In vivo inactivation of Nef ITAM motif of chimeric simian/human immunodeficiency virus SHIVsbg-YE correlates with absence of increased virulence in chinese rhesus macaques

AbstractSHIVsbg, expressing Vpu, Tat, Rev, and Env proteins of HIV-1 Lai, was shown to be infectious for rhesus macaques. In this study, we mutated SHIVsbg Nef amino acids 17–18 from RQ to YE, conferring to SHIVsbg-YE the ability to replicate in vitro in unstimulated macaque PBMC. Juvenile macaques inoculated intravenously or orally with SHIVsbg-YE developed persistent infection. All macaques lost weight during the first 17 weeks but recovered afterward. All animals developed a strong HIV-specific humoral immune response. Viruses isolated 2 years postinoculation lost the ability to replicate in unstimulated macaque PBMC. Point mutations or 33-bp-wide deletions in the nef ITAM motif were responsible for this phenotype and correlated with clinical improvement of the infected macaques. These data demonstrate that the ITAM domain is inactivated in animals developing an acute antiviral immune response and may be detrimental to viral replication, perhaps by interfering with other well-conserved functions of SIV Nef protein

A whole-genome sequence and transcriptome perspective on HER2-positive breast cancers.

HER2-positive breast cancer has long proven to be a clinically distinct class of breast cancers for which several targeted therapies are now available. However, resistance to the treatment associated with specific gene expressions or mutations has been observed, revealing the underlying diversity of these cancers. Therefore, understanding the full extent of the HER2-positive disease heterogeneity still remains challenging. Here we carry out an in-depth genomic characterization of 64 HER2-positive breast tumour genomes that exhibit four subgroups, based on the expression data, with distinctive genomic features in terms of somatic mutations, copy-number changes or structural variations. The results suggest that, despite being clinically defined by a specific gene amplification, HER2-positive tumours melt into the whole luminal-basal breast cancer spectrum rather than standing apart. The results also lead to a refined ERBB2 amplicon of 106 kb and show that several cases of amplifications are compatible with a breakage-fusion-bridge mechanism

Genome-Wide Association Study in a Lebanese Cohort Confirms PHACTR1 as a Major Determinant of Coronary Artery Stenosis

The manifestation of coronary artery disease (CAD) follows a well-choreographed series of events that includes damage of arterial endothelial cells and deposition of lipids in the sub-endothelial layers. Genome-wide association studies (GWAS) of multiple populations with distinctive genetic and lifestyle backgrounds are a crucial step in understanding global CAD pathophysiology. In this study, we report a GWAS on the genetic basis of arterial stenosis as measured by cardiac catheterization in a Lebanese population. The locus of the phosphatase and actin regulator 1 gene (PHACTR1) showed association with coronary stenosis in a discovery experiment with genome wide data in 1,949 individuals (rs9349379, OR = 1.37, p = 1.57×10−5). The association was replicated in an additional 2,547 individuals (OR = 1.31, p = 8.85×10−6), leading to genome-wide significant association in a combined analysis (OR = 1.34, p = 8.02×10−10). Results from this GWAS support a central role of PHACTR1 in CAD susceptibility irrespective of lifestyle and ethnic divergences. This association provides a plausible component for understanding molecular mechanisms involved in the formation of stenosis in cardiac vessels and a potential drug target against CAD

A whole-genome sequence and transcriptome perspective on HER2-positive breast cancers

HER2-positive breast cancer has long proven to be a clinically distinct class of breast cancers for which several targeted therapies are now available. However, resistance to the treatment associated with specific gene expressions or mutations has been observed, revealing the underlying diversity of these cancers. Therefore, understanding the full extent of the HER2-positive disease heterogeneity still remains challenging. Here we carry out an in-depth genomic characterization of 64 HER2-positive breast tumour genomes that exhibit four subgroups, based on the expression data, with distinctive genomic features in terms of somatic mutations, copy-number changes or structural variations. The results suggest that, despite being clinically defined by a specific gene amplification, HER2-positive tumours melt into the whole luminal-basal breast cancer spectrum rather than standing apart. The results also lead to a refined ERBB2 amplicon of 106 kb and show that several cases of amplifications are compatible with a breakage-fusion-bridge mechanism

Abdominal aortic aneurysm is associated with a variant in low-density lipoprotein receptor-related protein 1

Abdominal aortic aneurysm (AAA) is a common cause of morbidity and mortality and has a significant heritability. We carried out a genome-wide association discovery study of 1866 patients with AAA and 5435 controls and replication of promising signals (lead SNP with a p value < 1 × 10-5) in 2871 additional cases and 32,687 controls and performed further follow-up in 1491 AAA and 11,060 controls. In the discovery study, nine loci demonstrated association with AAA (p < 1 × 10-5). In the replication sample, the lead SNP at one of these loci, rs1466535, located within intron 1 of low-density-lipoprotein receptor-related protein 1 (LRP1) demonstrated significant association (p = 0.0042). We confirmed the association of rs1466535 and AAA in our follow-up study (p = 0.035). In a combined analysis (6228 AAA and 49182 controls), rs1466535 had a consistent effect size and direction in all sample sets (combined p = 4.52 × 10-10, odds ratio 1.15 [1.10-1.21]). No associations were seen for either rs1466535 or the 12q13.3 locus in independent association studies of coronary artery disease, blood pressure, diabetes, or hyperlipidaemia, suggesting that this locus is specific to AAA. Gene-expression studies demonstrated a trend toward increased LRP1 expression for the rs1466535 CC genotype in arterial tissues; there was a significant (p = 0.029) 1.19-fold (1.04-1.36) increase in LRP1 expression in CC homozygotes compared to TT homozygotes in aortic adventitia. Functional studies demonstrated that rs1466535 might alter a SREBP-1 binding site and influence enhancer activity at the locus. In conclusion, this study has identified a biologically plausible genetic variant associated specifically with AAA, and we suggest that this variant has a possible functional role in LRP1 expression

