Identification and characterization of a lysosomal transporter for small neutral amino acids

In eukaryotic cells, lysosomes represent a major site for macromolecule degradation. Hydrolysis products are eventually exported from this acidic organelle into the cytosol through specific transporters. Impairment of this process at either the hydrolysis or the efflux step is responsible of several lysosomal storage diseases. However, most lysosomal transporters, although biochemically characterized, remain unknown at the molecular level. In this study, we report the molecular and functional characterization of a lysosomal amino acid transporter (LYAAT-1), remotely related to a family of H+-coupled plasma membrane and synaptic vesicle amino acid transporters. LYAAT-1 is expressed in most rat tissues, with highest levels in the brain where it is present in neurons. Upon overexpression in COS-7 cells, the recombinant protein mediates the accumulation of neutral amino acids, such as γ-aminobutyric acid, l-alanine, and l-proline, through an H+/amino acid symport. Confocal microscopy on brain sections revealed that this transporter colocalizes with cathepsin D, an established lysosomal marker. LYAAT-1 thus appears as a lysosomal transporter that actively exports neutral amino acids from lysosomes by chemiosmotic coupling to the H+-ATPase of these organelles. Homology searching in eukaryotic genomes suggests that LYAAT-1 defines a subgroup of lysosomal transporters in the amino acid/auxin permease family.

Autoantibodies to calpastatin (an endogenous inhibitor for calcium-dependent neutral protease, calpain) in systemic rheumatic diseases.

We identified an autoantibody that reacts with calpastatin [an inhibitor protein of the calcium-dependent neutral protease calpain (EC 3.4.22.17)]. In early immunoblot studies, sera from patients with rheumatoid arthritis (RA) recognized unidentified 60-, 45-, and 75-kDa proteins in HeLa cell extracts. To identify these autoantigens, we used patient sera to clone cDNAs from a lambda gt11 expression library. We isolated clones of four genes that expressed fusion proteins recognized by RA sera. The 1.2-kb cDNA insert (termed RA-6) appeared to encode a polypeptide corresponding to the 60-kDa antigen from HeLa cells, since antibodies bound to the RA-6 fusion protein also reacted with a 60-kDa HeLa protein. The deduced amino acid sequence of the RA-6 cDNA was completely identical with the C-terminal 178 amino acids of human calpastatin except for one amino acid substitution. Patient sera that reacted with the RA-6 also bound pig muscle calpastatin, and a monoclonal antibody to human calpastatin recognized the RA-6 fusion protein, confirming the identity of RA-6 with calpastatin. Moreover, the purified RA-6 fusion protein inhibited the proteolytic activity of calpain, and IgG from a serum containing anti-calpastatin antibodies blocked the calpastatin activity of the RA-6 fusion protein. Immunoblots of the RA-6 product detected autoantibodies to calpastatin in 57% of RA patients; this incidence was significantly higher than that observed in other systemic rheumatic diseases, including systemic lupus erythematosus (27%), polymyositis/dermatomyositis (24%), systemic sclerosis (38%), and overlap syndrome (29%). Thus, anti-calpastatin antibodies are present most frequently in patients with RA and may participate in pathogenic mechanisms of rheumatic diseases.

Tecnologia construtiva de fachada em chapas delgadas estruturadas em light steel framing.

Aumento da produtividade, melhorias na qualidade dos produtos, redução de custos e de impactos ambientais são essenciais para a capacidade competitiva das empresas. A execução da fachada faz parte do caminho crítico da obra, por ser um subsistema que associa as funções de fechamento, acabamento, iluminação e ventilação e ainda por incorporar sistemas prediais; apresenta, por isso também, um alto custo direto em relação aos outros subsistemas do edifício. A tecnologia construtiva de fachadas em chapas delgadas com estrutura em Light Steel Framing (LSF) é uma alternativa viável para aumentar a produtividade e reduzir os prazos de obra, com qualidade e desempenho, e pode trazer benefícios em relação a atividades intensas em mão de obra como é o caso da alvenaria de vedação e de seus revestimentos. O presente trabalho tem por objetivo sistematizar e analisar o conhecimento relativo a essa tecnologia construtiva de fachada. O método adotado compreende revisão bibliográfica. Como contribuição, o trabalho reúne um conjunto organizado de informações sobre os principais sistemas disponíveis no mercado contemplando: a caracterização do sistema de fachada, de suas camadas e dos perfis leves de aço e a sistematização das principais avaliações técnicas de sistemas existentes em outros países, reunindo normas técnicas de produtos e de execução. Acredita-se que a reunião e organização das informações, antes dispersas em diversas referências, têm potencial para subsidiar o meio técnico para tomada de decisão quanto ao uso adequado da nova tecnologia.

Stability switches in a first-order complex neutral delay equation

Stability of the first-order neutral delay equation x’ (t) + ax’ (t – τ) = bx(t) + cx(t – τ) with complex coefficients is studied, by analyzing the existence of stability switches.

