Prediction and confirmation of a switch-like region within the N-terminal domain of hSIRT1

Many proteins display conformational changes resulting from allosteric regulation. Often only a few residues are crucial in conveying these structural and functional allosteric changes. These regions that undergo a significant change in structure upon receiving an input signal, such as molecular recognition, are defined as switch- like regions. Identifying these key residues within switch-like regions can help elucidate the mechanism of allosteric regulation and provide guidance for synthetic regulation. In this study, we combine a novel computational workflow with biochemical methods to identify a switch-like region in the N-terminal domain of human SIRT1 (hSIRT1), a lysine deacetylase that plays important roles in regulating cellular pathways. Based on primary sequence, computational methods predicted a region between residues 186–193 in hSIRT1 to exhibit switch-like behavior. Mutations were then introduced in this region and the resulting mutants were tested for allosteric reactions to resveratrol, a known hSIRT1 allosteric regulator. After fine-tuning the mutations based on comparison of known secondary structures, we were able to pinpoint M193 as the residue essential for allosteric regulation, likely by communicating the allosteric signal. Mutation of this residue maintained enzyme activity but abolished allosteric regulation by resveratrol. Our findings suggest a method to predict switch-like regions in allosterically regulated enzymes based on the primary sequence. If further validated, this could be an efficient way to identify key residues in enzymes for therapeutic drug targeting and other applications

Light Curves and Period Changes of Type II Cepheids in the Globular Clusters M3 and M5

Light curves in the B, V, and I_c passbands have been obtained for the type II Cepheids V154 in M3 and V42 and V84 in M5. Alternating cycle behavior, similar to that seen among RV Tauri variables, is confirmed for V84. Old and new observations, spanning more than a century, show that V154 has increased in period while V42 has decreased in period. V84, on the other hand, has shown large, erratic changes in period that do not appear to reflect the long term evolution of V84 through the HR diagram.Comment: 28 pages, 12 figure

The Stress Response Factors Yap6, Cin5, Phd1, and Skn7 Direct Targeting of the Conserved Co-Repressor Tup1-Ssn6 in S. cerevisiae

Maintaining the proper expression of the transcriptome during development or in response to a changing environment requires a delicate balance between transcriptional regulators with activating and repressing functions. The budding yeast transcriptional co-repressor Tup1-Ssn6 is a model for studying similar repressor complexes in multicellular eukaryotes. Tup1-Ssn6 does not bind DNA directly, but is directed to individual promoters by one or more DNA-binding proteins, referred to as Tup1 recruiters. This functional architecture allows the Tup1-Ssn6 to modulate the expression of genes required for the response to a variety of cellular stresses. To understand the targeting or the Tup1-Ssn6 complex, we determined the genomic distribution of Tup1 and Ssn6 by ChIP-chip. We found that most loci bound by Tup1-Ssn6 could not be explained by co-occupancy with a known recruiting cofactor and that deletion of individual known Tup1 recruiters did not significantly alter the Tup1 binding profile. These observations suggest that new Tup1 recruiting proteins remain to be discovered and that Tup1 recruitment typically depends on multiple recruiting cofactors. To identify new recruiting proteins, we computationally screened for factors with binding patterns similar to the observed Tup1-Ssn6 genomic distribution. Four top candidates, Cin5, Skn7, Phd1, and Yap6, all known to be associated with stress response gene regulation, were experimentally confirmed to physically interact with Tup1 and/or Ssn6. Incorporating these new recruitment cofactors with previously characterized cofactors now explains the majority of Tup1 targeting across the genome, and expands our understanding of the mechanism by which Tup1-Ssn6 is directed to its targets

The Membrane Fusion Step of Vaccinia Virus Entry Is Cooperatively Mediated by Multiple Viral Proteins and Host Cell Components

