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Characterization of mouse polycystic kidney disease and receptor for egg jelly gene and protein in heterologous and native system
No full textThe mammalian polycystic kidney disease (PKD) gene family comprises eight members whose role in cell physiology is still poorly understood. Two of the founding members of the family, PKD1 and PKD2, are responsible for the majority of cases of autosomal dominant polycystic kidney disease. While PKD1 is considered to be a cell-surface receptor, PKD2 functions as a cation channel and has been described both at the plasma membrane and intracellularly. The present study focuses on PKDREJ – polycystic kidney disease and receptor for egg jelly, a protein that contains both a large receptor-like extracellular part and a putative ion channel region. PKDREJ homology to suREJ proteins, which seem to play a role in the induction of the sperm acrosome reaction in sea urchin, gives a reason to propose the same scenario for PKDREJ in mammals. Although the PKDREJ gene was identified in 1999, its features and protein characteristics remained elusive up to now.
The present study investigates mouse PKDREJ gene and protein and provides the first data about its structure, expression and localization. It demonstrates that PKDREJ is highly expressed in testicular tissue and its expression there is restricted to germ cells and spermatozoa.
Upon heterologous expression in HEK 293, COS7, GC-1, GC-2 cells and Xenopus laevis oocytes PKDREJ is retained intracellularly in endoplasmatic compartments. Since many known receptors and ion channels can not be properly trafficked without special partner or adaptor proteins, while expressed in a heterologous system, co-expression experiments with mouse HspA2, PKD2L2 and human PKD1 and PKD2 were done. In all cases PKDREJ localization remained intracellular irrespective of whether it was co-expressed with other proteins or not.
The presence of the GPS domain in the PKDREJ amino acid structure gave a reason to speculate about the proteolytical cleavage within this site. The present work provides not only the biochemical characterisation of the PKDREJ protein, but also the first evidence about lack of the GPS processing. It shows that PKDREJ unlike PKD1 or suREJ3 does not undergo cleavage within G-protein-coupled receptor proteolytic site while being expressed in HEK293 cells or Xenopus laevis oocytes. Moreover, the explanation of this phenomenon, based on the structure of the GPS domain in PKDREJ and the autocatalytic mechanism of the cleavage, is proposed.
To investigate the PKDREJ protein in native system PKDREJ-specific polyclonal antibodies were developed and characterised. By means of indirect immunofluorescence it was possible to demonstrate PKDREJ localization in the acrosomal region and on the inner aspect of the falciform-shaped mouse sperm head. Additional immunoelectron microscopy with mouse sperm revealed the plasma membrane localization of the PKDREJ protein and confirmed the predicted protein topology with the N-terminus located extracellularly.
To examine the functional relevance of PKDREJ in terms of the induction of the sperm acrosome reaction capacitated mouse sperm was incubated with antibodies directed against the PKDREJ N-terminus. We have not observed antibody-induced Ca2+ influx or a significant increase in the fraction of acrosome-reacted sperm. Also, anti-PKDREJ antibodies failed to block ZP-induced acrosome reaction. These observations do not completely exclude a role of PKDREJ for the induction of the acrosome reaction, since the antibodies may neither be able to stabilize an active protein conformation required for transmembrane signaling nor to block relevant epitopes needed for ZP signaling. However, it may be also the first hint for another role of PKDREJ in sperm physiology.
Although all other PKD family members are widely expressed, PKDREJ expression was considered to be testis-specific. Investigating non-testicular tissue distribution of PKDREJ we were able to detect the protein in cardiomyocytes and smooth muscles of pulmonary blood vessels; the staining pattern of PKDREJ subcellular localization in both cell types corresponded to transverse tubules (t-tubules). Another site of non-testicular expression was discovered in epithelial cells of kidney and epididymis. In kidney epithelium PKDREJ was co-localized with acetylated -tubulin, a marker of primary cilia. These results open a new direction for the further investigation of PKDREJ that will provide deeper insight into its function and physiological relevance.Die polycystic kidney disease (PKD)-Genfamilie besteht aus acht Genen, deren Rolle in der Zellphysiologie weiterhin zum größten Teil unklar ist. Die zwei zuerst entdeckten Mitglieder dieser Familie, PKD1 und PKD2, sind für die meisten Fälle der autosomal dominanten polyzystischen Nierenerkrankung verantwortlich. Während PKD1 ein zellmembranständiger Rezeptor zu sein scheint, handelt es sich bei PKD2 um einen Kationenkanal, welcher sowohl in der Zellmembran als auch intrazellulär beschrieben wurde. Die vorliegende Studie fokussiert auf polycystic kidney disease and receptor for egg jelly (PKDREJ), ein Protein welches sowohl einen großen rezeptorähnlichen extrazellulären Teil als auch einen mutmaßlichen Ionenkanal beinhaltet. Die Homologie von PKDREJ zu sea urchin receptor for egg jelly (suREJ)- Proteinen, welche wahrscheinlich eine Rolle in der Induktion der akrosomalen Reaktion von Spermatozoen bei Seeigeln spielen, gibt Anlass zu der Spekulation, dass PKDREJ die gleiche Funktion bei Säugetieren erfüllt. Obwohl das PKDREJ-Gen schon 1999 entdeckt wurde, sind seine Eigenschaften und Proteinmerkmale bislang weitestgehend unklar.