Nitrate reduction at Pt(1 0 0) single crystals and preferentially oriented nanoparticles in neutral media

The electroreduction of nitrate on Pt(1 0 0) electrodes in phosphate buffer neutral solution, pH 7.2, is reported. The sensitivity of the reaction to the crystallographic order of the surface is studied through the controlled introduction of defects by using stepped surfaces with (1 0 0) terraces of different length separated by monoatomic steps, either with (1 1 1) or (1 1 0) symmetry. The results of this study show that nitrate reduction occurs mainly on the well defined (1 0 0) terraces in the potential region where H adsorption starts to decrease, allowing the nitrate anion to access the surface. Adsorbed NO has been detected as a stable intermediate in this media. An oxidation process observed at 0.8 V has been identified as leading to the formation of adsorbed NO and being responsible for a secondary reduction process observed in the subsequent negative scan. Using in situ FTIRS, ammonium was found to be the main product of nitrate reduction. This species can be oxidized at high potentials resulting in adsorbed NO and nitrate (probably with nitrite as intermediate).

Exploring the interfacial neutral pH region of Pt(111) electrodes

The interfacial properties of Pt(111) single crystal electrodes have been investigated in the pH range 3 < pH < 5 in order to obtain information about the acidity of electrosorbed water. Proper experimental conditions are defined to avoid local pH changes while maintaining the absence of specifically adsorbed anions and preserving the cleanliness of the solution. For this purpose, buffer solutions resulting from mixtures of NaF and HClO4 are used. Total charge curves are obtained at different pHs from the integration of the voltammetric currents in combination with CO charge displacement experiments. Analysis of the composition of the interphase as a function of the pH provides information for the understanding of the notion of interfacial pH.

#DressCodePM & Re-Framing The Niqab: News Sources, Hashtag Activism, and Media Representation

This thesis originates from my interest in exploring how minorities are using social media to talk back to mainstream media. This study examines whether hashtags that trend on Twitter may impact how news stories related to minorities are covered in Canadian media. The Canadian Prime Minister Stephen Harper stated the niqab was “rooted in a culture that is anti-women” on 10 March 2015. The next day #DressCodePM trended in response to the PM’s niqab remarks. Using network gatekeeping theory, this study examines the types of sources quoted in the media stories published on 10 and 11 March 2015. The study’s goal is to explore whether using tweet quotes leads to the representation of a more diverse range of news sources. The study compares the types of sources quoted in stories that covered Harper’s comments without mentioning #DressCodePM versus stories that mention #DressCodePM. This study also uses Tuen A. van Dijk’s methodology of asking “who is speaking, how often and how prominently?” in order to examine whose voices have been privileged and whose voices have been marginalized in covering the niqab in Canadian media from the 1970s and until the days following the PM’s remarks. Network gatekeeping theory is applied in this study to assess whether the gated gained more power after #DressCodePM trended. The case study’s findings indicates that Caucasian male politicians were predominantly used as news sources in covering stories related to the niqab for the past 38 years in the Globe and Mail. The sourcing pattern of favouring politicians continued in Canadian print and online media on 10 March 2015 following Harper’s niqab comments. However, ordinary Canadian women, including Muslim women, were used more often than politicians as news sources in the stories about #DressCodePM that were published on 11 March 2015. The gated media users were able to gain power and attract Canadian Media’s attention by widely spreading #DressCodePM. This study draws attention to the lack of diversity of sources used in Canadian political news stories, yet this study also shows it is possible for the gated media users to amplify their voices through hashtag activism.

Developing an Analytical Framework on Social Cohesion in Singapore. Reflections from the framing of social cohesion debates in the OECD and Europe. EU Centre Singapore Working Paper No. 17, November 2013

Official discourse in Singapore on social cohesion is often framed along the broad parameters of achieving racial and religious harmony. Many policies – formal and informal – and several laws evolved to manage these two aspects of society. Yet, as Singapore developed and with a much more complex socioeconomic environment both domestically and externally, there is perhaps a need to re-look the discourse and framework for discussing social cohesion. This paper takes a critical look at how the issue of social cohesion is framed in academic literature and policy discussions in Europe and the OECD, and tries to develop a broader analytical framework that could be useful in the Singapore context as it struggles with the multiple fault lines in society (beyond race and religion) that have emerged in the last decade or so.

Framing Banking Union in the Euro Area: Some empirical evidence. CEPS Working Document No. 389, 3 February 2014

Evidence shows that financial integration in the euro area is retrenching at a quicker pace than outside the union. Home bias persists: Governments compete on funding costs by supporting ‘their’ banks with massive state aids, which distorts the playing field and feeds the risk-aversion loop. This situation intensifies friction in credit markets, thus hampering the transmission of monetary policies and, potentially, economic growth. This paper discusses the theoretical foundations of a banking union in a common currency area and the legal and economic aspects of EU responses. As a result, two remedies are proposed to deal with moral hazard in a common currency area: a common (unlimited) financial backstop to a privately funded recapitalisation/resolution fund and a blanket prohibition on state aids.