Regulation of human argininosuccinate synthetase gene: Induction by positive-acting nuclear mechanism in canavanine-resistant cell variants

Nonhepatic human cell variants resistant to the arginine analog, canavanine, express argininosuccinate synthetase (AS) mRNA at levels 200-fold higher than parental cells without amplification of AS gene sequences. In this report we show that this regulation occurs in the nucleus prior to polyadenylation of AS precursor RNA and occurs through a positive-acting mechanism operating in canavanine-resistant cells. The half-life of cytoplasmic AS mRNA was estimated by blocking cellular transcription with actinomycin D. In both parental and canavanine-resistant variants of RPMI 2650 cells, the AS mRNA decayed with a half-life of 12–24 h, showing that cytoplasmic mRNA stabilization was not involved in this regulation. Quantification of AS RNA following cell fractionation showed that AS precursor RNA was present at greatly elevated amounts in the nuclei of canavanine-resistant cells. Similar results were obtained when nonpolyadenylated RNA was examined. Thus, the mechanism underlying high expression of AS mRNA in canavanine-resistant cells is an early nuclear event, and the processes of polyadenylation and transport of RNA to the cytoplasm are not involved. Intraspecific somatic cell hybrids were constructed to test whether the induction of AS mRNA was due to a gain of a function in canavanine-resistant cells or to a loss of a function in parental cells. Quantification of AS mRNA in hybrid cell lines showed that such cells contained high levels similar to those found in the canavanine-resistant parent. These findings show that the induction of AS mRNA is due to a positive-acting mechanism operating in the nucleus of canavanine-resistant cells .Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/45537/1/11188_2005_Article_BF01535071.pd

CAG expansion affects the expression of mutant huntingtin in the Huntington's disease brain

AbstractA trinucleotide repeat (CAG) expansion in the huntingtin gene causes Huntington's disease (HD). In brain tissue from HD heterozygotes with adult onset and more clinically severe juvenile onset, where the largest expansions occur, a mutant protein of equivalent intensity to wild-type huntingtin was detected in cortical synaptosomes, indicating that a mutant species is synthesized and transported with the normal protein to nerve endings. The increased size of mutant huntingtin relative to the wild type was highly correlated with CAG repeat expansion, thereby linking an altered electrophoretic mobility of the mutant protein to its abnormal function. Mutant huntingtin appeared in gray and white matter with no difference in expression in affected regions. The mutant protein was broader than the wild type and in 6 of 11 juvenile cases resolved as a complex of bands, consistent with evidence at the DNA level for somatic mosaicism. Thus, HD pathogenesis results from a gain of function by an aberrant protein that is widely expressed in brain and is harmful only to some neurons

Changes in cortical and striatal neurons predict behavioral and electrophysiological abnormalities in a transgenic murine model of Huntington\u27s disease

Neurons in Huntington\u27s disease exhibit selective morphological and subcellular alterations in the striatum and cortex. The link between these neuronal changes and behavioral abnormalities is unclear. We investigated relationships between essential neuronal changes that predict motor impairment and possible involvement of the corticostriatal pathway in developing behavioral phenotypes. We therefore generated heterozygote mice expressing the N-terminal one-third of huntingtin with normal (CT18) or expanded (HD46, HD100) glutamine repeats. The HD mice exhibited motor deficits between 3 and 10 months. The age of onset depended on an expanded polyglutamine length; phenotype severity correlated with increasing age. Neuronal changes in the striatum (nuclear inclusions) preceded the onset of phenotype, whereas cortical changes, especially the accumulation of huntingtin in the nucleus and cytoplasm and the appearance of dysmorphic dendrites, predicted the onset and severity of behavioral deficits. Striatal neurons in the HD mice displayed altered responses to cortical stimulation and to activation by the excitotoxic agent NMDA. Application of NMDA increased intracellular Ca(2+) levels in HD100 neurons compared with wild-type neurons. Results suggest that motor deficits in Huntington\u27s disease arise from cumulative morphological and physiological changes in neurons that impair corticostriatal circuitry

