Inheritable CRISPR based epigenetic modification in a fungus

The CRISPRoff system was recently introduced as a programmable epigenetic memory writer that can be used to silence genes in human cells. The system makes use of a dead Cas9 protein (dCas9) that is fused with the ZNF10 KRAB, Dnmt3A, and Dnmt3L protein domains. The DNA methylation resulting from the CRISPRoff system can be removed by the CRISPRon system that consists of dCas9 fused to the catalytic domain of Tet1. Here, the CRISPRoff and CRISPRon systems were applied for the first time in a fungus. The CRISPRoff system resulted in an inactivation up to 100 % of the target genes flbA and GFP in Aspergillus niger. Phenotypes correlated with the degree of gene silencing in the transformants and were stable when going through a conidiation cycle, even when the CRISPRoff plasmid was removed from the flbA silenced strain. Introducing the CRISPRon system in a strain in which the CRISPRoff plasmid was removed fully reactivated flbA showing a phenotype similar to that of the wildtype. Together, the CRISPRoff and CRISPRon systems can be used to study gene function in A. niger

The α-(1,3)-glucan synthase gene agsE impacts the secretome of Aspergillus niger

Aspergillus niger is widely used as a cell factory for the industrial production of enzymes. Previously, it was shown that deletion of α-1-3 glucan synthase genes results in smaller micro-colonies in liquid cultures of Aspergillus nidulans. Also, it has been shown that small wild-type Aspergillus niger micro-colonies secrete more protein than large mirco-colonies. We here assessed whether deletion of the agsC or agsE α-1-3 glucan synthase genes results in smaller A. niger micro-colonies and whether this is accompanied by a change in protein secretion. Biomass formation was not affected in the deletion strains but pH of the culture medium had changed from 5.2 in the case of the wild-type to 4.6 and 6.4 for ΔagsC and ΔagsE, respectively. The diameter of the ΔagsC micro-colonies was not affected in liquid cultures. In contrast, diameter of the ΔagsE micro-colonies was reduced from 3304 ± 338 µm to 1229 ± 113 µm. Moreover, the ΔagsE secretome was affected with 54 and 36 unique proteins with a predicted signal peptide in the culture medium of MA234.1 and the ΔagsE, respectively. Results show that these strains have complementary cellulase activity and thus may have complementary activity on plant biomass degradation. Together, α-1-3 glucan synthesis (in)directly impacts protein secretion in A. niger

Intracellular expression of Tat alters mitochondrial functions in T cells: a potential mechanism to understand mitochondrial damage during HIV-1 replication

HIV-1 replication results in mitochondrial damage that is enhanced during antiretroviral therapy (ART). The onset of HIV-1 replication is regulated by viral protein Tat, a 101-residue protein codified by two exons that elongates viral transcripts. Although the first exon of Tat (aa 1–72) forms itself an active protein, the presence of the second exon (aa 73–101) results in a more competent transcriptional protein with additional functions. Results: Mitochondrial overall functions were analyzed in Jurkat cells stably expressing full-length Tat (Tat101) or one-exon Tat (Tat72). Representative results were confirmed in PBLs transiently expressing Tat101 and in HIV-infected Jurkat cells. The intracellular expression of Tat101 induced the deregulation of metabolism and cytoskeletal proteins which remodeled the function and distribution of mitochondria. Tat101 reduced the transcription of the mtDNA, resulting in low ATP production. The total amount of mitochondria increased likely to counteract their functional impairment. These effects were enhanced when Tat second exon was expressed. Conclusions: Intracellular Tat altered mtDNA transcription, mitochondrial content and distribution in CD4+ T cells. The importance of Tat second exon in non-transcriptional functions was confirmed. Tat101 may be responsible for mitochondrial dysfunctions found in HIV-1 infected patients.We greatly appreciate the secretarial assistance of Mrs Olga Palao. This work was supported by FIPSE (360924/10), Spanish Ministry of Economy and Competitiveness (SAF2010-18388), Spanish Ministry of Health (EC11- 285), AIDS Network ISCIII-RETIC (RD12/0017/0015), Instituto de Salud Carlos III, Spanish Ministry of Economy and Competitiveness (FIS PI12/00506). The work of Sara Rodríguez-Mora is supported by a fellowship of Sara Borrell from Spanish Ministry of Economy and Competitiveness (2013). The work of María Rosa López-Huertas is supported by a fellowship of the European Union Programme Health 2009 (CHAARM).S

