Direct anabolic metabolism of three-carbon propionate to a six-carbon metabolite occurs in vivo across tissues and species

Anabolic metabolism of carbon in mammals is mediated via the one- and two-carbon carriers S-adenosyl methionine and acetyl-coenzyme A. In contrast, anabolic metabolism of three-carbon units via propionate has not been shown to extensively occur. Mammals are primarily thought to oxidize the three-carbon short chain fatty acid propionate by shunting propionyl-CoA to succinyl-CoA for entry into the TCA cycle. Here, we found that this may not be absolute as, in mammals, one nonoxidative fate of propionyl-CoA is to condense to two three-carbon units into a six-carbon trans-2-methyl-2-pentenoyl-CoA (2M2PE-CoA). We confirmed this reaction pathway using purified protein extracts provided limited substrates and verified the product via LC-MS using a synthetic standard. In whole-body in vivo stable isotope tracing following infusion of ¹³C-labeled valine at steady state, 2M2PE-CoA was found to form via propionyl-CoA in multiple murine tissues, including heart, kidney, and to a lesser degree, in brown adipose tissue, liver, and tibialis anterior muscle. Using ex vivo isotope tracing, we found that 2M2PE-CoA also formed in human myocardial tissue incubated with propionate to a limited extent. While the complete enzymology of this pathway remains to be elucidated, these results confirm the in vivo existence of at least one anabolic three- to six-carbon reaction conserved in humans and mice that utilizes propionate.     

Ethnic mobilization and the state: the Roma in Eastern Europe

The political mobilization of marginalized ethnic groups is a process indispensable for the realization of their political objectives. This article identifies the key criteria and conditions that promote and impede successful ethnic mobilization and analyses the determinants of state policy towards newly mobilizing ethnic minorities. The theoretical arguments receive empirical support from the findings of a comparative study of the Romani (Gypsy) minority in seven East European states.     

Cyclic Nucleotide Dependent Dephosphorylation of Regulator of G-Protein Signaling 18 in Human Platelets

Regulator of G-protein signaling 18 (RGS18) is a GTPase-activating protein that turns off Gq signaling in platelets. RGS18 is regulated by binding to the adaptor protein 14-3-3 via phosphorylated serine residues S49 and S218 on RGS18. In this study we confirm that thrombin, thromboxane A2, or ADP stimulate the interaction of RGS18 and 14-3-3 by increasing the phosphorylation of S49. Cyclic AMP- and cyclic GMP-dependent kinases (PKA, PKG) inhibit the interaction of RGS18 and 14-3-3 by phosphorylating S216. To understand the effect of S216 phosphorylation we studied the phosphorylation kinetics of S49, S216, and S218 using Phos-tag gels and phosphorylation site-specific antibodies in transfected cells and in platelets. Cyclic nucleotide-induced detachment of 14-3-3 from RGS18 coincides initially with double phosphorylation of S216 and S218. This is followed by dephosphorylation of S49 and S218. Dephosphorylation of S49 and S218 might be mediated by protein phosphatase 1 (PP1) which is linked to RGS18 by the regulatory subunit PPP1R9B (spinophilin). We conclude that PKA and PKG induced S216 phosphorylation triggers the dephosphorylation of the 14-3-3 binding sites of RGS18 in platelets.     

Quantitative Analysis of the Whole-Body Metabolic Fate of Branched-Chain Amino Acids

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Identification and characterization of carbapenem binding sites within the RND-transporter AcrB

Understanding the molecular determinants for recognition, binding and transport of antibiotics by multidrug efflux systems is important for basic research and useful for the design of more effective antimicrobial compounds. Imipenem and meropenem are two carbapenems whose antibacterial activity is known to be poorly and strongly affected by MexAB-OprM, the major efflux pump transporter in Pseudomonas aeruginosa. However, not much is known regarding recognition and transport of these compounds by AcrAB-TolC, which is the MexAB-OprM homologue in Escherichia coli and by definition the paradigm model for structural studies on efflux pumps. Prompted by this motivation, we unveiled the molecular details of the interaction of imipenem and meropenem with the transporter AcrB by combining computer simulations with biophysical experiments. Regarding the interaction with the two main substrate binding regions of AcrB, the so-called access and deep binding pockets, molecular dynamics simulations revealed imipenem to be more mobile than meropenem in the former, while comparable mobilities were observed in the latter. This result is in line with isothermal titration calorimetry, differential scanning experiments, and binding free energy calculations, indicating a higher affinity for meropenem than imipenem at the deep binding pocket, while both sharing similar affinities at the access pocket. Our findings rationalize how different physico-chemical properties of compounds reflect on their interactions with AcrB. As such, they constitute precious information to be exploited for the rational design of antibiotics able to evade efflux pumps.     

