Differential Epitope Tagging of Actin in Transformed Drosophila Produces Distinct Effects on Myofibril Assembly and Function of the Indirect Flight Muscle

We have tested the impact of tags on the structure and function of indirect flight muscle (IFM)-specific Act88F actin by transforming mutant Drosophila melanogaster, which do not express endogenous actin in their IFMs, with tagged Act88F constructs. Epitope tagging is often the method of choice to monitor the fate of a protein when a specific antibody is not available. Studies addressing the functional significance of the closely related actin isoforms rely almost exclusively on tagged exogenous actin, because only few antibodies exist that can discriminate between isoforms. Thereby it is widely presumed that the tag does not significantly interfere with protein function. However, in most studies the tagged actin is expressed in a background of endogenous actin and, as a rule, represents only a minor fraction of the total actin. The Act88F gene encodes the only Drosophila actin isoform exclusively expressed in the highly ordered IFM. Null mutations in this gene do not affect viability, but phenotypic effects in transformants can be directly attributed to the transgene. Transgenic flies that express Act88F with either a 6x histidine tag or an 11-residue peptide derived from vesicular stomatitis virus G protein at the C terminus were flightless. Overall, the ultrastructure of the IFM resembled that of the Act88F null mutant, and only low amounts of C-terminally tagged actins were found. In contrast, expression of N-terminally tagged Act88F at amounts comparable with that of wild-type flies yielded fairly normal-looking myofibrils and partially reconstituted flight ability in the transformants. Our findings suggest that the N terminus of actin is less sensitive to modifications than the C terminus, because it can be tagged and still polymerize into functional thin filaments.

Linking genome and proteome by mass spectrometry: Large-scale identification of yeast proteins from two dimensional gels

The function of many of the uncharacterized open reading frames discovered by genomic sequencing can be determined at the level of expressed gene products, the proteome. However, identifying the cognate gene from minute amounts of protein has been one of the major problems in molecular biology. Using yeast as an example, we demonstrate here that mass spectrometric protein identification is a general solution to this problem given a completely sequenced genome. As a first screen, our strategy uses automated laser desorption ionization mass spectrometry of the peptide mixtures produced by in-gel tryptic digestion of a protein. Up to 90% of proteins are identified by searching sequence data bases by lists of peptide masses obtained with high accuracy. The remaining proteins are identified by partially sequencing several peptides of the unseparated mixture by nanoelectrospray tandem mass spectrometry followed by data base searching with multiple peptide sequence tags. In blind trials, the method led to unambiguous identification in all cases. In the largest individual protein identification project to date, a total of 150 gel spots—many of them at subpicomole amounts—were successfully analyzed, greatly enlarging a yeast two-dimensional gel data base. More than 32 proteins were novel and matched to previously uncharacterized open reading frames in the yeast genome. This study establishes that mass spectrometry provides the required throughput, the certainty of identification, and the general applicability to serve as the method of choice to connect genome and proteome.

Assembly of large genomic segments in artificial chromosomes by homologous recombination in Escherichia coli

We developed a method for the reconstruction of a 100 kb DNA fragment into a bacterial artificial chromosome (BAC). The procedure makes use of iterative rounds of homologous recombination in Escherichia coli. Smaller, overlapping fragments of cloned DNA, such as cosmid clones, are required. They are transferred first into a temperature-sensitive replicon and then into the BAC of choice. We demonstrated the usefulness of this procedure by assembling a 90 kb genomic segment into an E.coli–Streptomyces artificial chromosome (ESAC). Using this procedure, ESACs are easy to handle and remarkably more stable than the starting cosmids.

Ethics and access to teaching materials in the medical library: the case of the Pernkopf atlas*

Conflicts can occur between the principle of freedom of information treasured by librarians and ethical standards of scientific research involving the propriety of using data derived from immoral or dishonorable experimentation. A prime example of this conflict was brought to the attention of the medical and library communities in 1995 when articles claiming that the subjects of the illustrations in the classic anatomy atlas, Eduard Pernkopf's Topographische Anatomie des Menschen, were victims of the Nazi holocaust. While few have disputed the accuracy, artistic, or educational value of the Pernkopf atlas, some have argued that the use of such subjects violates standards of medical ethics involving inhuman and degrading treatment of subjects or disrespect of a human corpse. Efforts were made to remove the book from medical libraries. In this article, the history of the Pernkopf atlas and the controversy surrounding it are reviewed. The results of a survey of academic medical libraries concerning their treatment of the Pernkopf atlas are reported, and the ethical implications of these issues as they affect the responsibilities of librarians is discussed.

