1,720,962 research outputs found
Anti-DNA antibodies bind to DNase I.
No full textPolyspecificity is a well-known property of the anti-DNA antibodies produced by autoimmune animals. In our search for antigen targets of anti-DNA antibodies within tissue extracts, we identified a 32-kD polypeptide that was recognized by a large panel of anti-DNA antibodies. Direct sequencing of this protein disclosed its identity with DNase I. 22 monoclonal anti-DNA antibodies bound to DNase I in direct and competitive immunoassays; out of 15 autoantibodies that did not bind DNA, none had the ability to bind DNase I. The ability of anti-DNA antibodies to interfere with DNase I enzymatic activity was evaluated in an assay based on the enzyme digestion of phage double strand DNA. Six monoclonal anti-single strand DNA antibodies that did not bind double strand DNA were tested in this assay. Three out of six inhibited DNase I-mediated digestion of phage DNA. The interaction of anti-DNA antibodies with DNase I was further investigated by testing their ability to bind a synthetic peptide that corresponds to the catalytic site of the molecule. 4 out of 22 anti-DNA antibodies bound the active site peptide; two of these had been shown to inhibit DNase I enzymatic activity. This report show that anti-DNA antibodies recognize both DNA and its natural ligand DNase I. Some anti-DNA antibodies inhibit DNase I enzymatic activity, thus displaying the potential to modulate DNA catabolism. The dual specificity of anti-DNA antibodies offers a clue for understanding the mechanisms that lead to anti-DNA antibody production in autoimmune animals
ABCD Meeting “Organelle Biogenesis and Signal Transduction”
No full textAn ubiquitin-proteasome-endocytic pathway but not autophagy orchestrates ErbB2 internalization and cleavage in HSP90-inhibited breast cancer cell
ABCD 2015, Biennial Congress of the Italian Association of cell biology and differentiation
No full textHSP90 inhibition induces ErbB2 cleavage and internalization simultaneoulsy with alterations in endosomal trafficking/maturation and autophag
3D Breast Cancer Spheroids Reveal Architecture-Dependent HER2 Expression and Signaling
No full textBackground: Three-dimensional (3D) culture systems offer a physiologically relevant alternative to monolayers for studying tumor organization, signaling, and drug response. HER2-positive breast cancers (BCa) account for 15–30% of BCa cases and benefit from HER2-targeted therapies, yet predictive in vitro models remain limited. Objective: To generate and compare 3D spheroids from two HER2+ BCa cell lines, SKBR3 and BT474, and investigate how 3D architecture influences HER2 distribution, intracellular signaling, and cellular organization. Methods: Spheroids were reproducibly generated from SKBR3 and BT474 cells and analyzed after 4 days of culture. Cell viability was evaluated using live/dead staining, HER2 distribution was assessed by confocal microscopy and quantified on cryosections, and protein expression/phosphorylation was measured by Western blotting. Epithelial and EMT markers were visualized by immunofluorescence, and ultrastructural features were examined by transmission electron microscopy (TEM). Results: Both cell lines formed viable spheroids with distinct architectures: SKBR3 spheroids were loose and heterogeneous, whereas BT474 spheroids were compact and highly spherical. Confocal and cryosection imaging showed consistent membrane HER2 localization with a progressive signal decrease toward the core of the spheroids, more pronounced in BT474. Western blotting revealed divergent HER2 expression and AKT phosphorylation: SKBR3 spheroids displayed increased HER2 but reduced pAKT, while BT474 spheroids showed reduced HER2 and pAKT levels. EpCAM and E-cadherin staining revealed cell line-specific epithelial organization, and TEM demonstrated differences in intercellular spacing and mitochondrial morphology, reflecting spheroid compactness. Conclusions: 3D architecture profoundly influences HER2 distribution, signaling, and structural organization in HER2+ BCa spheroids. This model provides a robust platform for investigating architecture-dependent molecular processes, with potential applications in drug response, receptor trafficking, and targeted therapy evaluation
71° CONGRESSO NAZIONALE SIAI, Società di Anatomia e Istologia, Taormina, 20 - 22 Settembre 2017
No full textThe receptor tyrosine kinase ERBB2 interacts with HSP90 and is overexpressed in aggressive breast cancers. Therapeutic HSP90 inhibitors, i.e. Geldanamycin (GA), target ERBB2 to degradation. We have previously shown that HSP90 is responsible for the missorting of recycling ERBB2 to degradation compartments. In this study, we used biochemical, immunofluorescence and electron microscopy techniques to demonstrate that in SKBR3 human breast cancer cells, GA strongly induces polyubiquitination and internalization of the full-length p185-ERBB2, and promotes its cleavage, with the formation of a p116-ERBB2 form in EEA1-positive endosomes (EE). p116-ERBB2 corresponds to a non-ubiquitinated, signaling-impaired, membrane-bound fragment, which is readily sorted to lysosomes and degraded. To define the sequence of events leading to p116-ERBB2 degradation, we first blocked the EE maturation/trafficking to late endosomes/lysosomes with wortmannin, and found an increase in GA-dependent formation of p116-ERBB2; we then inhibited the proteasome activity with MG-132 or lactacystin, and observed an efficient block of p185-ERBB2 cleavage, and its accumulation in EE, suggesting that p185-ERBB2 polyubiquitination is necessary for proteasome-dependent p116-ERBB2 generation occurring in EE. As polyubiquitination has also been implicated in autophagy-mediated degradation of ERBB2 under different experimental conditions, we exploited this possibility and demonstrate that GA strongly inhibits early autophagy, and reduces the levels of the autophagy markers atg5-12 and LC3-II, irrespective of GA-induced ERBB2 polyubiquitination, ruling out a GA-dependent autophagic degradation of ERBB2. In conclusion, we propose that HSP90 inhibition fosters ERBB2 polyubiquitination and proteasome-dependent generation of a non-ubiquitinated and inactive p116-ERBB2 form in EE, which is trafficked from altered EE to lysosomes
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
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