1,720,987 research outputs found
HIV-1 Nef expression inhibits the activity of a Ca2+-dependent K+ channel involved in the control of the resting potential in CEM lymphocytes
No full textThe HIV-1 Nef protein plays an important role in the development of the pathology associated with AIDS. Despite various studies that have dealt with different aspects of Nef function, the complete mechanism by which it alters the physiology of infected cells remains to be established. Nef can associate with cell membranes, therefore supporting the hypothesis that it might interact with membrane proteins as ionic channels and modify their electrical properties. By using the patch-clamp technique, we found that Nef expression determines a 25-mV depolarization of lymphoblastoid CEM cells. Both charybdotoxin (CTX) and the membrane-permeable Ca2+ chelator BAPTA/AM depolarized the membrane of native cells without modifying that of Nef-transfected cells. These data suggested that the resting potential in native CEM cells is settled by a CTX- and Ca2+-sensitive K+ channel (KCa,CTX), whose activity is absent in Nef-expressing cells. This was confirmed by direct measurements of whole-cell KCa,CTX currents. Single-channel recordings on excised patches showed that a KCa,CTX channel of 35 pS with a half-activation near 400 nM Ca2+ was present in both native and Nef-transfected cells. The measurements of free intracellular Ca2+ were not different in the two cell lines, but Nef-transfected cells displayed an increased Ca2+ content in ionomycin-sensitive stores. Taken together, these results indicate that Nef expression alters the resting membrane potential of the T lymphocyte cell line by inhibiting a KCa,CTX channel, possibly by intervening in the regulation of intracellular Ca2+ homeostasis
Biochemical and computational approaches to dissect the effect of MT-CYB pathogenic mutations on respiratory chain activity and assembly
No full textComplex III (ubiquinol:cytochrome c oxidoreductase) is a multisubunit membrane bound enzyme and in its native form is a symmetrical homodimer (CIII2). CIII2 is central for mitochondrial respiratory chain and is associated with different stoichiometry with Complex I and Complex IV to form supramolecular assemblies, called supercomplexes (SCs). Defects in CIII2 are rare and mostly associated with mutations in MT-CYB gene that encodes for one of the catalytic subunits, cytochrome b (cyt b). It has been suggested that pathogenic mutations in MT-CYB are mitigated when CIII2 is assembled in SCs [1]. Therefore, we applied biochemical approaches in human cellular models carrying pathogenic point mutations in cyt b to analyse the structural stability and enzymatic activity of CIII2. Our preliminary results showed that pathogenic mutation differently affected the kinetics of the assembly of CIII2 and its SCs after the treatment with a reversible mitochondrial translation inhibitor, suggesting a role of these mutations not only in CIII2 activity but also in its biogenesis. Interestingly, the rescue of the oxygen consumption profile was further delayed compared to the formation of enzymatic complexes. In addition, we applied the Protein Stability Prediction with a Gaussian Network Model (PSP-GNM) approach [2] to evaluate global changes in the unfolding Gibbs free energy change and study the effects of single amino acid mutations on cyt b stability on the available isolated and SC-bound CIII2 structures. Preliminary results indicate that some pathogenic mutations may affect the unfolding free energy of CIII2, stiffening the structure of the enzyme, in agreement with the reduction of CIII2 activity. This dual experimental and biocomputational approach may be very useful to dissect assembly processes and function of the respiratory chain to better understand the effect of these rare pathogenic mutations and to design new strategies for possible therapeutic options
Cybrid Cell Lines As a Model to Study the Effects of Mitochondrial DNA Mutations on the Mitochondrial Proteome.
No full textA distruptive mitochondrial DNA mutation in respiratory complex I: Two-dimensional electrophoresis analysis of mitochondrial proteome of cybrid cell lines.
No full textAnalysis of mitochondrial proteome of cybrid cell harbouring a disruptive mitochondrial DNA mutation in respiratory complex I.
No full textGoing 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
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