Biochemical, structural and functional studies on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) papain-like protease PLpro reveal that it regulates host antiviral responses by... Show moreBiochemical, structural and functional studies on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) papain-like protease PLpro reveal that it regulates host antiviral responses by preferentially cleaving the ubiquitin-like interferon-stimulated gene 15 protein (ISG15) and identify this protease as a potential therapeutic target for coronavirus disease 2019 (COVID-19).The papain-like protease PLpro is an essential coronavirus enzyme that is required for processing viral polyproteins to generate a functional replicase complex and enable viral spread(1,2). PLpro is also implicated in cleaving proteinaceous post-translational modifications on host proteins as an evasion mechanism against host antiviral immune responses(3-5). Here we perform biochemical, structural and functional characterization of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PLpro (SCoV2-PLpro) and outline differences with SARS-CoV PLpro (SCoV-PLpro) in regulation of host interferon and NF-kappa B pathways. SCoV2-PLpro and SCoV-PLpro share 83% sequence identity but exhibit different host substrate preferences; SCoV2-PLpro preferentially cleaves the ubiquitin-like interferon-stimulated gene 15 protein (ISG15), whereas SCoV-PLpro predominantly targets ubiquitin chains. The crystal structure of SCoV2-PLpro in complex with ISG15 reveals distinctive interactions with the amino-terminal ubiquitin-like domain of ISG15, highlighting the high affinity and specificity of these interactions. Furthermore, upon infection, SCoV2-PLpro contributes to the cleavage of ISG15 from interferon responsive factor 3 (IRF3) and attenuates type I interferon responses. Notably, inhibition of SCoV2-PLpro with GRL-0617 impairs the virus-induced cytopathogenic effect, maintains the antiviral interferon pathway and reduces viral replication in infected cells. These results highlight a potential dual therapeutic strategy in which targeting of SCoV2-PLpro can suppress SARS-CoV-2 infection and promote antiviral immunity. Show less
Data analysis workflows in many scientific domains have become increasingly complex and flexible. Here we assess the effect of this flexibility on the results of functional magnetic resonance... Show moreData analysis workflows in many scientific domains have become increasingly complex and flexible. Here we assess the effect of this flexibility on the results of functional magnetic resonance imaging by asking 70 independent teams to analyse the same dataset, testing the same 9 ex-ante hypotheses(1). The flexibility of analytical approaches is exemplified by the fact that no two teams chose identical workflows to analyse the data. This flexibility resulted in sizeable variation in the results of hypothesis tests, even for teams whose statistical maps were highly correlated at intermediate stages of the analysis pipeline. Variation in reported results was related to several aspects of analysis methodology. Notably, a meta-analytical approach that aggregated information across teams yielded a significant consensus in activated regions. Furthermore, prediction markets of researchers in the field revealed an overestimation of the likelihood of significant findings, even by researchers with direct knowledge of the dataset(2-5). Our findings show that analytical flexibility can have substantial effects on scientific conclusions, and identify factors that may be related to variability in the analysis of functional magnetic resonance imaging. The results emphasize the importance of validating and sharing complex analysis workflows, and demonstrate the need for performing and reporting multiple analyses of the same data. Potential approaches that could be used to mitigate issues related to analytical variability are discussed.The results obtained by seventy different teams analysing the same functional magnetic resonance imaging dataset show substantial variation, highlighting the influence of analytical choices and the importance of sharing workflows publicly and performing multiple analyses. Show less
Botvinik-Nezer, R.; Holzmeister, F.; Camerer, C.F.; Dreber, A.; Huber, J.; Johannesson, M.; ... ; Schonberg, T. 2020
Data analysis workflows in many scientific domains have become increasingly complex and flexible. Here we assess the effect of this flexibility on the results of functional magnetic resonance... Show moreData analysis workflows in many scientific domains have become increasingly complex and flexible. Here we assess the effect of this flexibility on the results of functional magnetic resonance imaging by asking 70 independent teams to analyse the same dataset, testing the same 9 ex-ante hypotheses(1). The flexibility of analytical approaches is exemplified by the fact that no two teams chose identical workflows to analyse the data. This flexibility resulted in sizeable variation in the results of hypothesis tests, even for teams whose statistical maps were highly correlated at intermediate stages of the analysis pipeline. Variation in reported results was related to several aspects of analysis methodology. Notably, a meta-analytical approach that aggregated information across teams yielded a significant consensus in activated regions. Furthermore, prediction markets of researchers in the field revealed an overestimation of the likelihood of significant findings, even by researchers with direct knowledge of the dataset(2-5). Our findings show that analytical flexibility can have substantial effects on scientific conclusions, and identify factors that may be related to variability in the analysis of functional magnetic resonance imaging. The results emphasize the importance of validating and sharing complex analysis workflows, and demonstrate the need for performing and reporting multiple analyses of the same data. Potential approaches that could be used to mitigate issues related to analytical variability are discussed.The results obtained by seventy different teams analysing the same functional magnetic resonance imaging dataset show substantial variation, highlighting the influence of analytical choices and the importance of sharing workflows publicly and performing multiple analyses. Show less
