1,720,979 research outputs found
HIV-1 Env and HLA-C interaction is crucial in modulating viral infectivity
Full text linkIntroduction HLA-C plays a crucial role in the progression of HIV-1 infection. Host genetic HLA-C variants, appear to be associated with a different ability to control HIV-1 infection. A higher HLA-C expression is associated with a better activation of cytotoxic T lymphocytes (CTLs) and of Killer Immunoglobulin like receptors (KIR) on NK-cells, which lead to a better HIV-1 infection control. Vice-versa, a lower HLA-C expression leads to a rapid progression toward AIDS. In addition, different HLA-C alleles present different binding stabilities to β2microglobulin (β2m)/peptide. Noteworthy, some HLA-C highly expressed/protective alleles are also stably bound to β2m/peptide, while some low expressed/non-protective variants present an unstable bond to β2m/peptide. Finally virions lacking HLA-C have reduced infectivity and increased susceptibility to neutralizing antibodies. Experiments In the present work, it was first characterized the association between HLA-C and HIV-1 Env. We investigated if HIV-1 infection involves HLA-C free chains or the heterotrimeric complex, and to this purpose the A3.01 cell line and its HIV-1-infected counterpart ACH-2, as well as PM1 cells, were used as in vitro infection model. HEK-293T β2m negative cells, generated using CRISPR/Cas9 system, were used to produce HIV-1 pseudoviruses and to test their infectivity. Then, the proportion between HLA-C associated to β2m and HLA-C presents as free chains on the cell surface was characterized on PBMC from healthy donors, bearing both Stable or Unstable HLA-C alleles. In addition, PBMC were tested for their ability to support HIV-1 infection in vitro. Results HIV-1 infection induces the appearance of HLA-C free chains on the surface of infected cells, which may be responsible for the increased HIV-1 infectivity. HIV-1 Env-pseudotyped viruses produced in the absence of β2m, thus lacking HLA-C on their envelope, were less infectious than those produced in the presence of β2m. By analysing PBMC from healthy donors, differences in HLA-C heterotrimers stability and HLA-C expression levels were found. Finally, it was reported that R5 HIV-1 virions produced by PBMC having Unstable HLA-C alleles were more infectious than those produced by PBMC having the Stable variants. Conclusions The outcome of HIV-1 infection might depend both on the HLA-C surface expression levels and on HLA-C/β2m/peptide binding stability. According to this model, PBMC carrying low expressed/Unstable HLA-C alleles have a high proportion of HLA-C free chains on their surface that raises viral infectivity and, at the same time, a low proportion of HLA-C heterotrimeric complexes which leads to a poor control of HIV-1 infection, and thus to a rapid progression toward AIDS.È noto che alcuni polimorfismi del sistema HLA giocano un ruolo cruciale nell’eziopatogenesi e nella prognosi di numerose malattie infettive, fra le quali l’AIDS (Sindrome da Immunodeficienza Acquisita). Recenti studi hanno evidenziato una forte correlazione fra i livelli di espressione di HLA-C e il controllo della replicazione del virus dell’immunodeficienza umana (HIV-1). Alti livelli di espressione sono stati correlati con un miglior controllo dell’infezione, mentre bassi livelli sono stati associati con una progressione più rapida della malattia. Inoltre, è noto che la molecola HLA-C, presente sull’envelope di HIV-1, in associazione con la glicoproteina Env, è in grado di aumentarne l’infettività. Il ruolo protettivo di alti livelli di espressione di HLA-C sembra essere in contraddizione con il ruolo dell’ HLA-C stesso nell’aumentare l’infettività virale quando incorporato nel virione. Ciò potrebbe essere dovuto alla presenza di diverse conformazioni dell’ HLA-C. È infatti noto che diverse varianti alleliche dell’HLA-C presentano una diversa stabilità di legame con la β2 microglobulina (β2m) e il peptide. In particolare, l’HLA-C può presentarsi associato alla β2m e al peptide, costituendo un complesso che svolge un ruolo chiave nell’attivazione del sistema immunitario, oppure come free chain, dissociato dal complesso. I primi risultati di questo lavoro hanno dimostrato che la proteina Env di HIV-1 è in grado di associarsi all’HLA-C quando presente nella conformazione di free chain. L’ipotesi testata nello studio prevede l’esistenza di un’associazione fra la suscettibilità all’infezione da HIV-1 e le diverse