A Solve-RD ClinVar-based reanalysis of 1522 index cases from ERN-ITHACA reveals common pitfalls and misinterpretations in exome sequencing

Purpose Within the Solve-RD project (https://solve-rd.eu/), the European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies aimed to investigate whether a reanalysis of exomes from unsolved cases based on ClinVar annotations could establish additional diagnoses. We present the results of the “ClinVar low-hanging fruit” reanalysis, reasons for the failure of previous analyses, and lessons learned. Methods Data from the first 3576 exomes (1522 probands and 2054 relatives) collected from European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies was reanalyzed by the Solve-RD consortium by evaluating for the presence of single-nucleotide variant, and small insertions and deletions already reported as (likely) pathogenic in ClinVar. Variants were filtered according to frequency, genotype, and mode of inheritance and reinterpreted. Results We identified causal variants in 59 cases (3.9%), 50 of them also raised by other approaches and 9 leading to new diagnoses, highlighting interpretation challenges: variants in genes not known to be involved in human disease at the time of the first analysis, misleading genotypes, or variants undetected by local pipelines (variants in off-target regions, low quality filters, low allelic balance, or high frequency). Conclusion The “ClinVar low-hanging fruit” analysis represents an effective, fast, and easy approach to recover causal variants from exome sequencing data, herewith contributing to the reduction of the diagnostic deadlock

Etude des étapes précoces de l'infection par le virus de l'Hépatite C (Mécanismes de réplication du génome viral)

Le virus de l'hépatite C (VHC) infecte 3% de la population mondiale. L'infection par ce virus devient chronique dans une grande majorité des cas puis peut évoluer vers une cirrhose ou un hépatocarcinome. Deux étapes successives sont indispensables à la réplication du génome de ce virus à ARN de polarité positive : (1) l'étape de traduction du brin génomique permet de produire toutes les protéines virales, puis, (2) le complexe de réplication, reconnaît la région 3' terminale du génome et permet la synthèse du brin complémentaire ou brin (-). Le brin (-) servira ensuite de matrice au complexe de réplication pour produire un nouveau génome ou ARN (+). Dans notre travail, certains aspects de la traduction et de la réplication du génome viral ont été abordés. Dans un premier temps, nous avons étudié la régulation de la traduction du génome du VHC. Nous avons montré que ni la région 3'(+) ni l'expression des protéines virales n'influençaient l'efficacité de traduction du génome viral. Afin de mieux comprendre le mécanisme de réplication de ce virus, nous avons, dans un deuxième temps, entrepris une étude structurale de l'extrémité 3' du brin (-). La structure secondaire de cette région d'ARN a été déterminée en solution, en utilisant des sondes chimiques et enzymatiques. Cette région très structurée est reconnue par le complexe de réplication. La connaissance de sa structure nous permettra donc, de proposer des hypothèses concernant l'importance de certains nucléotides ou groupes de nucléotides dans la réplication de ce virus. En parallèle, une étude des interactions existant entre les protéines non structurales du virus (protéines formant le complexe de réplication) a été entreprise et a montré qu'elles interagissaient toutes les unes avec les autres. La dernière partie de ce travail présente dans un contexte ex vivo, différentes approches développées au laboratoire afin d'étudier la réplication du VHC et en particulier la synthèse du brin (+) à partir du brin (-).The Hepatitis C Virus (HCV) infects 3% of the world population. Virus infection becomes chronic in a large majority of cases and can evolve to cirrhosis or hepatocarcinoma. Two successive steps are essential for the replication of this positive strand RNA virus: (1) the viral genome translation which produces all viral proteins, and the (2) further recognition of the 3' non translated region (or 3'(+)) by the replication complex which allows the complementary (or (-)) strand synthesis. The minus strand is then used to produce new genomic RNA (or (+)) strands. Our work is focused on some translation and replication aspects. First, translation regulation of the genomic RNA was analyzed. We show that neither the 3'(+) region nor the viral proteins modulate the translation efficiency. In order to learn more about replication mechanisms, we studied in solution, the 3'(-) end folding with enzymatic or chemical probes. This well structured region is recognized by the replication complex. This knowledge allows us thus to propose nucleotides or groups of nucleotides essential for the viral replication. In parallel, we analyzed protein/protein interactions between the non structural proteins (which form the replication complex). We show that all non structural proteins interact ones with the others. The last part of my study concerns various approaches developed in order to study the HCV replication, and in particular the (+) strand synthesis, in an ex vivo context.STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF

Transcription-Dependent Generation of a Specialized Chromatin Structure at the TCRβ Locus

International audienceno abstrac

Detection of Human Immunodeficiency Virus Type 1 Nef and CD4 Physical Interaction in Living Human Cells by Using Bioluminescence Resonance Energy Transfer

CD4 down-regulation by human immunodeficiency virus type 1 (HIV-1) Nef protein is a key function for virus virulence. This activity may be mediated by a direct Nef-CD4 interaction. We investigated the formation, in situ, of such a complex between proteins using bioluminescence resonance energy transfer technology and coimmunoprecipitations. Our data clearly demonstrate that Nef and CD4 interact in intact human cells. Moreover, our results clearly indicate that the dileucine motif of the CD4 cytoplasmic domain, critical for the Nef-induced CD4 down-regulation, is not implicated in the Nef/CD4 complex formation in the cellular context

Salles de sport: principes de planification

Bases de planification appropriées; dimensionnement des salles, terrains de jeu et locaux annexes. Indications concernant l’éclairage, la ventilation, l’accoustique etc