SummaryNitrogen metabolism of Mycobacterium tuberculosis (Mtb) is crucial for the survival of this important pathogen in its primary human host cell, the macrophage, but little is known about the source(s) and their assimilation within this intracellular niche. Here, we have developed 15N-flux spectral ratio analysis (15N-FSRA) to explore Mtb’s nitrogen metabolism; we demonstrate that intracellular Mtb has access to multiple amino acids in the macrophage, including glutamate, glutamine, aspartate, alanine, glycine, and valine; and we identify glutamine as the predominant nitrogen donor. Each nitrogen source is uniquely assimilated into specific amino acid pools, indicating compartmentalized metabolism during intracellular growth. We have discovered that serine is not available to intracellular Mtb, and we show that a serine auxotroph is attenuated in macrophages. This work provides a systems-based tool for exploring the nitrogen metabolism of intracellular pathogens and highlights the enzyme phosphoserine transaminase as an attractive target for the development of novel anti-tuberculosis therapies.

Single quasicrystals of Al?oPd2iMn9 were plastically deformed by 25% under compression at 750°C. Both these samples as well as control samples of as-grown and heat-treated material were investigated by transmission electron microscopy. It was found that upon deformation the dislocation density increases by about 2 orders of magnitude. This provides the first direct evidence for quasicrystal deformation by a dislocation mechanism. The dislocations were found to have twofold and fivefold Burgers vector directions in physical space.PACS numbers: 61.44.+p, 6L66.Dk, 61.72.Bb, 62.20.Fe In a number of ternary aluminum-transition metal alloy systems thermodynamically stable quasicrystalline phases occur which can be produced with high structural perfection [l]. Of these alloys Al-Pd-Mn is particularly attractive. It contains an icosahedral phase of F-type lattice structure, of which single quasicrystals can be grown by the Czochralski technique [2,3]. After the first observation of lattice defects in the form of dislocations in decagonal AI65CU20C015 and icosahedral Al65Cu2oFei5, similar observations were reported for other stable quasicrystalline phases [4][5][6][7][8][9]. These observations raised the question of the origin of the dislocations, in particular, whether they can be induced by plastic deformation. Indeed hardness and compression tests demonstrated a relatively high ductility which increases with temperature [10][11][12][13][14][15]. With the exception of a hardness test at room temperature [10], these studies were all carried out in polyquasicrystalline material. We are reporting here on the first high-temperature single quasicrystal deformation experiments. Transmission electron microscopy of deformed and undeformed single quasicrystals of Al7 0 Pd2r Mn9 indicates an increase in dislocation density by about 2 orders of magnitude after plastic deformation, providing the first direct evidence for quasicrystal deformation by a dislocation mechanism.A master alloy of composition Al7 0 Pd2iMn9 was prepared in an induction furnace under an Ar atmosphere. Employing the Czochralski technique the resulting ingot was used to grow a single quasicrystal 7 cm in length and 1 cm in diameter of [0/0,0/0,0/2] orientation (notation of Cahn, Schechtman, and Gratias [16]). Small columns of 3x3x7 mm 3 were cut from this with their long axis parallel to [0/0,0/0,0/2]. Deformation under compression along this axis was performed at 750 °C in an INSTRON 1122 machine in air at a deformation velocity of 0.05 mm/min. A second sample was placed on the lower piston of the machine in order to serve as a reference. Because of a reduced length it was not deformed but went through the same temperature program as the deformation sample. In the following this sample is referred to as the heat-treated sample. After a deformation of 25% the load was released and the samples were quenched in water. Specimens for investigation in the transmission electron microscope (TEM) were prepared from the deformed and the heat-treated samples was well...

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Developmental neurotoxicity (DNT) may be induced when chemicals disturb a key neurodevelopmental process, and many tests focus on this type of toxicity. Alternatively, DNT may occur when chemicals are cytotoxic only during a specific neurodevelopmental stage. The toxicant sensitivity is affected by the expression of toxicant targets and by resilience factors. Although cellular metabolism plays an important role, little is known how it changes during human neurogenesis, and how potential alterations affect toxicant sensitivity of mature vs. immature neurons. We used immature (d0) and mature (d6) LUHMES cells (dopaminergic human neurons) to provide initial answers to these questions. Transcriptome profiling and characterization of energy metabolism suggested a switch from predominantly glycolytic energy generation to a more pronounced contribution of the tricarboxylic acid cycle (TCA) during neuronal maturation. Therefore, we used pulsed stable isotope-resolved metabolomics (pSIRM) to determine intracellular metabolite pool sizes (concentrations), and isotopically non-stationary C-metabolic flux analysis (INSTC-MFA) to calculate metabolic fluxes. We found that d0 cells mainly use glutamine to fuel the TCA. Furthermore, they rely on extracellular pyruvate to allow continuous growth. This metabolic situation does not allow for mitochondrial or glycolytic spare capacity, i.e. the ability to adapt energy generation to altered needs. Accordingly, neuronal precursor cells displayed a higher sensitivity to several mitochondrial toxicants than mature neurons differentiated from them. In summary, this study shows that precursor cells lose their glutamine dependency during differentiation while they gain flexibility of energy generation and thereby increase their resistance to low concentrations of mitochondrial toxicants.

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