Effects of hospital facilities on patient outcomes after cancer surgery: an international, prospective, observational study

Background Early death after cancer surgery is higher in low-income and middle-income countries (LMICs) compared with in high-income countries, yet the impact of facility characteristics on early postoperative outcomes is unknown. The aim of this study was to examine the association between hospital infrastructure, resource availability, and processes on early outcomes after cancer surgery worldwide.Methods A multimethods analysis was performed as part of the GlobalSurg 3 study-a multicentre, international, prospective cohort study of patients who had surgery for breast, colorectal, or gastric cancer. The primary outcomes were 30-day mortality and 30-day major complication rates. Potentially beneficial hospital facilities were identified by variable selection to select those associated with 30-day mortality. Adjusted outcomes were determined using generalised estimating equations to account for patient characteristics and country-income group, with population stratification by hospital.Findings Between April 1, 2018, and April 23, 2019, facility-level data were collected for 9685 patients across 238 hospitals in 66 countries (91 hospitals in 20 high-income countries; 57 hospitals in 19 upper-middle-income countries; and 90 hospitals in 27 low-income to lower-middle-income countries). The availability of five hospital facilities was inversely associated with mortality: ultrasound, CT scanner, critical care unit, opioid analgesia, and oncologist. After adjustment for case-mix and country income group, hospitals with three or fewer of these facilities (62 hospitals, 1294 patients) had higher mortality compared with those with four or five (adjusted odds ratio [OR] 3.85 [95% CI 2.58-5.75]; p<0.0001), with excess mortality predominantly explained by a limited capacity to rescue following the development of major complications (63.0% vs 82.7%; OR 0.35 [0.23-0.53]; p<0.0001). Across LMICs, improvements in hospital facilities would prevent one to three deaths for every 100 patients undergoing surgery for cancer.Interpretation Hospitals with higher levels of infrastructure and resources have better outcomes after cancer surgery, independent of country income. Without urgent strengthening of hospital infrastructure and resources, the reductions in cancer-associated mortality associated with improved access will not be realised

Albiglutide and cardiovascular outcomes in patients with type 2 diabetes and cardiovascular disease (Harmony Outcomes): a double-blind, randomised placebo-controlled trial

Background: Glucagon-like peptide 1 receptor agonists differ in chemical structure, duration of action, and in their effects on clinical outcomes. The cardiovascular effects of once-weekly albiglutide in type 2 diabetes are unknown. We aimed to determine the safety and efficacy of albiglutide in preventing cardiovascular death, myocardial infarction, or stroke. Methods: We did a double-blind, randomised, placebo-controlled trial in 610 sites across 28 countries. We randomly assigned patients aged 40 years and older with type 2 diabetes and cardiovascular disease (at a 1:1 ratio) to groups that either received a subcutaneous injection of albiglutide (30–50 mg, based on glycaemic response and tolerability) or of a matched volume of placebo once a week, in addition to their standard care. Investigators used an interactive voice or web response system to obtain treatment assignment, and patients and all study investigators were masked to their treatment allocation. We hypothesised that albiglutide would be non-inferior to placebo for the primary outcome of the first occurrence of cardiovascular death, myocardial infarction, or stroke, which was assessed in the intention-to-treat population. If non-inferiority was confirmed by an upper limit of the 95% CI for a hazard ratio of less than 1·30, closed testing for superiority was prespecified. This study is registered with ClinicalTrials.gov, number NCT02465515. Findings: Patients were screened between July 1, 2015, and Nov 24, 2016. 10 793 patients were screened and 9463 participants were enrolled and randomly assigned to groups: 4731 patients were assigned to receive albiglutide and 4732 patients to receive placebo. On Nov 8, 2017, it was determined that 611 primary endpoints and a median follow-up of at least 1·5 years had accrued, and participants returned for a final visit and discontinuation from study treatment; the last patient visit was on March 12, 2018. These 9463 patients, the intention-to-treat population, were evaluated for a median duration of 1·6 years and were assessed for the primary outcome. The primary composite outcome occurred in 338 (7%) of 4731 patients at an incidence rate of 4·6 events per 100 person-years in the albiglutide group and in 428 (9%) of 4732 patients at an incidence rate of 5·9 events per 100 person-years in the placebo group (hazard ratio 0·78, 95% CI 0·68–0·90), which indicated that albiglutide was superior to placebo (p<0·0001 for non-inferiority; p=0·0006 for superiority). The incidence of acute pancreatitis (ten patients in the albiglutide group and seven patients in the placebo group), pancreatic cancer (six patients in the albiglutide group and five patients in the placebo group), medullary thyroid carcinoma (zero patients in both groups), and other serious adverse events did not differ between the two groups. There were three (<1%) deaths in the placebo group that were assessed by investigators, who were masked to study drug assignment, to be treatment-related and two (<1%) deaths in the albiglutide group. Interpretation: In patients with type 2 diabetes and cardiovascular disease, albiglutide was superior to placebo with respect to major adverse cardiovascular events. Evidence-based glucagon-like peptide 1 receptor agonists should therefore be considered as part of a comprehensive strategy to reduce the risk of cardiovascular events in patients with type 2 diabetes. Funding: GlaxoSmithKline