The Early-Onset Myocardial Infarction Associated PHACTR1 Gene Regulates Skeletal and Cardiac Alpha-Actin Gene Expression

The phosphatase and actin regulator 1 (PHACTR1) locus is a very commonly identified hit in genome-wide association studies investigating coronary artery disease and myocardial infarction (MI). However, the function of PHACTR1 in the heart is still unknown. We characterized the mechanisms regulating Phactr1 expression in the heart, used adenoviral gene delivery to investigate the effects of Phactr1 on cardiac function, and analyzed the relationship between MI associated PHACTR1 allele and cardiac function in human subjects. Phactr1 mRNA and protein levels were markedly reduced (60%, P<0.01 and 90%, P<0.001, respectively) at 1 day after MI in rats. When the direct myocardial effects of Phactr1 were studied, the skeletal α-actin to cardiac α-actin isoform ratio was significantly higher (1.5-fold, P<0.05) at 3 days but 40% lower (P<0.05) at 2 weeks after adenovirus-mediated Phactr1 gene delivery into the anterior wall of the left ventricle. Similarly, the skeletal α-actin to cardiac α-actin ratio was lower at 2 weeks in infarcted hearts overexpressing Phactr1. In cultured neonatal cardiac myocytes, adenovirus-mediated Phactr1 overexpression for 48 hours markedly increased the skeletal α-actin to cardiac α-actin ratio, this being associated with an enhanced DNA binding activity of serum response factor. Phactr1 overexpression exerted no major effects on the expression of other cardiac genes or LV structure and function in normal and infarcted hearts during 2 weeks’ follow-up period. In human subjects, MI associated PHACTR1 allele was not associated significantly with cardiac function (n = 1550). Phactr1 seems to regulate the skeletal to cardiac α-actin isoform ratio.     

A Quantitative and Dynamic Model of the Arabidopsis Flowering Time Gene Regulatory Network

Various environmental signals integrate into a network of floral regulatory genes leading to the final decision on when to flower. Although a wealth of qualitative knowledge is available on how flowering time genes regulate each other, only a few studies incorporated this knowledge into predictive models. Such models are invaluable as they enable to investigate how various types of inputs are combined to give a quantitative readout. To investigate the effect of gene expression disturbances on flowering time, we developed a dynamic model for the regulation of flowering time in Arabidopsis thaliana. Model parameters were estimated based on expression time-courses for relevant genes, and a consistent set of flowering times for plants of various genetic backgrounds. Validation was performed by predicting changes in expression level in mutant backgrounds and comparing these predictions with independent expression data, and by comparison of predicted and experimental flowering times for several double mutants. Remarkably, the model predicts that a disturbance in a particular gene has not necessarily the largest impact on directly connected genes. For example, the model predicts that SUPPRESSOR OF OVEREXPRESSION OF CONSTANS (SOC1) mutation has a larger impact on APETALA1 (AP1), which is not directly regulated by SOC1, compared to its effect on LEAFY (LFY) which is under direct control of SOC1. This was confirmed by expression data. Another model prediction involves the importance of cooperativity in the regulation of APETALA1 (AP1) by LFY, a prediction supported by experimental evidence. Concluding, our model for flowering time gene regulation enables to address how different quantitative inputs are combined into one quantitative output, flowering time.     

Quelques remarques sur l'holisme dans la pensée grecque

Summary The conception of the world with Greek thinkers generally differs from contemporary thought in the idea of the limitation of the world whose essence manifests itself to speculative thinking. The cosmology which culminates inPlato's Timaios attributes to the whole of being the properties of finiteness, uniqueness, life and perfection.Plato's holism subsumes inanimate and living nature under the idea of cosmic life, the parts of which are constituted by life-phenomena given in experience. His world-conception, constructed from within and intuitively, which ranges phenomena in descending order, is opposed to the atomism ofDemocritos which reduces the higher structures to the one equal material of all things. This opposition manifests itself in all essential points: toDemocritos there are worlds in infinite number, only accumulations of atoms are real, uniqueness and perfection are lacking as well as the idea of Life as a central reality.Modern research foregoes the attempt to determine the whole speculatively, but holistic and atomistic ways of thinking, after having been dogmatically applied by the Greeks, continue to exist as opposed types of mental attitude. Contemporary holism may be empirical and limited in application, but it is directed by the same intuition of life articulating itself organically as wasPlato's. It would be incorrect, however, to characterize holism as synthetical, since all synthetics is based upon analysis. The bounds of synthesis are never fixed ones, nor does it ever determine essence. Holism's way of subsuming single life-phenomena under more comprehensive wholes is an intuitional one, the connections which it creates between phenomena are not external ones, but apprehended intuitively in analogy to inner experience.—In distinction to atomism, which only admits as real atoms and their mutual relations, holism poses groups of unities, classified in grades, the higher of which dominate the lower, without debasing them to elements. As metaphysics is concerned to discover a connection between Nature and human life, it must be highly interested in the empirical demonstration, such as holism offers, of connections of life, surpassing individual being.

