bluuebunny/crossref_metadata_2025_split huggingface dataset:
Changes in the distribution of fitness effects and adaptive mutational spectra following a single first step towards adaptation
Authors: Aggeli Dimitra , Li Yuping , Sherlock Gavin 
Published: June 2020

AbstractThe fitness effects of random mutations are contingent upon the genetic and environmental contexts in which they occur, and this contributes to the unpredictability of evolutionary outcomes at the molecular level. Despite this unpredictability, the rate of adaptation in homogeneous environments tends to decrease over evolutionary time, due to diminishing returns epistasis, causing relative fitness gains to be predictable over the long term. Here, we studied the extent of diminishing returns epistasis and the changes in the adaptive mutational spectra after yeast populations have already taken their first adaptive mutational step. We used three distinct adaptive clones that arose under identical conditions from a common ancestor, from which they diverge by a single point mutation, to found populations that we further evolved. We followed the evolutionary dynamics of these populations by lineage tracking and determined adaptive outcomes using fitness assays and whole genome sequencing. We found compelling evidence for diminishing returns: fitness gains during the 2ndstep of adaptation are smaller than those of the 1ststep, due to a compressed distribution of fitness effects in the 2ndstep. We also found strong evidence for historical contingency at the genic level: the beneficial mutational spectra of the 2nd-step adapted genotypes differ with respect to their ancestor and to each other, despite the fact that the three founders’ 1st-step mutations provided their fitness gains due to similar phenotypic improvements. While some targets of selection in the second step are shared with those seen in the common ancestor, other targets appear to be contingent on the specific first step mutation, with more phenotypically similar founding clones having more similar adaptive mutational spectra. Finally, we found that disruptive mutations, such as nonsense and frameshift, were much more common in the first step of adaptation, contributing an additional way that both diminishing returns and historical contingency are evident during 2ndstep adaptation.
Single-cell transcriptomic analysis of SARS-CoV-2 reactive CD4+T cells
Authors: Meckiff Benjamin J. , Ramírez-Suástegui Ciro , Fajardo Vicente , Chee Serena J , Kusnadi Anthony , Simon Hayley , Grifoni Alba , Pelosi Emanuela , Weiskopf Daniela , Sette Alessandro , Ay Ferhat , Seumois Grégory , Ottensmeier Christian H , Vijayanand Pandurangan 
Published: June 2020

ABSTRACTThe contribution of CD4+T cells to protective or pathogenic immune responses to SARS-CoV-2 infection remains unknown. Here, we present large-scale single-cell transcriptomic analysis of viral antigen-reactive CD4+T cells from 32 COVID-19 patients. In patients with severe disease compared to mild disease, we found increased proportions of cytotoxic follicular helper (TFH) cells and cytotoxic T helper cells (CD4-CTLs) responding to SARS-CoV-2, and reduced proportion of SARS-CoV-2 reactive regulatory T cells. Importantly, the CD4-CTLs were highly enriched for the expression of transcripts encoding chemokines that are involved in the recruitment of myeloid cells and dendritic cells to the sites of viral infection. Polyfunctional T helper (TH)1 cells and TH17 cell subsets were underrepresented in the repertoire of SARS-CoV-2-reactive CD4+T cells compared to influenza-reactive CD4+T cells. Together, our analyses provide so far unprecedented insights into the gene expression patterns of SARS-CoV-2 reactive CD4+T cells in distinct disease severities.
Single-cell profiling of Ebola virus infectionin vivoreveals viral and host transcriptional dynamics
Authors: Kotliar Dylan , Lin Aaron E. , Logue James , Hughes Travis K. , Khoury Nadine M. , Raju Siddharth S. , Wadsworth Marc H. , Chen Han , Kurtz Jonathan R. , Dighero-Kemp Bonnie , Bjornson Zach B. , Mukherjee Nilanjan , Sellers Brian A. , Tran Nancy , Bauer Matthew R. , Adams Gordon C. , Adams Ricky , Rinn John L. , Melé Marta , Nolan Garry P. , Barnes Kayla G. , Hensley Lisa E. , McIlwain David R. , Shalek Alex K. , Sabeti Pardis C. , Bennett Richard S. 
Published: June 2020