A Unique Population of Ventral Tegmental Area Neurons Inhibits the Lateral Habenula to Promote Reward

Lateral habenula (LHb) neurons convey aversive and negative reward conditions through potent indirect inhibition of ventral tegmental area (VTA) dopaminergic neurons. While VTA dopaminergic neurons reciprocally project to the LHb, the electrophysiological properties and the behavioral consequences associated with selective manipulations of this circuit are unknown. Here, we identify a novel inhibitory input to the LHb arising from a unique population of VTA neurons expressing dopaminergic markers. Optogenetic activation of this circuit resulted in no detectable dopamine release in LHb brain slices. Instead, stimulation produced GABA-mediated inhibitory synaptic transmission, which suppressed the firing of postsynaptic LHb neurons in brain slices and increased the spontaneous firing rate of VTA dopaminergic neurons in vivo. Furthermore, in vivo activation of this pathway produced reward-related phenotypes that were dependent on intra-LHb GABAA receptor signaling. These results suggest that non-canonical inhibitory signaling by these hybrid dopaminergic-GABAergic neurons act to suppress LHb output under rewarding conditions

The Flagellum of Pseudomonas aeruginosa Is Required for Resistance to Clearance by Surfactant Protein A

Surfactant protein A (SP-A) is an important lung innate immune protein that kills microbial pathogens by opsonization and membrane permeabilization. We investigated the basis of SP-A-mediated pulmonary clearance of Pseudomonas aeruginosa using genetically-engineered SP-A mice and a library of signature-tagged P. aeruginosa mutants. A mutant with an insertion into flgE, the gene that encodes flagellar hook protein, was preferentially cleared by the SP-A(+/+) mice, but survived in the SP-A(-/-) mice. Opsonization by SP-A did not play a role in flgE clearance. However, exposure to SP-A directly permeabilized and killed the flgE mutant, but not the wild-type parental strain. P. aeruginosa strains with mutation in other flagellar genes, as well as mucoid, nonmotile isolates from cystic fibrosis patients, were also permeabilized by SP-A. Provision of the wild-type fliC gene restored the resistance to SP-A-mediated membrane permeabilization in the fliC-deficient bacteria. In addition, non-mucoid, motile revertants of CF isolates reacquired resistance to SP-A-mediated membrane permeability. Resistance to SP-A was dependent on the presence of an intact flagellar structure, and independent of flagellar-dependent motility. We provide evidence that flagellar-deficient mutants harbor inadequate amounts of LPS required to resist membrane permeabilization by SP-A and cellular lysis by detergent targeting bacterial outer membranes. Thus, the flagellum of P. aeruginosa plays an indirect but important role resisting SP-A-mediated clearance and membrane permeabilization

Lex Maritima in a changing world: development and prospect of rules governing carriage of goods by sea

This chapter examines the attempts to unifying law governing carriage of goods by sea and the background to these attempts over the past hundred years or so. It finds that a repetition of the current mode of negotiating static conventions will not unify these rules. Moreover, from historic and legal perspectives, the attempts to unify the international carriage of goods by sea regimes in the past century have remained transitional. The active players have shifted from private entrepreneurs to government delegates. This research probes into the new trade practice for the shipping industry in the twenty-first century and argues that new ‘landscape’ calls for innovative modifications of the conventional approach to unifying carriage of goods by sea rules. This research also forecasts the prospects of the Rotterdam Rules and discusses several countries’ current attitudes, including the UK, the Netherlands, Scandinavian countries and, particularly, the USA

Hitomi (ASTRO-H) X-ray Astronomy Satellite

The Hitomi (ASTRO-H) mission is the sixth Japanese x-ray astronomy satellite developed by a large international collaboration, including Japan, USA, Canada, and Europe. The mission aimed to provide the highest energy resolution ever achieved at E  >  2  keV, using a microcalorimeter instrument, and to cover a wide energy range spanning four decades in energy from soft x-rays to gamma rays. After a successful launch on February 17, 2016, the spacecraft lost its function on March 26, 2016, but the commissioning phase for about a month provided valuable information on the onboard instruments and the spacecraft system, including astrophysical results obtained from first light observations. The paper describes the Hitomi (ASTRO-H) mission, its capabilities, the initial operation, and the instruments/spacecraft performances confirmed during the commissioning operations for about a month

