1,720,968 research outputs found
Mechanistic study on the DNA methyltransferase DNMT3A
Get PDFThe phenotypical and functional diversity of mammalian cell types can be attributed to a large extent to epigenetic signals that determine and stabilize gene expression profiles. One of the most important types of epigenetic signals is DNA methylation. This modification is set early in development by the de novo DNA methyltransferases DNMT3A and DNMT3B, and is found predominantly at the C5 position of cytosine bases in a CpG dinucleotide context. The accurate setting of DNA methylation patterns is critical for normal development and is determined by the precise recruitment and control of DNMT activity on chromatin. In this work, four main directions of research were undertaken, with the ultimate goal of shedding novel mechanistic insights into the mechanism of DNMT3A, its regulation by chromatin signals and interaction partners, as well as the dysregulation of this enzyme in cancer. Furthermore, the potential of DNMT3A to generate 3-methylcytosine as a side reaction was explored.
The DNA methyltransferase DNMT3A has been shown to multimerize on DNA and to form large multimeric protein/DNA fibers. However, it has also been postulated that this enzyme can methylate DNA in a processive manner, a property incompatible with fiber formation. By using a dedicated set of biochemical experiments, I was able to show that the DNA methylation rate of DNMT3A increases more than linearly with increasing enzyme concentration on a long DNA substrate, but not on a short 30-mer oligonucleotide, which cannot accommodate DNMT3A polymers. Methylation experiments over a range of enzyme concentrations and with substrates containing one or two CpG sites did not provide evidence for a processive mechanism. The addition of a catalytically inactive DNMT3A mutant was found to increase the DNA methylation rate by DNMT3A on the long substrate but not on the short one. Together, these data clearly indicate that DNMT3A binds to DNA in a cooperative reaction and the formation of protein/DNA fibers increases the DNA methylation rate. These results contribute mechanistic insights into the mode by which DNA methylation patterns are established during development.
The second project dealt with characterizing the effects of the R882H exchange on DNMT3A. The R882H mutation is found in the DNA binding interface of DNMT3A and is frequently observed in acute myeloid leukemia (AML). By establishing a double-tag affinity purification system, I was able to show that the mutation only leads to a minor reduction in overall DNA methylation activity in mixed R882H/wildtype DNMT3A complexes. However, a pronounced change in flanking sequence preference of the DNMT3A-R882H mutant was found. Accordingly, a substrate designed to contain the target CpG site flanked by sequences preferred by R882H was better methylated by the variant than by the wildtype enzyme. Together, these data strongly argue against a dominant-negative effect of the R882H mutation and rather propose a site-specific gain-of-activity effect. These findings are in agreement with a recently determined structure of DNMT3A in complex with DNA and they might explain the high prevalence of this specific point mutation in AML.
The third project was built on previous data from the lab, documenting a strong and direct interaction between the ADD domain of DNMT3A and the TRD domain of the 5mC reading protein MECP2. These experiments revealed that through its binding, MECP2 allosterically stabilizes the autoinhibitory conformation of DNMT3A, resulting in a strong inhibition of enzymatic activity in vitro. The interaction between these two proteins and its associated inhibition could be disrupted by unmodified histone H3. In my work, I further validated the interaction between the ADD and the TRD domains by size exclusion chromatography. Also, by generating cell lines with stable over-expression of MECP2, I could show that MECP2 inhibits DNMT3A activity in cells. Together, the data from this study offer unprecedented insights into the regulation of DNMT3A by the combined action of chromatin modifications and interaction partners. Accordingly, depending on the modification status of the H3 tail at the target site, MECP2 can act as either a repressor or activator of DNA methylation.
The last project dealt with the coevolution between DNA methylation and DNA repair systems, a very exciting topic that was addressed in close collaboration with the laboratory of Dr. Peter Sarkies (MRC London). By performing in vitro methylation experiments with the catalytic domain of DNMT3A, I could show that, in addition to 5mC, DNMT3A can also introduce 3mC, a modification which represents an alkylation damage of DNA. This study provides a new evolutionary perspective on the loss of DNA methylation that is observed in many species
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Get PDF“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Get PDFWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Get PDFWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
Author-wise bibliometric analysis based on entropy.
No full textAuthor-wise bibliometric analysis based on entropy.</p
Author Under Sail The Imagination of Jack London, 1893-1902
No full textIn Author Under Sail, Jay Williams offers the first complete literary biography of Jack London as a professional writer engaged in the labor of writing. It examines the authorial imagination in London's work, the use of imagination in both his fiction and nonfiction, and the ways he defined imagination in the creative process in his business dealings with his publishers, editors, and agents. In this first volume of a two-volume biography, Williams traverses the years 1893 to 1902, from London's "Story of a Typhoon" to The People of the Abyss. The Jack London who emerges in the pages of Author Under Sail is a writer whose partnership with publishers, most notably his productive alliance with George Brett of Macmillan, was one of the most formative in American literary history. London pioneered many author models during the heyday of realism and naturalism, blurring the boundaries of these popular genres by focusing on absorption and theatricality and the representation of the seen and unseen. London created an impassioned, sincere, and extremely personal realism unlike that of other American writers of the time. Author Under Sail is a literary tour de force that reveals the full range of London as writer, creative citizen, and entrepreneur at the same time it sheds light on the maverick side of machine-age literature.Intro -- Title Page -- Copyright Page -- Dedication -- Contents -- Acknowledgments -- Introduction -- 1. Spirit Truth -- 2. From Absorption to Theatricality and Back Again -- 3. "I Will Build a New Present" -- 4. Sons as Authors -- 5. Fathers as Publishers -- 6. The Daughter as Author -- 7. Lovers as Authors -- 8. At Sea with the Family -- 9. Yellow News, Yellow Stories -- 10. The Return Home -- Notes -- Bibliography -- Index -- About Jay WilliamsIn Author Under Sail, Jay Williams offers the first complete literary biography of Jack London as a professional writer engaged in the labor of writing. It examines the authorial imagination in London's work, the use of imagination in both his fiction and nonfiction, and the ways he defined imagination in the creative process in his business dealings with his publishers, editors, and agents. In this first volume of a two-volume biography, Williams traverses the years 1893 to 1902, from London's "Story of a Typhoon" to The People of the Abyss. The Jack London who emerges in the pages of Author Under Sail is a writer whose partnership with publishers, most notably his productive alliance with George Brett of Macmillan, was one of the most formative in American literary history. London pioneered many author models during the heyday of realism and naturalism, blurring the boundaries of these popular genres by focusing on absorption and theatricality and the representation of the seen and unseen. London created an impassioned, sincere, and extremely personal realism unlike that of other American writers of the time. Author Under Sail is a literary tour de force that reveals the full range of London as writer, creative citizen, and entrepreneur at the same time it sheds light on the maverick side of machine-age literature.Description based on publisher supplied metadata and other sources.Electronic reproduction. Ann Arbor, Michigan : ProQuest Ebook Central, YYYY. Available via World Wide Web. Access may be limited to ProQuest Ebook Central affiliated libraries
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