SummaryEbola virus (EBOV) causes epidemics with high case fatality rates, yet remains understudied due to the challenge of experimentation in high-containment and outbreak settings. To better understand EBOV infectionin vivo, we used single-cell transcriptomics and CyTOF-based single-cell protein quantification to characterize peripheral immune cell activity during EBOV infection in rhesus monkeys. We obtained 100,000 transcriptomes and 15,000,000 protein profiles, providing insight into pathogenesis. We find that immature, proliferative monocyte-lineage cells with reduced antigen presentation capacity replace conventional circulating monocyte subsets within days of infection, while lymphocytes upregulate apoptosis genes and decline in abundance. By quantifying viral RNA abundance in individual cells, we identify molecular determinants of tropism and examine temporal dynamics in viral and host gene expression. Within infected cells, we observe that EBOV down-regulatesSTAT1mRNA and interferon signaling, and up-regulates putative pro-viral genes (e.g.,DYNLL1andHSPA5), nominating cellular pathways the virus manipulates for its replication. Overall, this study sheds light on EBOV tropism, replication dynamics, and elicited immune response, and provides a framework for characterizing interactions between hosts and emerging viruses in a maximum containment setting.
An electron transfer competent structural ensemble of membrane-bound cytochrome P450 1A1 and cytochrome P450 oxidoreductase
Authors: Mukherjee Goutam , Nandekar Prajwal P. , Wade Rebecca C. 
Published: June 2020

AbstractCytochrome P450 (CYP) heme monooxygenases require two electrons for their catalytic cycle. For mammalian microsomal CYPs, key enzymes for xenobiotic metabolism and steroidogenesis and important drug targets and biocatalysts, the electrons are transferred by NADPH-cytochrome P450 oxidoreductase (CPR). No structure of a mammalian CYP-CPR complex has been solved experimentally, hindering understanding of the determinants of electron transfer (ET), which is often rate-limiting for CYP reactions. Here, we investigated the interactions between membrane-bound CYP 1A1, an antitumor drug target, and CPR by a multiresolution computational approach. We find that upon binding to CPR, the CYP 1A1 catalytic domain becomes less embedded in the membrane and reorients, indicating that CPR may affect ligand passage to the CYP active site. Despite the constraints imposed by membrane binding, we identify several arrangements of CPR around CYP 1A1 that are compatible with ET. In the complexes, the interactions of the CPR FMN domain with the proximal side of CYP 1A1 are supplemented by more transient interactions of the CPR NADP domain with the distal side of CYP 1A1. Computed ET rates and pathways agree well with available experimental data and suggest why the CYP-CPR ET rates are low compared to those of soluble bacterial CYPs.
Crystal structure of inhibitor-bound human MSPL that can activate high pathogenic avian influenza
Authors: Ohno Ayako , Maita Nobuo , Tabata Takanori , Nagano Hikaru , Arita Kyohei , Ariyoshi Mariko , Uchida Takayuki , Nakao Reiko , Ulla Anayt , Sugiura Kosuke , Kishimoto Koji , Teshima-Kondo Shigetada , Okumura Yuushi , Nikawa Takeshi 
Published: June 2020

AbstractInfection of certain influenza viruses is triggered when its hemagglutinin (HA) is cleaved by host cell proteases such as proprotein convertases and type II transmembrane serine proteases (TTSP). HA with a monobasic motif is cleaved by trypsin-like proteases, including TMPRSS2 and HAT, while the multibasic motif found in high pathogenicity avian influenza HA is cleaved by furin, PC5/6, or MSPL. MSPL belongs to the TMPRSS family and preferentially cleaves [R/K]-K-K-R↓ sequences. Here, we solved the crystal structure of the extracellular region of human MSPL in complex with an irreversible substrate-analog inhibitor. The structure revealed three domains clustered around the C-terminal α-helix of the SPD. The inhibitor structure and its putative model show that the P1-Arg inserts into the S1 pocket, whereas the P2-Lys and P4-Arg interacts with the Asp/Glu-rich 99-loop that is unique to MSPL. Based on the structure of MSPL, we also constructed a homology model of TMPRSS2, which is essential for the activation of the SARS-CoV-2 spike protein and infection. The model may provide the structural insight for the drug development for COVID-19.
Autoregulatory control of microtubule binding in the oncogene, doublecortin-like kinase 1
Authors: Rogers Melissa M. , Ramkumar Amrita , Downing Ashlyn M. , Bodin Hannah , Castro Julia , Nowakowski Dan W. , Ori-McKenney Kassandra M. 
Published: June 2020