The James Webb Space Telescope Mission

Twenty-six years ago a small committee report, building on earlier studies, expounded a compelling and poetic vision for the future of astronomy, calling for an infrared-optimized space telescope with an aperture of at least $4m$. With the support of their governments in the US, Europe, and Canada, 20,000 people realized that vision as the $6.5m$ James Webb Space Telescope. A generation of astronomers will celebrate their accomplishments for the life of the mission, potentially as long as 20 years, and beyond. This report and the scientific discoveries that follow are extended thank-you notes to the 20,000 team members. The telescope is working perfectly, with much better image quality than expected. In this and accompanying papers, we give a brief history, describe the observatory, outline its objectives and current observing program, and discuss the inventions and people who made it possible. We cite detailed reports on the design and the measured performance on orbit.Comment: Accepted by PASP for the special issue on The James Webb Space Telescope Overview, 29 pages, 4 figure

The Athena X-ray Integral Field Unit: a consolidated design for the system requirement review of the preliminary definition phase

The Athena X-ray Integral Unit (X-IFU) is the high resolution X-ray spectrometer studied since 2015 for flying in the mid-30s on the Athena space X-ray Observatory. Athena is a versatile observatory designed to address the Hot and Energetic Universe science theme, as selected in November 2013 by the Survey Science Committee. Based on a large format array of Transition Edge Sensors (TES), X-IFU aims to provide spatially resolved X-ray spectroscopy, with a spectral resolution of 2.5 eV (up to 7 keV) over a hexagonal field of view of 5 arc minutes (equivalent diameter). The X-IFU entered its System Requirement Review (SRR) in June 2022, at about the same time when ESA called for an overall X-IFU redesign (including the X-IFU cryostat and the cooling chain), due to an unanticipated cost overrun of Athena. In this paper, after illustrating the breakthrough capabilities of the X-IFU, we describe the instrument as presented at its SRR (i.e. in the course of its preliminary definition phase, so-called B1), browsing through all the subsystems and associated requirements. We then show the instrument budgets, with a particular emphasis on the anticipated budgets of some of its key performance parameters, such as the instrument efficiency, spectral resolution, energy scale knowledge, count rate capability, non X-ray background and target of opportunity efficiency. Finally, we briefly discuss the ongoing key technology demonstration activities, the calibration and the activities foreseen in the X-IFU Instrument Science Center, touch on communication and outreach activities, the consortium organisation and the life cycle assessment of X-IFU aiming at minimising the environmental footprint, associated with the development of the instrument. Thanks to the studies conducted so far on X-IFU, it is expected that along the design-to-cost exercise requested by ESA, the X-IFU will maintain flagship capabilities in spatially resolved high resolution X-ray spectroscopy, enabling most of the original X-IFU related scientific objectives of the Athena mission to be retained. The X-IFU will be provided by an international consortium led by France, The Netherlands and Italy, with ESA member state contributions from Belgium, Czech Republic, Finland, Germany, Poland, Spain, Switzerland, with additional contributions from the United States and Japan.The French contribution to X-IFU is funded by CNES, CNRS and CEA. This work has been also supported by ASI (Italian Space Agency) through the Contract 2019-27-HH.0, and by the ESA (European Space Agency) Core Technology Program (CTP) Contract No. 4000114932/15/NL/BW and the AREMBES - ESA CTP No.4000116655/16/NL/BW. This publication is part of grant RTI2018-096686-B-C21 funded by MCIN/AEI/10.13039/501100011033 and by “ERDF A way of making Europe”. This publication is part of grant RTI2018-096686-B-C21 and PID2020-115325GB-C31 funded by MCIN/AEI/10.13039/501100011033