Multiomic profiling of several cohorts of patients treated with immune checkpoint blockade highlights the presence and potential role of B cells and tertiary lymphoid structures in promoting... Show moreMultiomic profiling of several cohorts of patients treated with immune checkpoint blockade highlights the presence and potential role of B cells and tertiary lymphoid structures in promoting therapy response.Treatment with immune checkpoint blockade (ICB) has revolutionized cancer therapy. Until now, predictive biomarkers(1-10) and strategies to augment clinical response have largely focused on the T cell compartment. However, other immune subsets may also contribute to anti-tumour immunity(11-15), although these have been less well-studied in ICB treatment(16). A previously conducted neoadjuvant ICB trial in patients with melanoma showed via targeted expression profiling(17) that B cell signatures were enriched in the tumours of patients who respond to treatment versus non-responding patients. To build on this, here we performed bulk RNA sequencing and found that B cell markers were the most differentially expressed genes in the tumours of responders versus non-responders. Our findings were corroborated using a computational method (MCP-counter(18)) to estimate the immune and stromal composition in this and two other ICB-treated cohorts (patients with melanoma and renal cell carcinoma). Histological evaluation highlighted the localization of B cells within tertiary lymphoid structures. We assessed the potential functional contributions of B cells via bulk and single-cell RNA sequencing, which demonstrate clonal expansion and unique functional states of B cells in responders. Mass cytometry showed that switched memory B cells were enriched in the tumours of responders. Together, these data provide insights into the potential role of B cells and tertiary lymphoid structures in the response to ICB treatment, with implications for the development of biomarkers and therapeutic targets. Show less
Velden, D.L. van der; Hoes, L.R.; Wijngaart, H. van der; Henegouwen, J.M.V.; Werkhoven, E. van; Roepman, P.; ... ; Voest, E.E. 2019
Cell-free DNA in the blood provides a non-invasive diagnostic avenue for patients with cancer'. However, characteristics of the origins and molecular features of cell-free DNA are poorly understood... Show moreCell-free DNA in the blood provides a non-invasive diagnostic avenue for patients with cancer'. However, characteristics of the origins and molecular features of cell-free DNA are poorly understood. Here we developed an approach to evaluate fragmentation patterns of cell-free DNA across the genome, and found that profiles of healthy individuals reflected nucleosomal patterns of white blood cells, whereas patients with cancer had altered fragmentation profiles. We used this method to analyse the fragmentation profiles of 236 patients with breast, colorectal, lung, ovarian, pancreatic, gastric or bile duct cancer and 245 healthy individuals. A machine learning model that incorporated genome-wide fragmentation features had sensitivities of detection ranging from 57% to more than 99% among the seven cancer types at 98% specificity, with an overall area under the curve value of 0.94. Fragmentation profiles could be used to identify the tissue of origin of the cancers to a limited number of sites in 75% of cases. Combining our approach with mutation-based cell-free DNA analyses detected 91% of patients with cancer. The results of these analyses highlight important properties of cell-free DNA and provide a proof-of-principle approach for the screening, early detection and monitoring of human cancer. Show less
Sikora, M.; Pitulko, V.V.; Sousa, V.C.; Allentoft, M.E.; Vinner, L.; Rasmussen, S.; ... ; Willerslev, E. 2019
Mergers of neutron stars are known to be associated with short gamma-ray bursts(1-4). If the neutron-star equation of state is sufficiently stiff (that is, the pressure increases sharply as the... Show moreMergers of neutron stars are known to be associated with short gamma-ray bursts(1-4). If the neutron-star equation of state is sufficiently stiff (that is, the pressure increases sharply as the density increases), at least some such mergers will leave behind a supramassive or even a stable neutron star that spins rapidly with a strong magnetic field(5-8) (that is, a magnetar). Such a magnetar signature may have been observed in the form of the X-ray plateau that follows up to half of observed short gamma-ray bursts(9,10). However, it has been expected that some X-ray transients powered by binary neutron-star mergers may not be associated with a short gamma-ray burst(11,12). A fast X-ray transient (CDF-S XT1) was recently found to be associated with a faint host galaxy, the redshift of which is unknown(13). Its X-ray and host-galaxy properties allow several possible explanations including a short gamma-ray burst seen off-axis, a low-luminosity gamma-ray burst at high redshift, or a tidal disruption event involving an intermediate-mass black hole and a white dwarf(13). Here we report a second X-ray transient, CDF-S XT2, that is associated with a galaxy at redshift z = 0.738 (ref.(14)). The measured light curve is fully consistent with the X-ray transient being powered by a millisecond magnetar. More intriguingly, CDF-S XT2 lies in the outskirts of its star-forming host galaxy with a moderate offset from the galaxy centre, as short gamma-ray bursts often do(15,16). The estimated event-rate density of similar X-ray transients, when corrected to the local value, is consistent with the event-rate density of binary neutron-star mergers that is robustly inferred from the detection of the gravitational-wave event GW170817. Show less
