Differential susceptibility of mitochondrial complex II to inhibition by oxaloacetate in brain and heart

Mitochondrial Complex II is a key mitochondrial enzyme connecting the tricarboxylic acid (TCA) cycle and the electron transport chain. Studies of complex II are clinically important since new roles for this enzyme have recently emerged in cell signalling, cancer biology, immune response and neurodegeneration. Oxaloacetate (OAA) is an intermediate of the TCA cycle and at the same time is an inhibitor of complex II with high affinity (Kd ~ 10− 8 M). Whether or not OAA inhibition of complex II is a physiologically relevant process is a significant, but still controversial topic. We found that complex II from mouse heart and brain tissue has similar affinity to OAA and that only a fraction of the enzyme in isolated mitochondrial membranes (30.2 ± 6.0% and 56.4 ± 5.6% in the heart and brain, respectively) is in the free, active form. Since OAA could bind to complex II during isolation, we established a novel approach to deplete OAA in the homogenates at the early stages of isolation. In heart, this treatment significantly increased the fraction of free enzyme, indicating that OAA binds to complex II during isolation. In brain the OAA-depleting system did not significantly change the amount of free enzyme, indicating that a large fraction of complex II is already in the OAA-bound inactive form. Furthermore, short-term ischemia resulted in a dramatic decline of OAA in tissues, but it did not change the amount of free complex II. Our data show that in brain OAA is an endogenous effector of complex II, potentially capable of modulating the activity of the enzyme.

Molecular Mechanism of the Response to Hydrogen Sulfide in C. elegans

Thesis (Ph.D.)--University of Washington, 2016-08

Macrophage adaptation leads to parallel evolution of genetically diverseEscherichia colismall-colony variants with increased fitness in vivo and antibiotic collateral sensitivity

Small-colony variants (SCVs) are commonly observed in evolution experiments and clinical isolates, being associated with antibiotic resistance and persistent infections. We recently observed the repeated emergence of Escherichia coli SCVs during adaptation to the interaction with macrophages. To identify the genetic targets underlying the emergence of this clinically relevant morphotype, we performed whole-genome sequencing of independently evolved SCV clones. We uncovered novel mutational targets, not previously associated with SCVs (e.g. cydA, pepP) and observed widespread functional parallelism. All SCV clones had mutations in genes related to the electron-transport chain. As SCVs emerged during adaptation to macrophages, and often show increased antibiotic resistance, we measured SCV fitness inside macrophages and measured their antibiotic resistance profiles. SCVs had a fitness advantage inside macrophages and showed increased aminoglycoside resistance in vitro, but had collateral sensitivity to other antibiotics (e.g. tetracycline). Importantly, we observed similar results in vivo. SCVs had a fitness advantage upon colonization of the mouse gut, which could be tuned by antibiotic treatment: kanamycin (aminoglycoside) increased SCV fitness, but tetracycline strongly reduced it. Our results highlight the power of using experimental evolution as the basis for identifying the causes and consequences of adaptation during host-microbe interactions.

Exploring complex chemical systems: insights from mass spectrometry, electrochemistry and continuous cell culture

The work presented herein covers a broad range of research topics and so, in the interest of clarity, has been presented in a portfolio format. Accordingly, each chapter consists of its own introductory material prior to presentation of the key results garnered, this is then proceeded by a short discussion on their significance. In the first chapter, a methodology to facilitate the resolution and qualitative assessment of very large inorganic polyoxometalates was designed and implemented employing ion-mobility mass spectrometry. Furthermore, the potential of this technique for ‘mapping’ the conformational space occupied by this class of materials was demonstrated. These claims are then substantiated by the development of a tuneable, polyoxometalate-based calibration protocol that provided the necessary platform for quantitative assessments of similarly large, but unknown, polyoxometalate species. In addition, whilst addressing a major limitation of travelling wave ion mobility, this result also highlighted the potential of this technique for solution-phase cluster discovery. The second chapter reports on the application of a biophotovoltaic electrochemical cell for characterising the electrogenic activity inherent to a number of mutant Synechocystis strains. The intention was to determine the key components in the photosynthetic electron transport chain responsible for extracellular electron transfer. This would help to address the significant lack of mechanistic understanding in this field. Finally, in the third chapter, the design and fabrication of a low-cost, highly modular, continuous cell culture system is presented. To demonstrate the advantages and suitability of this platform for experimental evolution investigations, an exploration into the photophysiological response to gradual iron limitation, in both the ancestral wild type and a randomly generated mutant library population, was undertaken. Furthermore, coupling random mutagenesis to continuous culture in this way is shown to constitute a novel source of genetic variation that is open to further investigation.

