The stress polarity signaling (SPS) pathway serves as a marker and a target in the leaky gut barrier: implications in aging and cancer.

The gut barrier separates trillions of microbes from the largest immune system in the body; when compromised, a "leaky" gut barrier fuels systemic inflammation, which hastens the progression of chronic diseases. Strategies to detect and repair the leaky gut barrier remain urgent and unmet needs. Recently, a stress-polarity signaling (SPS) pathway has been described in which the metabolic sensor, AMP-kinase acts via its effector, GIV (also known as Girdin) to augment epithelial polarity exclusively under energetic stress and suppresses tumor formation. Using murine and human colon-derived organoids, and enteroid-derived monolayers (EDMs) that are exposed to stressors, we reveal that the SPS-pathway is active in the intestinal epithelium and requires a catalytically active AMP-kinase. Its pharmacologic augmentation resists stress-induced collapse of the epithelium when challenged with microbes or microbial products. In addition, the SPS-pathway is suppressed in the aging gut, and its reactivation in enteroid-derived monolayers reverses aging-associated inflammation and loss of barrier function. It is also silenced during progression of colorectal cancers. These findings reveal the importance of the SPS-pathway in the gut and highlights its therapeutic potential for treating gut barrier dysfunction in aging, cancer, and dysbiosis

AMPK alpha 1-induced RhoA phosphorylation mediates vasoprotective effect of estradiol

OBJECTIVE: Estradiol (E2) mediates numerous beneficial effects assigned to estrogens, but whereas mechanisms have been described at the endothelial level, direct effects on vascular smooth muscle cells (VSMC) are poorly documented. As evidence accumulates regarding the role of RhoA in vascular pathophysiology and the benefit of RhoA-Rho associated protein kinase (Rock) pathway inhibition, we analyzed if E2 could inhibit it in VSMC. METHODS AND RESULTS: We show that in VSMC, E2 inhibits the RhoA-Rock pathway in a time- and concentration-dependent manner. The inhibition of RhoA-Rock pathway results from E2-induced phosphorylation of the Ser188 of RhoA. Using pharmacological, transfection, and in vitro phosphorylation experiments, we demonstrate that AMP-activated protein kinase subunit alpha 1 (AMPKalpha1) is activated by estrogen receptor stimulation and catalyzes RhoA phosphorylation induced by E2. Ex vivo, ovariectomy leads to an increase in the amplitude of phenylephrine- or serotonine-induced contractions of aortic rings in wild-type mice but not in AMPKalpha1-knock-out mice or E2-supplemented animals. These functional effects were correlated with a reduced level of RhoA phosphorylation in the aorta of ovariectomized female, male, and AMPKalpha1 knock-out mice. CONCLUSION: Our work thus defines AMPKalpha1 as (1) a new kinase for RhoA and (2) a new mediator of the vasoprotective effects of estrogen

A <i>Tbc1d1</i>^Ser231Ala-knockin mutation partially impairs AICAR- but not exercise-induced muscle glucose uptake in mice

Aims/hypothesis: TBC1D1 (tre-2/USP6, BUB2, cdc16 domain family member 1) is a Rab GTPase-activating protein (RabGAP) that has been implicated in regulating GLUT4 trafficking. TBC1D1 can be phosphorylated by the AMP-activated protein kinase (AMPK) on Ser(231), which consequently interacts with 14-3-3 proteins. Given the key role for AMPK in regulating insulin-independent muscle glucose uptake, we hypothesised that TBC1D1-Ser(231) phosphorylation and/or 14-3-3 binding may mediate AMPK-governed glucose homeostasis.Methods: Whole-body glucose homeostasis and muscle glucose uptake were assayed in mice bearing a Tbc1d1 (Ser231Ala)-knockin mutation or harbouring skeletal muscle-specific Ampkα1/α2 (also known as Prkaa1/2) double-knockout mutations in response to an AMPK-activating agent, 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR). Exercise-induced muscle glucose uptake and exercise capacity were also determined in the Tbc1d1 (Ser231Ala)-knockin mice.Results: Skeletal muscle-specific deletion of Ampkα1/a2 in mice prevented AICAR-induced hypoglycaemia and muscle glucose uptake. The Tbc1d1 (Ser231Ala)-knockin mutation also attenuated the glucose-lowering effect of AICAR in mice. Glucose uptake and cell surface GLUT4 content were significantly lower in muscle isolated from the Tbc1d1 (Ser231Ala)-knockin mice upon stimulation with a submaximal dose of AICAR. However, this Tbc1d1 (Ser231Ala)-knockin mutation neither impaired exercise-induced muscle glucose uptake nor affected exercise capacity in mice.Conclusions/interpretation: TBC1D1-Ser(231) phosphorylation and/or 14-3-3 binding partially mediates AMPK-governed glucose homeostasis and muscle glucose uptake in a context-dependent manner.</p