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Zusammenfassung Die griechische Weltauffassung unterscheidet sich van der modernen durch den Gedanken der Begrenztheit des Weltalls, das sich dem spekulativen Blick seinem Wesen nach erschliesst. Die inPlaton's Timaios gipfelnde Kosmologie bestimmt das Weltall nach den Wesensgesichtspunkten der Endlichkeit, der Einzigkeit, der Lebendigkeit und der Vollkommenheit.Platon's Holismus bezieht die leblose und die lebende Natur unter die Idee des Lebens des Kosmos, dessen Teile die in der Erfahrung gegebenen Lebenserscheinungen sind. Sein von innen her und intuitiv aufgebautes Weltbild, das die Erscheinungen von oben herab hierarchisch ordnet, tritt dem AtomismusDemokrits gegenüber, der in umgekehrtem Verfahren die höheren Strukturen auf den einen Baustoff aller Dinge reduziert. Der Gegensatz zum Holismus offenbart sich in allen Bestimmungen: fürDemokrit sind der Kosmoi unbegrenzt viele, es gibt nur Ansammlungen von Atomen, es fehlen Einzigkeit und Vollkommenheit, sowie ein Begriff des Lebens als des zentralen Geschehens.Die neuzeitliche Forschung verzichtet auf die spekulativen Bestimmungen des Ganzen, aber der bei den Griechen dogmatische Gegensatz holistischer und atomistischer Betrachtungsweise lebt fort als Gegensatz geistiger Einstellung. Der neuzeitliche Holismus ist ein empirischer, beschränkter, aber bei ihm liegt dieselbe Intuition des sich organisch gestaltenden Gesamtlebens zu Grunde wei beiPlaton. Nur wäre es, unrichtig, diesen Holismus als synthetisch zu bezeichnen. Synthetisches Denken erhebt sich immer auf analytischem Boden. Jede Synthese hat eine offene Grenze. Die Art aber, wie der Holismus die vereinzelten Lebenserscheinungen auf umfassendere Ganzheiten bezieht, ist eine intuitive. Das Band der Erscheinungen wird nicht durch äusseres Beziehen gestiftet, sondern geschaut nach Analogie einer inneren Erfahrung.Im Gegensatz zum Atomismus, der als real nur die Atome und deren Verhältnisse gelten lässt, erkennt der Holismus stufenartig aufgebaute Einheitsarten an, in denen die höheren die niedrigeren beherrschen, ohne sie zu Elementen herabzudrücken. Die Metaphysik, auf der Suche nach dem Zusammenhang zwischen dem Menschenleben und der Natur ist an der empirischen Aufweisung das Einzelne übersteigender Lebensverbände wie sie der Holismus zu Tage fordert aufs Höchste interessiert.

     

Terahertz Selective Emission Enhancement from a Metasurface-Coupled Photoconductive Emitter in Quasi-Near-Field Zone

We present a THz emission enhancement of 41 times at 0.92 THz from a metasurface made of T-shaped resonators excited in a quasi-near-field zone. Such a metasurface has an intrinsic transmission minimum with Q factor of 4 at 1.25 THz under far-field excitation. When this metasurface is coupled onto the backside of a 625-μm-thick photoconductive emitter, the metasurface is below the Fraunhofer distance to the excitation source. As such, one broad enhancement around 0.47 THz and another extremely narrow enhancement at 0.92 THz in the emission spectrum are observed owing to a quasi-near-field excitation. Theoretically, the Q factor of the latter is up to 307, which is limited by the spectral resolution in experiment. The numerical simulations indicate that the T-shaped resonators serve as an array of plasmonic antennas resulting in the aforementioned emission enhancement of THz radiation.

     

Notch Is a Critical Component of the Mouse Somitogenesis Oscillator and Is Essential for the Formation of the Somites

Segmentation of the vertebrate body axis is initiated through somitogenesis, whereby epithelial somites bud off in pairs periodically from the rostral end of the unsegmented presomitic mesoderm (PSM). The periodicity of somitogenesis is governed by a molecular oscillator that drives periodic waves of clock gene expression caudo-rostrally through the PSM with a periodicity that matches somite formation. To date the clock genes comprise components of the Notch, Wnt, and FGF pathways. The literature contains controversial reports as to the absolute role(s) of Notch signalling during the process of somite formation. Recent data in the zebrafish have suggested that the only role of Notch signalling is to synchronise clock gene oscillations across the PSM and that somite formation can continue in the absence of Notch activity. However, it is not clear in the mouse if an FGF/Wnt-based oscillator is sufficient to generate segmented structures, such as the somites, in the absence of all Notch activity. We have investigated the requirement for Notch signalling in the mouse somitogenesis clock by analysing embryos carrying a mutation in different components of the Notch pathway, such as Lunatic fringe (Lfng), Hes7, Rbpj, and presenilin1/presenilin2 (Psen1/Psen2), and by pharmacological blocking of the Notch pathway. In contrast to the fish studies, we show that mouse embryos lacking all Notch activity do not show oscillatory activity, as evidenced by the absence of waves of clock gene expression across the PSM, and they do not develop somites. We propose that, at least in the mouse embryo, Notch activity is absolutely essential for the formation of a segmented body axis.