The Crystal Structure of the SV40 T-Antigen Origin Binding Domain in Complex with DNA

DNA replication is initiated upon binding of “initiators” to origins of replication. In simian virus 40 (SV40), the core origin contains four pentanucleotide binding sites organized as pairs of inverted repeats. Here we describe the crystal structures of the origin binding domain (obd) of the SV40 large T-antigen (T-ag) both with and without a subfragment of origin-containing DNA. In the co-structure, two T-ag obds are oriented in a head-to-head fashion on the same face of the DNA, and each T-ag obd engages the major groove. Although the obds are very close to each other when bound to this DNA target, they do not contact one another. These data provide a high-resolution structural model that explains site-specific binding to the origin and suggests how these interactions help direct the oligomerization events that culminate in assembly of the helicase-active dodecameric complex of T-ag

Effect of Hydrocortisone on Mortality and Organ Support in Patients With Severe COVID-19: The REMAP-CAP COVID-19 Corticosteroid Domain Randomized Clinical Trial.

Importance: Evidence regarding corticosteroid use for severe coronavirus disease 2019 (COVID-19) is limited. Objective: To determine whether hydrocortisone improves outcome for patients with severe COVID-19. Design, Setting, and Participants: An ongoing adaptive platform trial testing multiple interventions within multiple therapeutic domains, for example, antiviral agents, corticosteroids, or immunoglobulin. Between March 9 and June 17, 2020, 614 adult patients with suspected or confirmed COVID-19 were enrolled and randomized within at least 1 domain following admission to an intensive care unit (ICU) for respiratory or cardiovascular organ support at 121 sites in 8 countries. Of these, 403 were randomized to open-label interventions within the corticosteroid domain. The domain was halted after results from another trial were released. Follow-up ended August 12, 2020. Interventions: The corticosteroid domain randomized participants to a fixed 7-day course of intravenous hydrocortisone (50 mg or 100 mg every 6 hours) (n = 143), a shock-dependent course (50 mg every 6 hours when shock was clinically evident) (n = 152), or no hydrocortisone (n = 108). Main Outcomes and Measures: The primary end point was organ support-free days (days alive and free of ICU-based respiratory or cardiovascular support) within 21 days, where patients who died were assigned -1 day. The primary analysis was a bayesian cumulative logistic model that included all patients enrolled with severe COVID-19, adjusting for age, sex, site, region, time, assignment to interventions within other domains, and domain and intervention eligibility. Superiority was defined as the posterior probability of an odds ratio greater than 1 (threshold for trial conclusion of superiority >99%). Results: After excluding 19 participants who withdrew consent, there were 384 patients (mean age, 60 years; 29% female) randomized to the fixed-dose (n = 137), shock-dependent (n = 146), and no (n = 101) hydrocortisone groups; 379 (99%) completed the study and were included in the analysis. The mean age for the 3 groups ranged between 59.5 and 60.4 years; most patients were male (range, 70.6%-71.5%); mean body mass index ranged between 29.7 and 30.9; and patients receiving mechanical ventilation ranged between 50.0% and 63.5%. For the fixed-dose, shock-dependent, and no hydrocortisone groups, respectively, the median organ support-free days were 0 (IQR, -1 to 15), 0 (IQR, -1 to 13), and 0 (-1 to 11) days (composed of 30%, 26%, and 33% mortality rates and 11.5, 9.5, and 6 median organ support-free days among survivors). The median adjusted odds ratio and bayesian probability of superiority were 1.43 (95% credible interval, 0.91-2.27) and 93% for fixed-dose hydrocortisone, respectively, and were 1.22 (95% credible interval, 0.76-1.94) and 80% for shock-dependent hydrocortisone compared with no hydrocortisone. Serious adverse events were reported in 4 (3%), 5 (3%), and 1 (1%) patients in the fixed-dose, shock-dependent, and no hydrocortisone groups, respectively. Conclusions and Relevance: Among patients with severe COVID-19, treatment with a 7-day fixed-dose course of hydrocortisone or shock-dependent dosing of hydrocortisone, compared with no hydrocortisone, resulted in 93% and 80% probabilities of superiority with regard to the odds of improvement in organ support-free days within 21 days. However, the trial was stopped early and no treatment strategy met prespecified criteria for statistical superiority, precluding definitive conclusions. Trial Registration: ClinicalTrials.gov Identifier: NCT02735707