Basal Autophagy Induction Without AMP-Activated Protein Kinase Under Low Glucose Conditions

When ATP levels in a cell decrease, various homeostatic intracellular mechanisms initiate attempts to restore ATP levels. As a prominent energy sensor, AMP-activated protein kinase (AMPK) represents one molecular gauge that links energy levels to regulation of anabolic and catabolic processes to restore energy balance. Although pharmacological studies have suggested that an AMPK activator, AIC AR (5-aminoimidazole-4-carboxamide ribonucleoside) may link AMPK activation to autophagy, a process that can provide short-term energy within the cell, AICAR can have AMPK-independent effects. Therefore, using a genetic-based approach we investigated the role of AMPK in cellular energy balance. We demonstrate that genetically altered cells, mouse embryonic fibroblasts (MEFs), lacking functional AMPK, display altered energy balance under basal conditions and die prematurely under low glucose-serum starvation challenge. These AMPK mutant cells appear to be abnormally reliant on autophagy under low glucose basal conditions, and therefore cannot rely further on autophagy like wild-type cells during further energetic stress and instead undergo apoptosis. This data suggests that AMPK helps regulate basal energy levels under low glucose. Further, AMPK mutant cells show increased basal phosphorylation of p53 at serine 15, a residue phosphorylated under glucose deprivation. We propose that cells lacking AMPK function have altered p53 activity that may help sensitize these cells to apoptosis under energetic stress

T4 RNA Ligase 2 truncated active site mutants: improved tools for RNA analysis

<p>Abstract</p> <p>Background</p> <p>T4 RNA ligases 1 and 2 are useful tools for RNA analysis. Their use upstream of RNA analyses such as high-throughput RNA sequencing and microarrays has recently increased their importance. The truncated form of T4 RNA ligase 2, comprising amino acids 1-249 (T4 Rnl2tr), is an attractive tool for attachment of adapters or labels to RNA 3'-ends. Compared to T4 RNA ligase 1, T4 Rnl2tr has a decreased ability to ligate 5'-PO₄ends in single-stranded RNA ligations, and compared to the full-length T4 Rnl2, the T4 Rnl2tr has an increased activity for joining 5'-adenylated adapters to RNA 3'-ends. The combination of these properties allows adapter attachment to RNA 3'-ends with reduced circularization and concatemerization of substrate RNA.</p> <p>Results</p> <p>With the aim of further reducing unwanted side ligation products, we substituted active site residues, known to be important for adenylyltransferase steps of the ligation reaction, in the context of T4 Rnl2tr. We characterized the variant ligases for the formation of unwanted ligation side products and for activity in the strand-joining reaction.</p> <p>Conclusions</p> <p>Our data demonstrate that lysine 227 is a key residue facilitating adenylyl transfer from adenylated ligation donor substrates to the ligase. This reversal of the second step of the ligation reaction correlates with the formation of unwanted ligation products. Thus, T4 Rn2tr mutants containing the K227Q mutation are useful for reducing undesired ligation products. We furthermore report optimal conditions for the use of these improved T4 Rnl2tr variants.</p