AbstractThe microtubule-associated protein (MAP), doublecortin-like kinase 1 (DCLK1), is highly expressed in a range of cancers and is a prominent therapeutic target for the development of kinase inhibitors. However, the physiological roles of its kinase activity and how DCLK1 kinase activity is regulated remain elusive. Here we employ in vitro reconstitution with purified proteins to analyze the role of DCLK1 kinase activity in regulating microtubule binding. We find that DCLK1 autophosphorylates a single residue within its C-terminal tail to restrict its kinase activity and prevent aberrant hyperphosphorylation within its microtubule-binding domain. Removal of the C-terminal tail or mutation of this residue causes an increase in phosphorylation largely within the doublecortin 2 (DC2) domain, which dramatically reduces the microtubule affinity of DCLK1. Therefore, autophosphorylation at specific sites within DCLK1 have diametric effects on the molecule’s ability to associate with microtubules. Overall, our results suggest a mechanism by which DCLK1 modulates its own kinase activity to tune its microtubule binding affinity, providing molecular insights into a unique form of autoregulatory control over microtubule binding activity within the broader family of MAPs. These results provide useful molecular insights for future therapeutic efforts related to DCLK1’s role in cancer development and progression.
The impact of interneurons on nonlinear synaptic integration in the neocortex
Authors: Dorsett Christopher , Philpot Benjamin , Smith Spencer LaVere , Smith Ikuko T. 
Published: June 2020

AbstractExcitatory inputs arriving at the dendrites of a neuron can engage active mechanisms that nonlinearly amplify the depolarizing currents. Interneuron-mediated inhibition can modulate this active process, in a subtype-dependent manner. For example, dendrite-targeting inhibition is hypothesized to increase the amplitude of synaptic input required to activate voltage-dependent nonlinear synaptic integration. To examine how inhibition influences active synaptic integration, we optogenetically manipulated the activity of two different subtypes of interneurons: dendrite-targeting somatostatin-expressing (SOM) and perisomatic-targeting parvalbumin-expressing (PV) interneurons. In acute slices of mouse primary visual cortex, electrical stimulation evoked nonlinear synaptic integration that depended onN-methyl-D-aspartate (NMDA) receptors. Optogenetic activation of SOM neurons in conjunction with electrical stimulation resulted in predominantly divisive inhibitory gain control, reducing the magnitude of the nonlinear response without affecting its threshold. PV activation, on the other hand, had a minimal effect on dendritic nonlinearities, resulting in a small subtractive effect. Furthermore, we found that mutual inhibition among SOM interneurons was strong and more prevalent than previously thought, while mutual inhibition among PV interneurons was minimal. These results challenge previous models of inhibitory modulation of active synaptic integration. The major effect of SOM inhibition is not a shift in threshold for activation of nonlinear integration, but rather a decrease the amplitude of the nonlinear response.
Ultra-rapid metagenotyping of the human gut microbiome
Authors: Shi Zhou Jason , Dimitrov Boris , Zhao Chunyu , Nayfach Stephen , Pollard Katherine S. 
Published: June 2020

AbstractSequence variation is used to quantify population structure and identify genetic determinants of phenotypes that vary within species. In the human microbiome and other environments, single nucleotide polymorphisms (SNPs) are frequently detected by aligning metagenomic sequencing reads to catalogs of genes or genomes. But this requires high-performance computing and enough read coverage to distinguish SNPs from sequencing errors. We solved these problems by developing the GenoTyper for Prokaytotes (GT-Pro), a suite of novel methods to catalog SNPs from genomes and use exact k-mer matches to perform ultra-fast reference-based SNP calling from metagenomes. Compared to read alignment, GT-Pro is more accurate and two orders of magnitude faster. We discovered 104 million SNPs in 909 human gut species, characterized their global population structure, and tracked pathogenic strains. GT-Pro democratizes strain-level microbiome analysis by making it possible to genotype hundreds of metagenomes on a personal computer.Software availabilityGT-Pro is available athttps://github.com/zjshi/gt-pro.
WHOLE-GENOME SEQUENCING ANDDE NOVOASSEMBLY OF A 2019 NOVEL CORONAVIRUS (SARS-COV-2) STRAIN ISOLATED IN VIETNAM
Authors: Lam Le Tung , Hieu Nguyen Trung , Trang Nguyen Hong , Thuong Ho Thi , Linh Tran Huyen , Tien Luu Thuy , Thao Nguyen Thi Ngoc , Loan Huynh Thi Kim , Quang Pham Duy , Quang Luong Chan , Thang Cao Minh , Thuong Nguyen Vu , Ha Hoang , Ha Chu Hoang , Lan Phan Trong , Hai Truong Nam 
Published: June 2020