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

DisLog: A Separation Logic for Disentanglement - Artifact

This is the artifact corresponding to the article entitled "DisLog: A Separation Logic for Disentanglement", and its associated documentation

DisLog: A Separation Logic for Disentanglement

International audienceDisentanglement is a run-time property of parallel programs that facilitates task-local reasoning about the memory footprint of parallel tasks. In particular, it ensures that a task does not access any memory locations allocated by another concurrently executing task. Disentanglement can be exploited, for example, to implement a high-performance parallel memory manager, such as in the MPL (MaPLe) compiler for Parallel ML. Prior research on disentanglement has focused on the design of optimizations, either trusting the programmer to provide a disentangled program or relying on runtime instrumentation for detecting and managing entanglement. This paper provides the first static approach to verify that a program is disentangled: it contributes DisLog, a concurrent separation logic for disentanglement. DisLog enriches concurrent separation logic with the notions necessary for reasoning about the fork-join structure of parallel programs, allowing the verification that memory accesses are effectively disentangled. A large class of programs, including race-free programs, exhibit memory access patterns that are disentangled "by construction". To reason about these patterns, the paper distills from DisLog an almost standard concurrent separation logic, called DisLog+. In this high-level logic, no specific reasoning about memory accesses is needed: functional correctness proofs entail disentanglement. The paper illustrates the use of DisLog and DisLog+ on a range of case studies, including two different implementations of parallel deduplication via concurrent hashing. All our results are mechanized in the Coq proof assistant using Iris

Approches sensorielle et analytique de l’arôme fruité des vins rouges. Influence majeure de la souche de levures.

Article dans une revue professionnelleLa typicité se définit comme la propriété d’appartenance à un type distingué et identifié par un groupe humain de référence (Casabianca et al., 2005). Ainsi, il est depuis longtemps observé de façon empirique l’existence d’une expression fruitée des vins rouges, et, de façon plus spécifique aux vins de Bordeaux, de notes évoquant les fruits rouges et noirs comme la framboise, la cerise ou encore le cassis. Récemment, l’existence d’un espace sensoriel propre à ces vins a été démontrée (Pineau, 2007). Ces notes fruitées, non détectées dans le moût, sont révélées au cours des différentes étapes de vinification et d’élaboration des vins. Les fermentations alcoolique et malolactique constituent des étapes clés dans la libération ou la genèse des arômes du vin. Plusieurs travaux ont tenté de déterminer l’influence des souches de levures ou de bactéries lactiques (BL) sur la modulation organoleptique du vin sans qu’aucun consensus ne puisse encore être établi (Álvarez-Pérez et al., 2012 ; McDaniel et al., 1987; Henick-Kling et al., 1994 ; Antalick et al., 2012). L’interprétation moléculaire de cette spécificité des vins rouges de Bordeaux est rendue difficile par le manque de connaissances sur les composés aromatiques responsables de ces nuances. Depuis plusieurs décennies, de nombreux travaux se sont attachés à la caractérisation moléculaire des composés volatils du vin. Les premières études réalisées sur des vins blancs ont permis de faire l’association entre certains arômes caractéristiques et une ou plusieurs molécules d’intérêt (Marais, 1983 ; Tominaga et al., 1998). De nombreux auteurs ont abordé l’étude de l’arôme fruité des vins rouges en tentant d’identifier les composés aromatiques clés responsables de ces odeurs (Moio et Etievant, 1995 ; Kotseridis et Baumes, 2000 ; Campo et al., 2007 ; Siebert et al., 2008). S’il n’a pas encore été possible d’identifier des molécules spécifiques, expliquant à elles seules la typicité de l’arôme fruité, l’existence de tels composés ne peut pour autant être définitivement exclue. Néanmoins, plusieurs travaux récents ont démontré l’importance des interactions perceptives entre plusieurs familles d’odorants tels que des composés d’origine variétale comme les norisoprénoïdes à 13 atomes de carbone (Pineau et al., 2007), les lactones (Loscos et al., 2007) ou les thiols (Tominaga et al., 1998), ou des composés fermentaires comme les esters (Lytra et al., 2012). Par ailleurs, plusieurs études ont visé à déterminer l’impact de micro-organismes fermentaires sur la modulation des teneurs en divers marqueurs potentiels de l’arôme fruité. Là encore, aucun consensus n’a pu réellement être mis en évidence. La variation de la note fruitée des vins rouges semble impliquer des phénomènes plus complexes qu’un simple effet souche et pourrait résulter d’effets liés aux propriétés de la matrice, combinés à l’influence de la souche de levures et/ou de la souche de BL. Au vu de ces dernières acquisitions, nous avons choisi, dans le cadre de ce travail, d’étudier l’impact organoleptique relatif des levures et des BL sur les nuances de l’arôme fruité de plusieurs vins rouges bordelais. Une approche par l’analyse chimique et sensorielle de vins produits dans des conditions de fermentation classique ou de microvinification a été effectuée pour trois sites en Bordelais sur deux millésimes. Il a été choisi de travailler sur le Cabernet- Sauvignon. Plusieurs couples levures/BL ont été sélectionnés et implantés dans des cuves de quelques hectolitres à plusieurs dizaines d’hectolitres