Diese Studie untersucht das murine PKDREJ-Gen und -Protein und beschreibt erstmalig dessen Struktur, Expression und Lokalisation. Es wird gezeigt, dass PKDREJ stark in testikulärem Gewebe exprimiert wird und diese Expression auf Keimzellen und Spermatozoen beschränkt ist.
Nach heterologer Expression in HEK 293-, COS7-, GC-1- und GC-2-Zellen sowie in Xenopus laevis-Eizellen verbleibt PKDREJ intrazellulär in endoplasmatischen Kompartimenten. Da viele bekannte Rezeptoren und Ionenkanäle, ohne spezielle Partner- oder „Adaptor“-Proteine nicht bestimmungsgemäß transportiert werden, wenn man sie in heterologen Systemen exprimiert, wurden Koexpressions-Experimente mit murinem HspA2, einem testisspezifischen Chaperon, PKD2L2, einem PKD2 eng verwandten Ionenkanal, und menschlichem PKD1 und PKD2 durchgeführt. In allen Fällen verblieb PKDREJ intrazellulär, unabhängig davon, ob mit oder ohne Koexpression anderer Proteine.
Das Vorhandensein einer GPS-Domäne in der PKDREJ-Aminosäuresequenz gab Anlass, die proteolytische Spaltung in dieser Region zu untersuchen. Dabei charakterisiert diese Arbeit nicht nur biochemische Eigenschaften des PKDREJ-Proteins, sondern gibt auch erste Hinweise auf eine fehlende proteolytische Abspaltung der N-terminalen Proteindomäne. In HEK293-Zellen oder Xenopus laevis-Oozyten exprimiertes PKDREJ wird im Gegensatz zu PKD1 oder suREJ3 nicht in seiner GPS-Domäne gespalten. Außerdem wird eine Erklärung dieses Phänomens, basierend auf der Struktur der GPS-Domäne von PKDREJ und dem autokatalytischem Mechanismus der Spaltung, vorgeschlagen.
Um das PKDREJ-Protein in einem nativen System zu untersuchen, wurden PKDREJ-spezifische polyklonale Antikörper entwickelt und charakterisiert. Durch indirekte Immunofluoreszenz war es möglich, PKDREJ in der akrosomalen Region und am inneren Anteil des sichelförmigen murinen Spermienkopfes zu lokalisieren. Zusätzliche Immun-Elektronenmikroskopie mit murinen Spermien zeigte die Lokalisation des PKDREJ-Proteins in der Plasmamembran und sicherte die vorausgesagte Protein-Topologie mit einem extrazellulär liegenden N-Terminus.
Um die funktionelle Relevanz von PKDREJ in Hinblick auf die Induktion der akrosomalen Reaktion zu zeigen wurden kapazitierte murine Spermien mit Antikörpern gegen das N-terminale Ende von PKDREJ-Protein inkubiert. Wir konnten keinen antikörperinduzierten Ca2+-Einstrom oder eine signifikante Steigerung des Anteils akrosomreagierter Spermien beobachten. Des weiteren blockierten anti-PKDREJ Antikörper die Zona pellucida (ZP)-induzierte akrosomale Reaktion nicht. Diese Beobachtungen schließen eine Rolle von PKDREJ in der Akrosomreaktion nicht vollständig aus, da die Antikörper eventuell nicht in der Lage sind, eine aktive Proteinkonformation, eine Voraussetzung für die transmembranäre Signalkette, zu stabilisieren oder die relevanten Epitope, die für ZP-Signaltransduktion benötigt werden, zu blockieren. Dennoch könnte diese Arbeit der erste Hinweis einer anderen, noch unbekannten Funktion von PKDREJ in der Physiologie von Spermien sein.
Obwohl die Mitglieder der PKD-Familie vielerorts exprimiert werden, galt PKDREJ bislang als hodenspezifisch. Wir konnten auf Proteinebene PKDREJ allerdings in Herzmuskelzellen und Lungenblutgefäßen nachweisen, wobei das Färbemuster von PKDREJ in beiden Zelltypen auf eine Lokalisation an den transversen Tubuli schließen lässt. Weitere nicht-testikuläre Expression wurde in Nierenepithelzellen und im Epididymis entdeckt. Im Nierenepithel war PKDREJ mit acetyliertem α-Tubilin kolokalisiert, einem Marker von primären Zilien. Diese Resultate bilden die Grundlage für weiterführende Untersuchungen zur Funktion und physiologischen Relevanz von PKDREJ
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe 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
Full text link“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
Full text linkWe 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
Full text linkWe 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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