Ferramentas de apoio à implementação da sustentabilidade na gestão da cadeia de abastecimento

O presente trabalho tem como principal objetivo contribuir para o desenvolvimento de um modelo de gestão da cadeia de abastecimento sustentável, posteriormente operacionalizado num conjunto de empresas. O principal elemento diferenciador do modelo apresentado é o seu cariz operacional, focado na fase da implementação com a integração de um conjunto de práticas de apoio à sustentabilidade. Para responder às pressões da conjuntura económica atual, das alterações climáticas, da escassez de recursos e das desigualdades sociais é necessário desenvolver de forma consolidada e abrangente um novo paradigma de gestão nas empresas. Muitas destas pressões fazem-se sentir nas atividades da Gestão da Cadeia de Abastecimento. O grande desafio é conseguir que as empresas obtenham bons resultados económicos, sociais e ambientais. A sustentabilidade tem sido abordada como a área de estudo donde deverá emergir este novo paradigma de gestão. Atendendo a esta problemática, a principal questão de investigação do presente trabalho é “Como se implementa a Sustentabilidade na Gestão da Cadeia de Abastecimento?” A metodologia de investigação partiu da revisão da literatura que permitiu estruturar um conjunto de pressupostos teóricos, estruturados num modelo conceptual sobre a implementação da sustentabilidade na Gestão da Cadeia de Abastecimento. O modelo foi aplicado em dois grupos de estudos empíricos: Análise Qualitativa de Relatórios de Sustentabilidade publicados por seis empresas com atividade em Portugal (Sonae; Lipor, Galp; EDP; Portucel e AutoEuropa); e o desenvolvimento de dois Estudos de Caso nas empresas Bosch Termotecnologia e Gestamp Aveiro. Os resultados permitiram o desenvolvimento de um Modelo Teórico de Implementação da Sustentabilidade na Gestão da Cadeia de Abastecimento. Bem como, um modelo de classificação das ferramentas de apoio à implementação da sustentabilidade adequadas a cada etapa que constitui o modelo de implementação. No desenvolvimento deste trabalho, acreditou-se que o caminho da sustentabilidade é possível e tangível. Os modelos desenvolvidos explicam que a integração da sustentabilidade se enceta pela estruturação da área da sustentabilidade na organização, prosseguindo com o processo de implementação constituído por quatro etapas: Envolvimento, Execução, Monitorização e Comunicação. A implementação necessita de ser abrangente a toda a cadeia de valor e apoiada num conjunto de ferramentas adequadas a cada fase de implementação.

Life without complex I: proteome analyses of an Arabidopsis mutant lacking the mitochondrial NADH dehydrogenase complex

The mitochondrial NADH dehydrogenase complex (complex I) is of particular importance for the respiratory chain in mitochondria. It is the major electron entry site for the mitochondrial electron transport chain (mETC) and therefore of great significance for mitochondrial ATP generation. We recently described an Arabidopsis thaliana double-mutant lacking the genes encoding the carbonic anhydrases CA1 and CA2, which both form part of a plant-specific 'carbonic anhydrase domain' of mitochondrial complex I. The mutant lacks complex I completely. Here we report extended analyses for systematically characterizing the proteome of the ca1ca2 mutant. Using various proteomic tools, we show that lack of complex I causes reorganization of the cellular respiration system. Reduced electron entry into the respiratory chain at the first segment of the mETC leads to induction of complexes II and IV as well as alternative oxidase. Increased electron entry at later segments of the mETC requires an increase in oxidation of organic substrates. This is reflected by higher abundance of proteins involved in glycolysis, the tricarboxylic acid cycle and branched-chain amino acid catabolism. Proteins involved in the light reaction of photosynthesis, the Calvin cycle, tetrapyrrole biosynthesis, and photorespiration are clearly reduced, contributing to the significant delay in growth and development of the double-mutant. Finally, enzymes involved in defense against reactive oxygen species and stress symptoms are much induced. These together with previously reported insights into the function of plant complex I, which were obtained by analysing other complex I mutants, are integrated in order to comprehensively describe 'life without complex I'.

