Strategies and good practice for sustainable and liveable cities of tomorrow

This contribution deals with the question, what makes cities sustainable and integrative, and suggests an approach for "liveable cities of tomorrow" designed to sustain mobility. The liveable city of tomorrow needs to meet both ecological and social requirements in an integrative approach. To design urban patterns appropriate or “sustainable mobility” based on a concept of mobility defined as the number of accessible destinations (different to that for “fossil mobility” defined as the ability to cover distances) is a key element of such an approach. Considering the limited reserves of fossil fuels and the long lifetime of the built structure, mobility needs to rely on modes independent of fossil fuels (public transport and pedestrians) to make it sustainable and the urban pattern needs to be developed appropriately for these modes. Crucial for the success of public transport is the location of buildings within the catchment area of stops. An attractive urban environment for pedestrians is characterised by short distances in a compact settlement with appropriate/qualified urban density and mixed land use as well as by attractive public space. This, complemented by an integrative urban development on the quarter level including neighbourhood management with a broad spectrum of activity areas (social infrastructure, integration of diverse social and ethnic groups, health promotion, community living, etc.), results in increased liveability. The role of information technology in this context is to support a sustainable use of the built structures by organisational instruments. Sustainable and liveable communities offer many benefits for health, safety and well-being of their inhabitants.

Comparative analysis of passenger transport sustainability in European cities

Sustainable development in its three dimensions – economic, social and environmental – has become a major concern on an international scale. The problem is global, but must be solved locally. Most of the world’s population lives in cities that act as centres of economic growth and productivity, but which – if they develop in the wrong direction – can cause social inequalities, or irreversibly harm the environment. Urban transport causes a number of negative impacts that can affect sustainability targets. The objective of this study is to propose an analysis of sustainability of urban passenger transport systems based on available indicators in most cities. This will serve to benchmark the practices of different cities and manage their transport systems. This work involves the creation of composite indicators (CI) to measure the sustainability of urban passenger transport systems. The methodology is applied to 23 European cities. The indicators are based on a benchmarking approach, and the evaluation of each aspect in each case therefore depends on the performance of the whole sample. The CI enabled us to identify which characteristics have the greatest influence on the sustainability of a city’s transport system, and to establish transport policies that could potentially improve its shortcomings. Finally, the cities are clustered according to the values obtained from the CIs, and thus according to the weaknesses and strengths of their transport systems.

Exploring users inter-temporal preferences for measuring transport social equity effects

There exist different ways for defining a welfare function. Traditionally, welfare economic theory foundation is based on the Net Present Value (NPV) calculation where the time dependent preferences of considered agents are taken into account. However, the time preferences, remains a controversial subject. Currently, the traditional approach employs a unique discount rate for various agents. Nevertheless, this way of discounting appears inconsistent with sustainable development. New research work suggests that the discount rate may not be a homogeneous value. The discount rates may change following the individual’s preferences. A significant body of evidence suggests that people do not behave following a constant discount rate. In fact, UK Government has quickly recognized the power of the arguments for time-varying rates, as it has done in its official guidance to Ministries on the appraisal of investments and policies. Other authors deal with not just time preference but with uncertainty about future income (precautionary saving). In a situation in which economic growth rates are similar across time periods, the rationale for declining social optimal discount rates is driven by the preferences of the individuals in the economy, rather than expectations of growth. However, these approaches have been mainly focused on long-term policies where intergenerational risks may appear. The traditional cost-benefit analysis (CBA) uses a unique discount rate derived from market interest rates or investment rates of return for discounting the costs and benefits of all social agents included in the CBA. However, recent literature showed that a more adequate measure of social benefit is possible by using different discount rates including inter-temporal preferences rate of users, private investment discount rate and intertemporal preferences rate of government. Actually, the costs of opportunity may differ amongst individuals, firms, governments, or society in general, as do the returns on savings. In general, the firms or operators require an investment rate linked to the current return on savings, while the discount rate of consumers-users depends on their time preferences with respect of the current and the future consumption, as well as society can take into account the intergenerational well-being, adopting a lower discount rate for today’s generation. Time discount rate of social actors (users, operators, government and society) places a lower value in a future gain, but the uncertainty about future income strongly determines the individual preferences. These time and uncertainty depends on preferences and should be integrated into a transport policy formulation that may have significant social impacts. The discount rate of a user cannot be the same than the operator’s discount rate. The preferences of both are different. In addition, another school of thought suggests that people, such as a social group, may have different attitudes towards future costs and benefits. Particularly, the users have different discount rates related to their income. Some research work tried to modify user discount rates using a compensating weight which represents the inverse of household income level. The inter-temporal preferences are a proxy of the willingness to pay during the time. Its consideration is important in order to make acceptable or not a policy or investment