ABSTRACTThe pandemic COVID-19 caused by the zoonotic virus SARS-CoV-2 has devastated countries worldwide, infecting more than 4.5 million people and leading to more than 300,000 deaths. Whole genome sequencing (WGS) is an effective tool to monitor emerging strains and provide information for intervention, thus help to inform outbreak control decisions. Here, we reported the first effort to sequence andde novoassemble the whole genome of SARS-CoV-2 using PacBio’s SMRT sequencing technology in Vietnam. We also presented the annotation results and a brief analysis of the variants found in our SARS-CoV-2 strain, which was isolated from a Vietnamese patient. The sequencing was successfully completed andde novoassembled in less than 30 hours, resulting in one contig with no gap and a length of 29,766 bp. All detected variants as compared to the NCBI reference were highly accurate as confirmed by Sanger sequencing. The results have shown the potential of long read sequencing to provide high quality WGS data to support public health responses, and advance understanding of this and future pandemics.
Cellular and Cytokine Responses in the Granulomas of Asymptomatic Cattle naturally infected withMycobacterium bovisin Ethiopia
Authors: Tulu Begna , Martineau Henny M , Zewude Aboma , Desta Fekadu , Jolliffe David A , Abebe Markos , Tolera Taye , Belay Mulugeta , Martineau Adrian R , Ameni Gobena 
Published: June 2020

ABSTRACTCellular (CD3+ T cell and CD68+ macrophages), cytokines (IFN-γ+ and TNF-α+) and effector molecule (iNOS+) responses were evaluated in the lymph nodes and tissue of cattle naturally infected withMycobacterium bovis. Detailed post mortem and immunonohistochemical examinations of lesions were performed on 16 cows positive for single intradermal cervical comparative tuberculin test (SICCTT) and identified from dairy farms located around the Addis Ababa City. The severity of the gross lesion was significantly higher (p=0.003) inM. bovisculture positive (n=12) cows than in culture negative (n=4). Immunohistochemical techniques showed that the mean percentage labelling of CD3+ T cells decreased as the stage of granuloma increased from stage I to stage IV in culture positive cows (p<0.001). On the other hand, the proportional fraction of CD68+ macrophages and the concentrations of IFN-γ+, TNF-α+ and iNOS+ increased significantly from stage I to stage IV (p< 0.001) in culture positive cows. At the early stage of the granuloma, the culture negative cows showed significantly higher mean proportions of CD68+ macrophages (p=0.03) as well as the concentrations of iNOS+ (p=0.007) compared to culture positive cows. Similarly, at advanced granuloma stages, culture negative cows demonstrated significantly higher mean proportions of CD3+ T cells (p< 0.001) compared to culture positive cows. Thus, the present study demonstrated that following natural infection of cows withM. bovis, as the stage of granuloma increases from stage I to stage IV, the proportion of CD3+ cells decreases while the immunolabeling fraction of CD68+ macrophages, IFN-γ+, TNF-α and iNOS+ increases.
Group A Streptococcus Infection of the Nasopharynx Requires Proinflammatory Signaling Through the Interleukin-1 Receptor
Authors: LaRock Doris L. , Russell Raedeen , Johnson Anders F. , Wilde Shyra , LaRock Christopher N. 
Published: June 2020