Modern trafik – En studie kring cykelfrämjandet i Umeå kommun

Purpose: To determine the progress in Umeå concerning the furtherance of bicycle traffic in the city and to investigate the possibility of a similar progress in Jönköping. Method: Semi structured interviews with officials, politicians and enthusiasts in Umeå and Jönköping, document analysis of documents provided by the municipalities and observations of Umeå city center. Findings: Umeå’s mindset around a sustainable society have been identified and the vast difference between the two cities is the time that they’ve actively worked with the bike issue and how public officials prioritize while planning. Umeå’s large advantage in their decision making is that officials and politicians are working towards the same goal and are united in the bike issue. This makes decisions regarding bicycle promotion easier to make. Due to the difference in conditions between the cities, the writers states that Umeå’s way of planning their traffic not necessarily is the right way for Jönköping to plan theirs. Implications: In order to develop it’s bicycle network, Jönköping must dare to let go of their old way of thinking. Instead of thinking that car traffic is necessary in a city, they should have the courage to take difficult decisions that benefit bicycle promotion and thus develop a sustainable transport system. Limitations: The study does not address the economic aspects that are involved in the development of an attractive bicycle network. Also the environmental issues are only dealt with in a thorough level, as it’s necessary to fulfill the study’s objectives. The study’s purpose is considered to be achieved but the result could have been even more developed if more parties were interviewed. Keywords: Bicycle path, bicycle, sustainable transport planning, sustainable society, sustainability, Umeå, Jönköping 

The role of urban consolidation centres in sustainable freight transport

The chapter considers the role that urban consolidation centres (UCCs) can play in helping to reduce goods vehicle traffic and its environmental impacts in urban areas. A UCC is a logistics facility that is situated in relatively close proximity to the area that it serves. Goods destined for this area are dropped off at the UCC, and are sorted and consolidated onto goods vehicles for delivery to their final destinations, often using environmentally friendly vehicles such as electric and gas-powered goods vehicles, and electrically-assisted cycles. The development of UCCs since their first appearance in the 1970s is reviewed and the various types of UCC categorised. The freight transport and logistics advantages offered by UCCs are discussed. Case studies of four recent UCC trials are included. The objectives, operational and financial aspects and impacts of these UCCs are compared and their critical success factors identified.

Developing a sustainability assessment model : the sustainable infrastructure, land-use, environment and transport model

Measuring the comparative sustainability levels of cities, regions, institutions and projects is an essential procedure in creating sustainable urban futures. This paper introduces a new urban sustainability assessment model: “The Sustainable Infrastructure, Land-use, Environment and Transport Model (SILENT)”. The SILENT Model is an advanced geographic information system and indicator-based comparative urban sustainability indexing model. The model aims to assist planners and policy makers in their daily tasks in sustainable urban planning and development by providing an integrated sustainability assessment framework. The paper gives an overview of the conceptual framework and components of the model and discusses the theoretical constructs, methodological procedures, and future development of this promising urban sustainability assessment model.

Sustainable urban and transport development for transportation disadvantaged : a review

Around the world, particularly in North America and Australia, urban sprawl combined with low density suburban development has caused serious accessibility and mobility problems, especially for those who do not own a motor vehicle or have access to public transportation services. Sustainable urban and transportation development is seen crucial in solving transportation disadvantage problems in urban settlements. However, current urban and transportation models have not been adequately addressed unsustainable urban transportation problems that transportation disadvantaged groups overwhelmingly encounter, and the negative impacts on the disadvantaged have not been effectively considered. Transportation disadvantaged is a multi-dimensional problem that combines demographic, spatial and transportation service dimensions. Nevertheless, most transportation models focusing on transportation disadvantage only employ demographic and transportation service dimensions and do not take spatial dimension into account. This paper aims to investigate the link between sustainable urban and transportation development and spatial dimension of the transportation disadvantage problem. The paper, for that purpose, provides a thorough review of the literature and identifies a set of urban, development and policy characteristics to define spatial dimension of the transportation disadvantage problem. This paper presents an overview of these urban, development and policy characteristics that have significant relationships with sustainable urban and transportation development and travel inability, which are also useful in determining transportation disadvantaged populations.

Developing sustainable housing through building utilities and transport innovation

Sustainability concerns every citizen. Housing affordability and sustainable solutions are being highlighted in research and practice in many parts of the world. This paper discusses the development of a Commuter Energy and Building Utilities System (CEBUS) in sustainable housing projects as a means of bridging the gap between current median house pricing and target affordable house pricing for low income earners. Similar scales of sustainable housing development cannot be achieved through independent application of current best practice methods in ecologically sustainable development strategies or transit oriented development master plans. This paper presents the initial stage of research on first capital and ongoing utilities and transport cost savings available from these sustainable design methods. It also outlines further research and development of a CEBUS Dynamic Simulation Model and Conceptual Framework for the Australian property development and construction industry.