The Pursuit of Balanced Income An Economic Perspective on Wealth

The purpose of this thesis is to determine if households with moderate incomes have the opportunity to acquire pecuniary wealth. In an environment of increasing globalization, labor-eliminating technology, and an overburdened entitlement system, the position of wealth is necessary for households with a penchant for early retirement and particularly for those that will inevitably succumb to involuntary retirement. In these writings, qualitative and quantitative descriptions of wealth are offered; two microeconomic metrics are introduced to identify baseline wealth and to gauge proximity to this value; and strategies are devised to close the gap between these metrics. The construct of the economic intertemporal household budget was used as the basis for this research whereby conventional measures of wealth, i.e. net worth and liquid net worth, have been coupled with historical and empirical household data to define and extrapolate wealth and wealth proximity metrics. The major finding is wealth is a dynamic variable contingent upon consumption level rather than income or saving levels. And because households exercise a greater degree of autonomy over consumption, moderate earners are afforded the same opportunities as high earners. The position of wealth is necessary because it sustains consumption in the event of an unwelcomed retirement and it can assuage reliance on entitlement programs. For the majority of households, wealth is a matter of choice.

Internet Civil Liberties in the U.S. and the Protection of the Modern Terrorist

This capstone examines the civil liberties of the modern terrorist and explicates the right to freedom of speech for terrorist organizations and their use of the internet. Terrorist organizations use the internet to promote ideas, recruit members, organize the flow of information, and coordinate actions. During the war on terror the US Patriot Act became law allowing for U.S. government censorship and surveillance of internet traffic and many believe these acts are a threat to civil liberties. Terrorist organizations have the right to express their views, however unpopular, and censoring or restricting web sites diminishes civil liberties for all, which democracies and liberal societies are founded upon.

An Independent Art Movement: The Independent Artist, Community Television, and the Age of Information

The history of community television shows that it has been a home to activist and non-profit organizations that have created programs focused on freedom of speech. This project proposes that community television is also a place where artists can have freedom of artistic expression. The reflective paper reviews the creation of my film designed to inform and attract artists to community television. In it I critically reflect on the artistic, technical, artistic/technical, and production changes made throughout my journey from being a visual artist to becoming a video-artist. The reflective paper, along with the film, act as a wake-up call to artists who are unaware of community television and the advantages it has to offer them.

The negative effects of failed service recoveries

Research has shown that more than half of attempted recovery efforts fail, producing a ‘double deviation’ effect. Surprisingly, these double deviation effects have received little attention in marketing literature. This paper examines what happens after these critical encounters, which behavior or set of behaviors the customers are prone to follow and how customers’ perceptions of the firm’s recovery efforts influence these behaviors. For the analysis of choice of the type of response (complaining, exit, complaining and exit, and no-switching), we estimate multinomial Logit models with random coefficients (RCL). The results of our study show that magnitude of service failure, explanations, apologies, perceived justice, angry and frustration felt by the customer, and satisfaction with service recovery have a significant effect on customers’ choice of the type of response. Implications from the findings are offered.

Management of social networks in the educational process

The research developed in this work consists in proposing a set of techniques for management of social networks and their integration into the educational process. The proposals made are based on assumptions that have been proven with simple examples in a real scenario of university teaching. The results show that social networks have more capacity to spread information than educational web platforms. Moreover, educational social networks are developed in a context of freedom of expression intrinsically linked to Internet freedom. In that context, users can write opinions or comments which are not liked by the staff of schools. However, this feature can be exploited to enrich the educational process and improve the quality of their achievement. The network has covered needs and created new ones. So, the figure of the Community Manager is proposed as agent in educational context for monitoring network and aims to channel the opinions and to provide a rapid response to an academic problem.

spotlight europe #2013/05 - December 2013: Unblocking the lifeline of talent

Against the background of demographic decline and growing economic competitiveness from emerging economies, this Spotlight published in cooperation with the Centre for European Policy Studies looks into the potential of increased intra-EU labour mobility. It will examine the ‘German case’ on EU labour mobility. It proposes ideas on how to better foster a European fair deal on talent, one that would benefit the EU as a whole. It concludes with a proposal on how to increase the benefits of the freedom of movement.

Unblocking the Lifeline of Talent. CEPS Policy Brief No. 306, 6 December 2013

Against the background of looming demographic decline, the departure of the baby-boom generation from European labour markets and growing economic competitiveness from emerging economies, this CEPS Policy Brief, published jointly with the Bertelsmann Stiftung, looks into the potential benefits of increased intra-EU labour mobility. The authors examine the ‘German case’ on EU labour mobility, digging below the surface of the aggregate data. They offer proposals on how to foster a European fair deal on talent, one that would benefit the EU as a whole. The paper concludes with policy recommendations on how to increase the potential benefits of the freedom of movement for both individual EU citizens and for the EU as a whole.