Oval Domes: History, Geometry and Mechanics

An oval dome may be defined as a dome whose plan or profile (or both) has an oval form. The word Aoval@ comes from the latin Aovum@, egg. Then, an oval dome has an egg-shaped geometry. The first buildings with oval plans were built without a predetermined form, just trying to close an space in the most economical form. Eventually, the geometry was defined by using arcs of circle with common tangents in the points of change of curvature. Later the oval acquired a more regular form with two axis of symmetry. Therefore, an “oval” may be defined as an egg-shaped form, doubly symmetric, constructed with arcs of circle; an oval needs a minimum of four centres, but it is possible also to build polycentric ovals. The above definition corresponds with the origin and the use of oval forms in building and may be applied without problem until, say, the XVIIIth century. Since then, the teaching of conics in the elementary courses of geometry made the cultivated people to define the oval as an approximation to the ellipse, an “imperfect ellipse”: an oval was, then, a curve formed with arcs of circles which tries to approximate to the ellipse of the same axes. As we shall see, the ellipse has very rarely been used in building. Finally, in modern geometrical textbooks an oval is defined as a smooth closed convex curve, a more general definition which embraces the two previous, but which is of no particular use in the study of the employment of oval forms in building. The present paper contains the following parts: 1) an outline the origin and application of the oval in historical architecture; 2) a discussion of the spatial geometry of oval domes, i. e., the different methods employed to trace them; 3) a brief exposition of the mechanics of oval arches and domes; and 4) a final discussion of the role of Geometry in oval arch and dome design

AMPK:a nutrient and energy sensor that maintains energy homeostasis

AMP-activated protein kinase (AMPK) is a crucial cellular energy sensor. Once activated by falling energy status, it promotes ATP production by increasing the activity or expression of proteins involved in catabolism while conserving ATP by switching off biosynthetic pathways. AMPK also regulates metabolic energy balance at the whole-body level. For example, it mediates the effects of agents acting on the hypothalamus that promote feeding and entrains circadian rhythms of metabolism and feeding behaviour. Finally, recent studies reveal that AMPK conserves ATP levels through the regulation of processes other than metabolism, such as the cell cycle and neuronal membrane excitability

Perceptions of key stakeholders regarding national federation sport policy: The case of the French Rugby Union

© 2017 Informa UK Limited, trading as Taylor & Francis Group. This article proposes a framework that explains how a national association sport policy is operationally formalized in relation with the various perceptions of its internal stakeholders. This is an important issue, as a lack of engagement from these stakeholders may result in a limited impact of such a sport policy. Considering the latter as a management tool, a case study of the French Rugby Union (FFR) was built using archival material and interviews. We demonstrated that a national sport federation policy, when viewed as a management tool, is the consequence of a conceptual framework, consisting of a formal substrate, a managerial philosophy and a simplified representation of the actors’ role [Hatchuel, A., and Weil, B. (1992). L’expert et le système [The expert and the system]. Paris: Economica]. As part of an intervention-research within the FFR, 45 semi-structured interviews were conducted with both national and regional actors with various statutes. The analysis of these interviews shows both agreement and contradictions, categorized through the conceptual framework used to explain that a “one size fits all” approach should enable a degree of regional freedom for sport governing bodies that include organizational membership

Phosphatase and tensin homologue: a therapeutic target for SMA

Spinal muscular atrophy (SMA) is one of the most common juvenile neurodegenerative diseases, which can be associated with child mortality. SMA is caused by a mutation of ubiquitously expressed gene, Survival Motor Neuron1 (SMN1), leading to reduced SMN protein and the motor neuron death. The disease is incurable and the only therapeutic strategy to follow is to improve the expression of SMN protein levels in motor neurons. Significant numbers of motor neurons in SMA mice and SMA cultures are caspase positive with condensed nuclei, suggesting that these cells are prone to a process of cell death called apoptosis. Searching for other potential molecules or signaling pathways that are neuroprotective for central nervous system (CNS) insults is essential for widening the scope of developmental medicine. PTEN, a Phosphatase and Tensin homologue, is a tumor suppressor, which is widely expressed in CNS. PTEN depletion activates anti-apoptotic factors and it is evident that the pathway plays an important protective role in many neurodegenerative disorders. It functions as a negative regulator of PIP3/AKT pathway and thereby modulates its downstream cellular functions through lipid phosphatase activity. Moreover, previous reports from our group demonstrated that, PTEN depletion using viral vector delivery system in SMN delta7 mice reduces disease pathology, with significant rescue on survival rate and the body weight of the SMA mice. Thus knockdown/depletion/mutation of PTEN and manipulation of PTEN medicated Akt/PKB signaling pathway may represent an important therapeutic strategy to promote motor neuron survival in SMA