Sustainable housing: family experiences with supply chain ethics

Climate change, resource depletion and increasing urbanization are converging global issues that are challenging the way we design, construct and operate buildings. The housing sector is a significant contributor to these global issues through consumption of limited resources, waste generation and disposal (solid, liquid and atmospheric waste) and negative human health impacts (Senick 2006). Although the design and construction of ‘sustainable housing’ would appear to be an obvious and technically feasible solution, there remains multi-faceted issues affecting the delivery of sustainable housing (Holloway and Bunker 2006). Two fundamental issues - what makes a house sustainable, and to what extent regulation should be used to deliver sustainability - have been, and continue to be, debated at multiple levels in society. Despite personal, professional and political views on these issues, three key characteristics of the whole housing supply chain require fundamental change if we are to successfully address sustainability challenges (Birkeland 2008). These include: fragmentation; established methods, practices and processes, and the relationships between players. A more in-depth understanding of the role of ethics (values, beliefs and standards) and potential ethical conflicts within the supply chain will assist in better defining the nature of the fundamental changes required...

Corporate social and environmental responsibility in relation to the agricultural sector and the food supply chain : a Case Study of Waitrose

Waitrose has a strong commitment to organic farming but also uses products from 'conventional' farms. At the production stage, Waitrose own-label products are fully traceable, GM-free and all suppliers undergo a detailed assessment programme based on current best practice. Crop suppliers to Waitrose operate an authenticity programme to certify that each assignment is GM-free and produce is screened for pesticide residues. Waitrose sources conventional crops grown from 'Integrated Crop Management Systems' (ICMS) using best horticultural practices. The 'Assured Product' scheme regulates all UK produce to ICMS standards and these audits are being extended worldwide. Business is withdrawn from suppliers who fail the audit. In relation to this, Waitrose has increased its Fairtrade range as in its view 'Buying these products provides direct additional benefit to workers in the developing countries where they are produced and assists marginal producers by giving them access to markets they would not otherwise have'. Currently, Waitrose is developing its own sustainable timber assessment criteria. For livestock, protocols are in place to ensure that animals are reared under the 'most natural conditions possible' and free range produce is offered where animals have access to open space although some produce is not from free-range animals. Waitrose also use a 'Hazards Analysis Critical Points' system to identify food safety hazards that occur at any stage from production to point of sale and to ensure that full measures are in place to control them. In addition, mechanisms have been implemented to reduce fuel use and hence reduce CO2 emissions in the transport of products and staff, and to increase the energy use efficiency of refrigeration systems which account for approximately 60% of Waitrose energy use.

Sustainable, safe, smart—three key elements of Singapore’s evolving transport policies

Sustainability, safety and smartness are three key elements of a modern transportation system. This study illustrates various policy directions and initiatives of Singapore to address how its transportation system is progressing in light of these three components. Sustainability targets economical efficiency, environmental justice and social equity by including policies for integrating land use and transport planning, ensuring adequate transport supply measures, managing travel demand efficiently, and incorporating environment-friendly strategies. Safety initiatives of its transportation system aim to minimize injuries and incidents of all users including motorists, public transport commuters, pedestrians, and bicyclists. Smartness incorporates qualities like real time sensing, fast processing and decision making, and automated action-taking into its control, monitoring, information management and revenue collection systems. Various policy implications and technology applications along these three directions reveal that smart technologies facilitate implementation of policies promoting sustainability and safety. The Singapore experience could serve as a good reference for other cities in promoting a transportation system that is sustainable, safe and smart.

BIM for sustainable whole-of-life transport infrastructure asset management

The US National Institute of Standards and Technology (NIST) showed that, in 2004, owners and operations managers bore two thirds of the total industry cost burden from inadequate interoperability in construction projects from inception to operation, amounting to USD10.6 billion. Building Information Modelling (BIM) and similar tools were identified by Engineers Australia in 2005 as potential instruments to significantly reduce this sum, which in Australia could amount to total industry-wide cost burden of AUD12 billion. Public sector road authorities in Australia have a key responsibility in driving initiatives to reduce greenhouse gas emissions from the construction and operations of transport infrastructure. However, as previous research has shown the Environmental Impact Assessment process, typically used for project approvals and permitting based on project designs available at the consent stage, lacks Key Performance Indicators (KPIs) that include long-term impact factors and transfer of information throughout the project life cycle. In the building construction industry, BIM is widely used to model sustainability KPIs such as energy consumption, and integrated with facility management systems. This paper proposes that a similar use of BIM in early design phases of transport infrastructure could provide: (i) productivity gains through improved interoperability and documentation; (ii) the opportunity to carry out detailed cost-benefit analyses leading to significant operational cost savings; (iii) coordinated planning of street and highway lighting with other energy and environmental considerations; iv) measurable KPIs that include long-term impact factors which are transferable throughout the project life cycle; and (v) the opportunity for integrating design documentation with sustainability whole-of-life targets.