Search for anomalous couplings in boosted WW/WZ -> l nu q(q)over-bar production in proton-proton collisions at root s=8TeV

Peer reviewe

Measurement of t(t)over-bar normalised multi-differential cross sections in pp collisions at root s=13 TeV, and simultaneous determination of the strong coupling strength, top quark pole mass, and parton distribution functions

Peer reviewe

An embedding technique to determine ττ backgrounds in proton-proton collision data

An embedding technique is presented to estimate standard model tau tau backgrounds from data with minimal simulation input. In the data, the muons are removed from reconstructed mu mu events and replaced with simulated tau leptons with the same kinematic properties. In this way, a set of hybrid events is obtained that does not rely on simulation except for the decay of the tau leptons. The challenges in describing the underlying event or the production of associated jets in the simulation are avoided. The technique described in this paper was developed for CMS. Its validation and the inherent uncertainties are also discussed. The demonstration of the performance of the technique is based on a sample of proton-proton collisions collected by CMS in 2017 at root s = 13 TeV corresponding to an integrated luminosity of 41.5 fb(-1).Peer reviewe

Measurement of the Splitting Function in &ITpp &ITand Pb-Pb Collisions at root&ITsNN&IT=5.02 TeV

Data from heavy ion collisions suggest that the evolution of a parton shower is modified by interactions with the color charges in the dense partonic medium created in these collisions, but it is not known where in the shower evolution the modifications occur. The momentum ratio of the two leading partons, resolved as subjets, provides information about the parton shower evolution. This substructure observable, known as the splitting function, reflects the process of a parton splitting into two other partons and has been measured for jets with transverse momentum between 140 and 500 GeV, in pp and PbPb collisions at a center-of-mass energy of 5.02 TeV per nucleon pair. In central PbPb collisions, the splitting function indicates a more unbalanced momentum ratio, compared to peripheral PbPb and pp collisions.. The measurements are compared to various predictions from event generators and analytical calculations.Peer reviewe

Measurement of nuclear modification factors of gamma(1S)), gamma(2S), and gamma(3S) mesons in PbPb collisions at root s(NN)=5.02 TeV

The cross sections for ϒ(1S), ϒ(2S), and ϒ(3S) production in lead-lead (PbPb) and proton-proton (pp) collisions at √sNN = 5.02 TeV have been measured using the CMS detector at the LHC. The nuclear modification factors, RAA, derived from the PbPb-to-pp ratio of yields for each state, are studied as functions of meson rapidity and transverse momentum, as well as PbPb collision centrality. The yields of all three states are found to be significantly suppressed, and compatible with a sequential ordering of the suppression, RAA(ϒ(1S)) > RAA(ϒ(2S)) > RAA(ϒ(3S)). The suppression of ϒ(1S) is larger than that seen at √sNN = 2.76 TeV, although the two are compatible within uncertainties. The upper limit on the RAA of ϒ(3S) integrated over pT, rapidity and centrality is 0.096 at 95% confidence level, which is the strongest suppression observed for a quarkonium state in heavy ion collisions to date. © 2019 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). Funded by SCOAP3.Peer reviewe