Assessment methodology to make urban transport interchanges attractive for users

En las ciudades europeas, los patrones de movilidad son cada vez más complejos debido fundamentalmente a un crecimiento sostenido de la población así como a la tendencia de dispersión de los núcleos urbanos. En consecuencia, muchos de los usuarios del transporte público se ven obligados a combinar varios modos o servicios de transporte para completar sus viajes diarios. Por tanto, el mayor reto de las ciudades es conseguir una mejora e incremento en la movilidad mientras que al mismo tiempo se reducen problemas como la congestión, los accidentes y la contaminación (COM, 2006). Un principio básico para lograr una movilidad sostenible es reducir los inconvenientes y molestias derivados de la transferencia o ruptura del viaje. En este sentido, los intercambiadores de transporte público juegan un papel fundamental como nodos de la red urbana de transporte y la calidad del servicio prestado en ellos tiene una influencia directa sobre la experiencia diaria de los viajeros. Como señaló Terzis and Last (2002), un intercambiador de transportes urbano eficiente debe ser competitivo y al mismo tiempo, debe ser atractivo para los usuarios dado que sus experiencias físicas y sus reacciones psicológicas se ven influenciadas de manera significativa por el diseño y operación del intercambiador. Sin embargo, todavía no existen standards o normativas a nivel europeo que especifiquen como deberían ser estos intercambiadores. Esta tesis doctoral proporciona conocimientos y herramientas de análisis dirigidas a planificadores y gestores de los propios intercambiadores con el fin de entender mejor el funcionamiento de los intercambiadores y gestionar así los recursos disponibles. Así mismo, esta tesis identifica los factores clave en el diseño y operación de intercambiadores urbanos de transporte y proporciona algunas guías generales de planificación en base a ellos. Dado que las percepciones de los usuarios son particularmente importantes para definir políticas adecuadas para intercambiadores, se diseñó y se llevó a cabo en 2013 una encuesta de satisfacción al viajero en tres intercambiadores de transporte urbano europeos: Moncloa (Madrid, España), Kamppi (Helsinki, Finlandia) e Ilford Railway Station ( Londres, Reino Unido). En resumen, esta tesis pone de relieve la naturaleza ambivalente de los intercambiadores urbanos de transporte, es decir, como nodos de la red de transporte y como lugares en sí mismos donde los usuarios pasan tiempo dentro de ellos y propone algunas recomendaciones para hacer más atractivos los intercambiadores a los usuarios. Travel patterns in European urban areas are becoming increasingly complex due to a sustained increase in the urban population and the trend towards urban sprawl. Consequently, many public transport users need to combine several modes or transport services to complete their daily trips. Therefore, the challenge facing all major cities is how to increase mobility while at the same time reducing congestion, accididents and pollution (COM, 2006). Reducing the inconvenience inherent in transferring between modes is a basic principle for achieving sustainable mobility. In this regard, transport interchanges play a key role as urban transport network nodes, and the quality of the service provided in them has a direct influence on travellers' daily experience. As noted by Terzis and Last (2000), an efficient urban transport interchange must be competitive and, at the same time, be attractive for users given that their physical experiences and psychological reactions are significantly influenced by the design and operation of the interchange. However, yet there are no standards or regulations specifying the form these interchanges should take in Europe. This doctoral thesis provides knowledge and analysis tools addressed to developers and managers in order to understand better the performance of an urban transport interchange and manage the available resources properly. Likewise, key factors of the design and operation of urban transport interchanges are identified and some 'Planning guidelines' are proposed on the basis on them. Since the users' perceptions of their experience are particularly important for achieving the most appropriate policy measures for interchanges, an ad‐hoc travellers' satisfaction survey was designed and carried out in 2013 at three European transport interchanges: Moncloa (Madrid, Spain), Kamppi (Helsinki, Finland) and Ilford Railway Station (London, United Kingdom) In summary, this thesis highlights the ambivalent nature of the urban transport interchanges, i.e. as nodes within the transport network and as places where users spending time and proposes some policy recommendations in order to make urban transport interchanges attractive for users.

