integration of the housing market into an increasingly reconfigured welfare system has permeated into the politics of everyday life. It has been consistent with individuals remaking their political subjectivities in line with preferences for the type of conservative monetary policies that typically keep house price bubbles inflated

Hierarchical Hough all-sky search for periodic gravitational waves in LIGO S5 data

We describe a new pipeline used to analyze the data from the fifth science run (S5) of the LIGO detectors to search for continuous gravitational waves from isolated spinning neutron stars. The method employed is based on the Hough transform, which is a semi-coherent, computationally efficient, and robust pattern recognition technique. The Hough transform is used to find signals in the time-frequency plane of the data whose frequency evolution fits the pattern produced by the Doppler shift imposed on the signal by the Earth's motion and the pulsar's spin-down during the observation period. The main differences with respect to previous Hough all-sky searches are described. These differences include the use of a two-step hierarchical Hough search, analysis of coincidences among the candidates produced in the first and second year of S5, and veto strategies based on a $\chi^2$ test.Comment: 7 pages, 2 figures, Amaldi08 proceedings, submitted to JPC

Chemotaxis: a feedback-based computational model robustly predicts multiple aspects of real cell behaviour

The mechanism of eukaryotic chemotaxis remains unclear despite intensive study. The most frequently described mechanism acts through attractants causing actin polymerization, in turn leading to pseudopod formation and cell movement. We recently proposed an alternative mechanism, supported by several lines of data, in which pseudopods are made by a self-generated cycle. If chemoattractants are present, they modulate the cycle rather than directly causing actin polymerization. The aim of this work is to test the explanatory and predictive powers of such pseudopod-based models to predict the complex behaviour of cells in chemotaxis. We have now tested the effectiveness of this mechanism using a computational model of cell movement and chemotaxis based on pseudopod autocatalysis. The model reproduces a surprisingly wide range of existing data about cell movement and chemotaxis. It simulates cell polarization and persistence without stimuli and selection of accurate pseudopods when chemoattractant gradients are present. It predicts both bias of pseudopod position in low chemoattractant gradients and-unexpectedly-lateral pseudopod initiation in high gradients. To test the predictive ability of the model, we looked for untested and novel predictions. One prediction from the model is that the angle between successive pseudopods at the front of the cell will increase in proportion to the difference between the cell's direction and the direction of the gradient. We measured the angles between pseudopods in chemotaxing Dictyostelium cells under different conditions and found the results agreed with the model extremely well. Our model and data together suggest that in rapidly moving cells like Dictyostelium and neutrophils an intrinsic pseudopod cycle lies at the heart of cell motility. This implies that the mechanism behind chemotaxis relies on modification of intrinsic pseudopod behaviour, more than generation of new pseudopods or actin polymerization by chemoattractant