Peptides Containing Cyclin/Cdk-Nuclear Localization Signal Motifs Derived from Viral Initiator Proteins Bind to DNA When Unphosphorylated

A single phosphorylation event at T-antigen residue Thr124 regulates initiation of simian virus 40 DNA replication. To explore this regulatory process, a series of peptides were synthesized, centered on Thr124. These peptides contain a nuclear localization signal (NLS) and a recognition site for cyclin/Cdk kinases. When unphosphorylated, the “CDK/NLS” peptides inhibit T-antigen assembly and bind non-sequence specifically to DNA. However, these activities are greatly reduced upon phosphorylation of Thr124. Similar results were obtained by using peptides derived from the CDK/NLS region of bovine papillomavirus E1. Related studies indicate that residues in the NLS bind to DNA, whereas those in the CDK motif regulate binding. These findings are discussed in terms of the control of T-antigen double hexamer assembly and initiation of viral replication

Evidence for a Structural Relationship between BRCT Domains and the Helicase Domains of the Replication Initiators Encoded by the Polyomaviridae and Papillomaviridae Families of DNA Tumor Viruses▿ †

Studies of DNA tumor viruses have provided important insights into fundamental cellular processes and oncogenic transformation. They have revealed, for example, that upon expression of virally encoded proteins, cellular pathways involved in DNA repair and cell cycle control are disrupted. Herein, evidence is presented that BRCT-related regions are present in the helicase domains of the viral initiators encoded by the Polyomaviridae and Papillomaviridae viral families. Of interest, BRCT domains in cellular proteins recruit factors involved in diverse pathways, including DNA repair and the regulation of cell cycle progression. Therefore, the viral BRCT-related regions may compete with host BRCT domains for particular cellular ligands, a process that would help to explain the pleiotropic effects associated with infections with many DNA tumor viruses

Analyses of the Interaction between the Origin Binding Domain from Simian Virus 40 T Antigen and Single-Stranded DNA Provide Insights into DNA Unwinding and Initiation of DNA Replication

DNA helicases are essential for DNA metabolism; however, at the molecular level little is known about how they assemble or function. Therefore, as a model for a eukaryotic helicase, we are analyzing T antigen (T-ag) the helicase encoded by simian virus 40. In this study, nuclear magnetic resonance (NMR) methods were used to investigate the transit of single-stranded DNA (ssDNA) through the T-ag origin-binding domain (T-ag OBD). When the residues that interact with ssDNA are viewed in terms of the structure of a hexamer of the T-ag OBD, comprised of residues 131 to 260, they indicate that ssDNA passes over one face of the T-ag OBD and then transits through a gap in the open ring structure. The NMR-based conclusions are supported by an analysis of previously described mutations that disrupt critical steps during the initiation of DNA replication. These and related observations are discussed in terms of the threading of DNA through T-ag hexamers and the initiation of viral DNA replication