Functionality of major histocompatibility complex class II molecules in mice doubly deficient for invariant chain and H-2M complexes

By combining two previously generated null mutations, Ii° and M°, we produced mice lacking the invariant chain and H-2M complexes, both required for normal cell-surface expression of major histocompatibility complex class II molecules loaded with the usual diverse array of peptides. As expected, the maturation and transport of class II molecules, their expression at the cell surface, and their capacity to present antigens were quite similar for cells from Ii°M° double-mutant mice and from animals carrying just the Ii° mutation. More surprising were certain features of the CD4+ T cell repertoire selected in Ii°M° mice: many fewer cells were selected than in Ii+M° animals, and these had been purged of self-reactive specificities, unlike their counterparts in Ii+M° animals. These findings suggest (i) that the peptides carried by class II molecules on stromal cells lacking H-2M complexes may almost all derive from invariant chain and (ii) that H-2M complexes edit the peptide array displayed on thymic stromal cells in the absence of invariant chain, showing that it can edit, in vivo, peptides other than CLIP.

Cytoplasmic Dynein Heavy Chain 1b Is Required for Flagellar Assembly in Chlamydomonas

A second cytoplasmic dynein heavy chain (cDhc) has recently been identified in several organisms, and its expression pattern is consistent with a possible role in axoneme assembly. We have used a genetic approach to ask whether cDhc1b is involved in flagellar assembly in Chlamydomonas. Using a modified PCR protocol, we recovered two cDhc sequences distinct from the axonemal Dhc sequences identified previously. cDhc1a is closely related to the major cytoplasmic Dhc, whereas cDhc1b is closely related to the minor cDhc isoform identified in sea urchins, Caenorhabditis elegans, and Tetrahymena. The Chlamydomonas cDhc1b transcript is a low-abundance mRNA whose expression is enhanced by deflagellation. To determine its role in flagellar assembly, we screened a collection of stumpy flagellar (stf) mutants generated by insertional mutagenesis and identified two strains in which portions of the cDhc1b gene have been deleted. The two mutants assemble short flagellar stumps (<1–2 μm) filled with aberrant microtubules, raft-like particles, and other amorphous material. The results indicate that cDhc1b is involved in the transport of components required for flagellar assembly in Chlamydomonas.

A Specific Light Chain of Kinesin Associates with Mitochondria in Cultured Cells

The motor protein kinesin is implicated in the intracellular transport of organelles along microtubules. Kinesin light chains (KLCs) have been suggested to mediate the selective binding of kinesin to its cargo. To test this hypothesis, we isolated KLC cDNA clones from a CHO-K1 expression library. Using sequence analysis, they were found to encode five distinct isoforms of KLCs. The primary region of variability lies at the carboxyl termini, which were identical or highly homologous to carboxyl-terminal regions of rat KLC B and C, human KLCs, sea urchin KLC isoforms 1–3, and squid KLCs. To examine whether the KLC isoforms associate with different cytoplasmic organelles, we made an antibody specific for a 10-amino acid sequence unique to B and C isoforms. In an indirect immunofluorescence assay, this antibody specifically labeled mitochondria in cultured CV-1 cells and human skin fibroblasts. On Western blots of total cell homogenates, it recognized a single KLC isoform, which copurified with mitochondria. Taken together, these data indicate a specific association of a particular KLC (B type) with mitochondria, revealing that different KLC isoforms can target kinesin to different cargoes.

BiP and Immunoglobulin Light Chain Cooperate to Control the Folding of Heavy Chain and Ensure the Fidelity of Immunoglobulin Assembly

The immunoglobulin (Ig) molecule is composed of two identical heavy chains and two identical light chains (H2L2). Transport of this heteromeric complex is dependent on the correct assembly of the component parts, which is controlled, in part, by the association of incompletely assembled Ig heavy chains with the endoplasmic reticulum (ER) chaperone, BiP. Although other heavy chain-constant domains interact transiently with BiP, in the absence of light chain synthesis, BiP binds stably to the first constant domain (CH1) of the heavy chain, causing it to be retained in the ER. Using a simplified two-domain Ig heavy chain (VH-CH1), we have determined why BiP remains bound to free heavy chains and how light chains facilitate their transport. We found that in the absence of light chain expression, the CH1 domain neither folds nor forms its intradomain disulfide bond and therefore remains a substrate for BiP. In vivo, light chains are required to facilitate both the folding of the CH1 domain and the release of BiP. In contrast, the addition of ATP to isolated BiP–heavy chain complexes in vitro causes the release of BiP and allows the CH1 domain to fold in the absence of light chains. Therefore, light chains are not intrinsically essential for CH1 domain folding, but play a critical role in removing BiP from the CH1 domain, thereby allowing it to fold and Ig assembly to proceed. These data suggest that the assembly of multimeric protein complexes in the ER is not strictly dependent on the proper folding of individual subunits; rather, assembly can drive the complete folding of protein subunits.