ABSTRACTGroup AStreptococcus(GAS) is the etiologic agent of numerous high morbidity and high mortality diseases which commonly have a highly proinflammatory pathology. One factor contributing to this inflammation is the GAS protease SpeB, which directly activates the proinflammatory cytokine interleukin-1β (IL-1β), independent of the canonical inflammasome pathway. IL-1β drives neutrophil activation and recruitment that limits bacterial growth and invasion during invasive skin and soft tissue infections like necrotizing fasciitis. GAS also causes pharyngitis (strep throat), and the upper respiratory tract is its primary nidus for growth and transmission. Since the fitness selection for the species is likely primarily for this site, we examined the process of IL-1β activation in the murine nasopharynx. SpeB still activated IL-1β, which was required for neutrophil migration, but this inflammation instead increased GAS replication. Inhibiting IL-1β or depleting neutrophils, which both promote invasive infection, prevented GAS infection of the nasopharynx. Prior antibiotic exposure increased GAS growth in the murine nasopharynx, and antibiotics were sufficient to reverse the attenuation previously observed when IL-1β, neutrophils, or SpeB were not present to drive inflammation. Therefore, the same fundamental mechanism has opposing effects on virulence at different body sites. Invasive disease may be limited in part due to specific adaptations for inducing host inflammation that are beneficial for pharyngitis.IMPORTANCEOur previous reports showed that Group AStreptococcus(GAS) protease SpeB directly activates the host proinflammatory cytokine IL-1β and this restricts invasive skin infection. The upper respiratory tract is the primary site of GAS colonization and infection, but the host-pathogen interactions at this site are still largely unknown. We provide the first evidence that IL-1β-mediated inflammation promotes upper respiratory tract infection. This provides experimental evidence that the notable inflammation of strep throat, which presents with significant swelling, pain, and neutrophil influx, is not an ineffectual immune response, but rather is a GAS-directed remodeling of this niche for its pathogenic benefit.
Structure of the cysteine-rich domain ofPlasmodium falciparumP113 identifies the location of the RH5 binding site
Authors: Campeotto Ivan , Galaway Francis , Mehmood Shahid , Barfod Lea K. , Quinkert Doris , Kotraiah Vinayaka , Phares Timothy W. , Wright Katherine E. , Snijders Ambrosius P. , Draper Simon J. , Higgins Matthew. K. , Wright Gavin J. 
Published: June 2020

AbstractPlasmodium falciparumRH5 is a secreted parasite ligand that is essential for erythrocyte invasion through direct interaction with the host erythrocyte receptor basigin. RH5 forms a tripartite complex with two other secreted parasite proteins: CyRPA and RIPR, and is tethered to the surface of the parasite through membrane-anchored P113. Antibodies against RH5, CyRPA and RIPR inhibit parasite invasion, suggesting that vaccines containing these three components have the potential to prevent blood-stage malaria. To further explore the role of the P113-RH5 interaction, we selected monoclonal antibodies against P113 that were either inhibitory or non-inhibitory for RH5 binding. Using a Fab fragment as a crystallisation chaperone, we determined the crystal structure of the RH5-binding region of P113 and showed that it is composed of two domains with structural similarities to rhamnose-binding lectins. We identified the RH5 binding site on P113 by using a combination of hydrogen-deuterium exchange mass spectrometry and site directed mutagenesis. We found that a monoclonal antibody to P113 that bound to this interface and inhibited the RH5-P113 interaction did not inhibit parasite blood-stage growth. These findings provide further structural information on the protein interactions of RH5 and will be helpful in the development of blood-stage malaria vaccines that target RH5.ImportanceMalaria is a deadly infectious disease primarily caused by the parasitePlasmodium falciparum.It remains a major global health problem and there is no highly effective vaccine. A parasite protein called RH5 is centrally involved in the invasion of host red blood cells, making it – and the other parasite proteins it interacts with – promising vaccine targets. We recently identified a protein called P113 that binds RH5 suggesting that it anchors RH5 to the parasite surface. In this paper, we use structural biology to locate and characterize the RH5 binding region on P113. These findings will be important to guide the development of new anti-malarial vaccines to ultimately prevent this disease which affects some of the poorest people on the planet.
Covariation of Zinc Deficiency with COVID-19 Infections and Mortality in European Countries: Is Zinc Deficiency a Risk Factor for COVID-19?
Authors: Singh Samer 
Published: June 2020

AbstractVariables responsible for the differential COVID-19 pandemic severity among countries remain undefined. Zinc, a micronutrient required for immunocompetence, is found deficient in populations. We hypothesized the differential COVID-19 severity observed among European countries could be associated with the Zn-deficiency prevalence. The COVID-19 data from different stages of pandemic, i.e., 8 April, 12 and 26 May 2020, were analyzed for covariation with the estimated Zn-deficiency. A significant, relatively stable, but negative correlation of Zn-deficiency with cases per million for the time period [r(20): -0.4930 to -0.5335, p-value: 0.02720 to 0.0154] and a steady improvement of covariation with deaths per million [r(20): -0.4056; p-value: 0.0760 on 26 May 2020] was observed. Considering, Zinc’s key immunomodulatory role, widespread deficiency along with the self- and prescribed intervention in different target groups, e.g. children, women, elderly, carefully planned dedicated exploratory studies to understand the basis of the observed association are advisable.