Stigmatisation of EU mobile citizens: a ticking time bomb for the European project. EPC Commentary, 24 January 2014

The 2013 European Year of Citizens was profoundly marked by escalating attacks against one of the EU’s major achievement for EU citizens: freedom of movement. In April 2013, Home Affairs Ministers from Austria, Germany, the Netherlands and the UK were party to a letter claiming that “a significant number of new immigrants draw social assistance in the host countries, frequently without genuine entitlement, burdening host societies’ social welfare systems”. This letter laid the groundwork for a “battle plan”, presented by David Cameron in November, which aimed to make the free movement of persons “less free” and put forward the idea of capping “EU migration”. Furthermore, in December, the German conservative Christian Social Union (CSU) took up a similar petty political discourse. After the end of the transitional period for Romania and Bulgaria on 1 January 2014, the debate continues with Chuka Umunna (British Labour Party) proposing to restrict the freedom of movement to highly skilled EU citizens and to citizens in possession of a firm job offer. Alongside this, the German Chancellor, Angela Merkel announced the formation of a committee to investigate “poverty migration” in Germany. This wave of resentment has been more recently followed by the UK Prime Minister David Cameron, expressing his intention to re-negotiate EU law in order to be able to withdraw child benefits from EU citizens working in the UK, citing Polish citizens working in the UK as an example. Seeing this as a stigmatisation of the Polish population, the Polish foreign minister, Radosław Sikorski, qualified Cameron’s discourse as “unacceptable”. The debate over limiting freedom of movement has continuously escalated and reached a worrying level. With the EP elections approaching in May 2014, this debate is likely to become worse.

The future of human rights protection in the new Libya: prospects and challenges. Jean Monnet Occasional Paper No. 6/2014

There is no doubt that demand for the respect of human rights was one of the factors behind the Arab Spring and Libya is no exception. Four decades of absolute dictatorship headed by Muammar Gaddafi had been further tainted with gross violations of human rights of Libyan citizens and restrictions on their basic freedoms. Before the revolution, Libya was a country where no political parties were allowed. Freedom of expression and the press were extremely restricted. Reports about the country’s human rights violations published by a number of international organizations documented large scale human rights abuses at the hands of the Gaddafi regime. The 17 February 2011 revolution in Libya led to a turning point in the country’s history. The regime of Muammar Gaddafi which had dominated the country since 1969 eventually collapsed, leading to the beginning of the painful task of reconciliation and state building. Nonetheless it is estimated that more than 7000 prisoners are held captive by various militias and armed groups without due process. This in addition to thousands of internally displaced persons. State building involves the consolidation of a democratic state based on a democratic constitution. In 2011, a constitutional declaration was adopted to replace the one that had been in effect since 1969. This was intended as a stop-gap solution to allow the new political forces unleashed in the country time to write a new democratic constitution. To help consolidate the democratic state, three elements are required: that human rights be placed at its core; that these rights are truly implemented and applied; and lastly that the independence of the judiciary is safeguarded. For all this to happen it is also essential to strengthen education on human rights by encouraging non-governmental organizations to take a stronger role in promoting human rights. Libyan citizens can only avail themselves of these rights and strengthen their implementation if they know what they are and how they can benefit from their implementation

Internet “Freedom” in Turkey. EPC Commentary, 2 March 2015

From the Introduction. Fundamental rights and freedoms in Turkey have been in decline for the last few years, including internet freedom, with Turkey hardly meeting the EU’s Human Rights Guidelines on Freedom of Expression.

Recent Changes in European Welfare State Services: A Comparison of Child Care Politics in the U.K., Sweden, Germany, and the Netherlands. CES Germany & Europe Working Papers, No. 07.6, 1997

This paper examines recent policies and politics of services, in particular child care services in European welfare states. It is argued that social (care) services are becoming an increasingly political issue in postindustrial societies and are at the very center of welfare-state restructuring. Some countries have recently developed new policy pro­ grams for child care-but there are important differences among these programs. To understand these differences as well as some common features, the paper argues that it is necessary to examine the institutional organization of child care and short-term political factors as well as the rationales articulated in political debates to support or im­ pede various policies. The paper concludes that a comprehensive system of child care provisions is still far off in most countries, despite a rhetoric of choice and postindustrial care and labor-market patterns.