Early Endosomes Are Required for Major Histocompatiblity Complex Class II Transport to Peptide-loading Compartments

Antigen presentation to CD4+ T lymphocytes requires transport of newly synthesized major histocompatibility complex (MHC) class II molecules to the endocytic pathway, where peptide loading occurs. This step is mediated by a signal located in the cytoplasmic tail of the MHC class II-associated Ii chain, which directs the MHC class II-Ii complexes from the trans-Golgi network (TGN) to endosomes. The subcellular machinery responsible for the specific targeting of MHC class II molecules to the endocytic pathway, as well as the first compartments these molecules enter after exit from the TGN, remain unclear. We have designed an original experimental approach to selectively analyze this step of MHC class II transport. Newly synthesized MHC class II molecules were caused to accumulate in the Golgi apparatus and TGN by incubating the cells at 19°C, and early endosomes were functionally inactivated by in vivo cross-linking of transferrin (Tf) receptor–containing endosomes using Tf-HRP complexes and the HRP-insoluble substrate diaminobenzidine. Inactivation of Tf-containing endosomes caused a marked delay in Ii chain degradation, peptide loading, and MHC class II transport to the cell surface. Thus, early endosomes appear to be required for delivery of MHC class II molecules to the endocytic pathway. Under cross-linking conditions, most αβIi complexes accumulated in tubules and vesicles devoid of γ-adaptin and/or mannose-6-phosphate receptor, suggesting an AP1-independent pathway for the delivery of newly synthesized MHC class II molecules from the TGN to endosomes.

Cytoplasmic Dynein, the Dynactin Complex, and Kinesin Are Interdependent and Essential for Fast Axonal Transport

In axons, organelles move away from (anterograde) and toward (retrograde) the cell body along microtubules. Previous studies have provided compelling evidence that conventional kinesin is a major motor for anterograde fast axonal transport. It is reasonable to expect that cytoplasmic dynein is a fast retrograde motor, but relatively few tests of dynein function have been reported with neurons of intact organisms. In extruded axoplasm, antibody disruption of kinesin or the dynactin complex (a dynein activator) inhibits both retrograde and anterograde transport. We have tested the functions of the cytoplasmic dynein heavy chain (cDhc64C) and the p150Glued (Glued) component of the dynactin complex with the use of genetic techniques in Drosophila. cDhc64C and Glued mutations disrupt fast organelle transport in both directions. The mutant phenotypes, larval posterior paralysis and axonal swellings filled with retrograde and anterograde cargoes, were similar to those caused by kinesin mutations. Why do specific disruptions of unidirectional motor systems cause bidirectional defects? Direct protein interactions of kinesin with dynein heavy chain and p150Glued were not detected. However, strong dominant genetic interactions between kinesin, dynein, and dynactin complex mutations in axonal transport were observed. The genetic interactions between kinesin and either Glued or cDhc64C mutations were stronger than those between Glued and cDhc64C mutations themselves. The shared bidirectional disruption phenotypes and the dominant genetic interactions demonstrate that cytoplasmic dynein, the dynactin complex, and conventional kinesin are interdependent in fast axonal transport.

The Involvement of the Intermediate Chain of Cytoplasmic Dynein in Binding the Motor Complex to Membranous Organelles of Xenopus Oocytes

Cytoplasmic dynein is one of the major motor proteins involved in intracellular transport. It is a protein complex consisting of four subunit classes: heavy chains, intermediate chains (ICs), light intermediate chains, and light chains. In a previous study, we had generated new monoclonal antibodies to the ICs and mapped the ICs to the base of the motor. Because the ICs have been implicated in targeting the motor to cargo, we tested whether these new antibodies to the intermediate chain could block the function of cytoplasmic dynein. When cytoplasmic extracts of Xenopus oocytes were incubated with either one of the monoclonal antibodies (m74–1, m74–2), neither organelle movement nor network formation was observed. Network formation and membrane transport was blocked at an antibody concentration as low as 15 μg/ml. In contrast to these observations, no effect was observed on organelle movement and tubular network formation in the presence of a control antibody at concentrations as high as 0.5 mg/ml. After incubating cytoplasmic extracts or isolated membranes with the monoclonal antibodies m74–1 and m74–2, the dynein IC polypeptide was no longer detectable in the membrane fraction by SDS-PAGE immunoblot, indicating a loss of cytoplasmic dynein from the membrane. We used a panel of dynein IC truncation mutants and mapped the epitopes of both antibodies to the N-terminal coiled-coil domain, in close proximity to the p150Glued binding domain. In an IC affinity column binding assay, both antibodies inhibited the IC–p150Glued interaction. Thus these findings demonstrate that direct IC–p150Glued interaction is required for the proper attachment of cytoplasmic dynein to membranes.