varianti alleliche di HLA-C che possono essere preferibilmente presenti o come complesso trimerico o come free chain. Individui con varianti di HLA-C aventi una forte stabilità come trimero completo mostrerebbero una maggiore immunità contro HIV-1 e una ridotta infettività virale, mentre soggetti con varianti dell’HLA-C che facilmente si dissociano dalla β2m e dal peptide mostrerebbero una ridotta risposta immunitaria nei confronti di HIV-1 e la produzione di virioni maggiormente infettivi. Nel suo complesso, questo studio fornisce nuove informazioni che potrebbero rivelarsi utili per la progettazione di nuove strategie vaccinali e approcci terapeutici contro HIV-1
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
In silico structural prediction and in vitro functional validation to identify the ACOT8 regions involved in the interaction with HIV-1 Nef
No full textIntroductionHIV-1 Nef interacts with several cellular proteins, among which the human peroxisomal thioesterase 8 (ACOT8). This interaction appears to influence the CD4 down-regulation and might modulate lipid composition of membrane proteins during HIV-1 infection. The Nef regions involved in the association with ACOT8 have been experimentally characterized. The lack of structural information for ACOT8 limits the full comprehension of the biological role of the Nef/ACOT8 association relevant to HIV-1 infection.ResultsIn this work we modelled, through in silico predictions, the ACOT8 structure. A high charge complementarity was observed between Nef and ACOT8 surfaces. This allowed the identification of the ACOT8 aminoacids most likely involved in the interaction with Nef. They map in the Arg45-Phe55 and Arg86-Pro93 ACOT8 regions. Their role has been validated by in vitro assays through the development of ACOT8 deletion mutants. Immunofluorescence and co-immunoprecipitation analyses showed that the ACOT8 K91S mutation is sufficient to abrogate the interaction with Nef. In addition, the ACOT8 Arg45-Phe55 region, as well as the Arg86-Pro93 region, are involved in Nef binding.ConclusionsOur data demonstrate that the ACOT8 Lys91 plays a key role in the interaction with Nef. The observation that both ACOT8 Arg45-Phe55 and Arg86-Pro93 regions are determinant for Nef association suggests that the interaction involves a wider region on ACOT8 surface. These findings improve the comprehension of the association between HIV-1 Nef and ACOT8 and will help elucidating the biological meaning of their interaction
CRISPR/Cas9 as a tool for the study of the interactions between viruses and host
No full textCRISPR/Cas9 in virolog
CRISPR/Cas9 as a tool for studying the interactions between viral and cellular proteins
No full textThe CRISPR/Cas9 system is a new promising technique that allows editing of DNA
sequences in the cell genome. We used CRISPR/Cas9 to produce β2microglobulin (β2m) and
human thioesterase 8 (ACOT8) knock-out cell lines to study their role in HIV-1 infection. It
has been reported that HLA-C interacts with HIV-1 Env, increasing virus infectivity (D.
Zipeto, A. Beretta. 2012), while ACOT8 interacts with HIV-1 Nef, increasing Nef stability
(M. Serena et al. 2016).
The loss of β2m expression in 293T, HeLa-Lai (expressing HIV-1 Env) and parental HeLa
cells was assessed by western blot and flow cytometry; β2m negative cells were finally sorted.
Cytofluorimetric analysis indicated that, in the absence of β2m, HLA-C expression on the cell
membrane was abrogated, both in HeLa and in HeLa-Lai cells. The presence of HIV-1 Env
did not restore HLA-C on the cell surface, suggesting that HLA-C needs b2m to be expressed
on the cell membrane, where the Env/HLA-C interaction occurs. In addition, we observed that
HIV-1 viruses produced in 293T b2m negative cells were three fold less infectious than those
produced in the parental cells, suggesting that the presence of HLA-C on the cell surface is
required for the association with Env, thus resulting in an increased virus infectivity.
The loss of ACOT8 expression in 293T cells and in TZM-bl, a cell line highly sensitive to
infection with HIV-1, was assessed by western blot. Cells were clonally expanded and
ACOT8 knock out was further confirmed in immunoblot. The role of Nef/ACOT8 association
will be explored comparing the infectivity of HIV-1 viruses produced in the presence or in the
absence of ACOT8 (293T), as well as com
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
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