For many viruses, one or two proteins allow cell attachment and entry, which occurs through the plasma membrane or following endocytosis at low pH. In contrast, vaccinia virus (VACV) enters cells by both neutral and low pH routes; four proteins mediate cell attachment and twelve that are associated in a membrane complex and conserved in all poxviruses are dedicated to entry. The aim of the present study was to determine the roles of cellular and viral proteins in initial stages of entry, specifically fusion of the membranes of the mature virion and cell. For analysis of the role of cellular components, we used well characterized inhibitors and measured binding of a recombinant VACV virion containing Gaussia luciferase fused to a core protein; viral and cellular membrane lipid mixing with a self-quenching fluorescent probe in the virion membrane; and core entry with a recombinant VACV expressing firefly luciferase and electron microscopy. We determined that inhibitors of tyrosine protein kinases, dynamin GTPase and actin dynamics had little effect on binding of virions to cells but impaired membrane fusion, whereas partial cholesterol depletion and inhibitors of endosomal acidification and membrane blebbing had a severe effect at the later stage of core entry. To determine the role of viral proteins, virions lacking individual membrane components were purified from cells infected with members of a panel of ten conditional-lethal inducible mutants. Each of the entry protein-deficient virions had severely reduced infectivity and except for A28, L1 and L5 greatly impaired membrane fusion. In addition, a potent neutralizing L1 monoclonal antibody blocked entry at a post-membrane lipid-mixing step. Taken together, these results suggested a 2-step entry model and implicated an unprecedented number of viral proteins and cellular components involved in signaling and actin rearrangement for initiation of virus-cell membrane fusion during poxvirus entry

Explaining cross-cultural service interactions in tourism with Shenkar’s Cultural Friction

In this paper, we commence a new dialogue on cross-cultural research in tourism. Using Shenkar’s (2001) metaphor of cultural friction as the analytical framework, we examine crosscultural service interactions between guests and service-providers in a luxury hotel. Cultural friction departs from, and extends, the notion of ‘cultural distance’, as it recognises asymmetry in social-economic conditions and considers the goals and the influence of control and power between the interacting parties. We use the Critical Incident Technique and Narrative Inquiry as the data collection technique and analytical approach respectively. The findings reveal that guests and service-providers use a number of strategies to exert power and gain control during their interactions, including subjective essentialism and stereotyping, to achieve their goals. The implications for tourism and hospitality management include providing cross-cultural sensitivity training to service-providers, ensuring a cultural-diverse employee composition, and to foster cross-cultural understanding amongst employees. We further suggest to develop strategies to facilitate effective cross-cultural service interactions based on evidence about cultural norms, expectations and behaviours from specific cultural groups. Further research is recommended to connect specific interactions between the interacting parties to examine whether the various strategies used leads to effective cross-cultural communication

"A Time to Gather": A History of Jewish Archives in the Twentieth Century

At the opening of the twentieth century, Jewish scholars turned to archives as a primary source of Jewish history and culture, and created diverse archives of their own. It was to be, as one scholar put it, a "time to gather"—a time when Jews the world over worked to bring together the records of the Jewish past, but when the shared impulse to preserve the past led to intense conflict. This dissertation explores the landscape of twentieth-century Jewish archives, tracing a transnational network of archives and archivists in Germany, the United States, and Israel/Palestine. Rather than casting these archives as neutral oases of objectivity, this study examines them as highly political sites of struggle over control of Jewish culture and memory. It investigates Jews’ rising interest in archives and the proliferation of archival projects that followed, and excavates a tradition of comprehensive collecting and the resulting conflicts over who could "own" the past. A Time to Gather argues that both before the Holocaust and especially in its aftermath, the act of creating Jewish archives was just as much about the future as it was about the past. In the twentieth century, Jews in various parts of the world harbored dreams of "total archives" that would comprehensively document Jewish life. These aspirations fueled fierce competition, as centralizing historical materials was one way to project cultural hegemony and to shape the way that history would be written. Against this backdrop, the study examines major archives including, among others, the Gesamtarchiv der deutschen Juden, founded in Berlin in 1905, the Jewish Historical General Archives in Jerusalem (since 1969 the Central Archives for the History of the Jewish People), and the American Jewish Archives in Cincinnati, both of which opened in 1947. This work seeks to comprehend the scope of this "time to gather," when Jewish scholars and leaders on three continents looked to archives as an important source of history and an anchor for communal memory, and to examine the significance of archiving for the development of the discipline of Jewish history as well as the politics of Jewish culture

The PREVENT Study: Psychiatric Reporting to Ease Vehicular Events Near Traffic

This thesis analyses health services data for a group of 23,145 patients who were warned by their physicians regarding motor vehicle driving risks between April 1, 2006 and March 31, 2011. A three-year baseline interval prior to the warning was compared to the one year subsequent to the warning for the rate of involvement in motor vehicle crashes and for psychiatric hospitalization. The relative risk of involvement in a serious crash in the subsequent compared to the baseline interval was 0.69 (95% confidence interval 0.59 to 0.81, PM.Sc

IHS Podcast: A Time to Gather: Archives and the Control of Jewish Culture

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