Relative role of heme nitrosylation and β-cysteine 93 nitrosation in the transport and metabolism of nitric oxide by hemoglobin in the human circulation

To quantify the reactions of nitric oxide (NO) with hemoglobin under physiological conditions and to test models of NO transport on hemoglobin, we have developed an assay to measure NO–hemoglobin reaction products in normal volunteers, under basal conditions and during NO inhalation. NO inhalation markedly raised total nitrosylated hemoglobin levels, with a significant arterial–venous gradient, supporting a role for hemoglobin in the transport and delivery of NO. The predominant species accounting for this arterial–venous gradient is nitrosyl(heme)hemoglobin. NO breathing increases S-nitrosation of hemoglobin β-chain cysteine 93, however only to a fraction of the level of nitrosyl(heme)hemoglobin and without a detectable arterial–venous gradient. A strong correlation between methemoglobin and plasma nitrate formation was observed, suggesting that NO metabolism is a primary physiological cause of hemoglobin oxidation. Our results demonstrate that NO–heme reaction pathways predominate in vivo, NO binding to heme groups is a rapidly reversible process, and S-nitrosohemoglobin formation is probably not a primary transport mechanism for NO but may facilitate NO release from heme.

Analysis of Respiratory Chain Regulation in Roots of Soybean Seedlings1

Changes in the respiratory rate and the contribution of the cytochrome (Cyt) c oxidase and alternative oxidase (COX and AOX, respectively) were investigated in soybean (Glycine max L. cv Stevens) root seedlings using the 18O-discrimination method. In 4-d-old roots respiration proceeded almost entirely via COX, but by d 17 more than 50% of the flux occurred via AOX. During this period the capacity of COX, the theoretical yield of ATP synthesis, and the root relative growth rate all decreased substantially. In extracts from whole roots of different ages, the ubiquinone pool was maintained at 50% to 60% reduction, whereas pyruvate content fluctuated without a consistent trend. In whole-root immunoblots, AOX protein was largely in the reduced, active form at 7 and 17 d but was partially oxidized at 4 d. In isolated mitochondria, Cyt pathway and succinate dehydrogenase capacities and COX I protein abundance decreased with root age, whereas both AOX capacity and protein abundance remained unchanged. The amount of mitochondrial protein on a dry-mass basis did not vary significantly with root age. It is concluded that decreases in whole-root respiration during growth of soybean seedlings can be largely explained by decreases in maximal rates of electron transport via COX. Flux via AOX is increased so that the ubiquinone pool is maintained in a moderately reduced state.

The Small, Methionine-Rich Chloroplast Heat-Shock Protein Protects Photosystem II Electron Transport during Heat Stress1

Evidence suggests that the small chloroplast heat-shock protein (Hsp) is involved in plant thermotolerance but its site of action is unknown. Functional disruption of this Hsp using anti-Hsp antibodies or addition of purified Hsp to chloroplasts indicated that (a) this Hsp protects thermolabile photosystem II and, consequently, whole-chain electron transport during heat stress; and (b) this Hsp completely accounted for heat acclimation of electron transport in pre-heat-stressed plants. Therefore, this Hsp is a major adaptation to acute heat stress in plants.

Yeast Gga Coat Proteins Function with Clathrin in Golgi to Endosome Transport

Gga proteins represent a newly recognized, evolutionarily conserved protein family with homology to the “ear” domain of the clathrin adaptor AP-1 γ subunit. Yeast cells contain two Gga proteins, Gga1p and Gga2p, that have been proposed to act in transport between the trans-Golgi network and endosomes. Here we provide genetic and physical evidence that yeast Gga proteins function in trans-Golgi network clathrin coats. Deletion of Gga2p (gga2Δ), the major Gga protein, accentuates growth and α-factor maturation defects in cells carrying a temperature-sensitive allele of the clathrin heavy chain gene. Cells carrying either gga2Δ or a deletion of the AP-1 β subunit gene (apl2Δ) alone are phenotypically normal, but cells carrying both gga2Δ and apl2Δ are defective in growth, α-factor maturation, and transport of carboxypeptidase S to the vacuole. Disruption of both GGA genes and APL2 results in cells so severely compromised in growth that they form only microcolonies. Gga proteins can bind clathrin in vitro and cofractionate with clathrin-coated vesicles. Our results indicate that yeast Gga proteins play an important role in cargo-selective clathrin-mediated protein traffic from the trans-Golgi network to endosomes.