bluuebunny/crossref_metadata_2025 huggingface dataset:
Distinctive functional regime of endogenous lncRNAs in dark regions of human genome
Authors: Wang Anyou , Hai Rong
Published: December 2020

Eukaryotic genomes gradually gain noncoding regions when advancing evolution and human genome actively transcribes >90% of its noncoding regions1, suggesting their criticality in evolutionary human genome. Yet <1% of them have been functionally characterized2, leaving most human genome in dark. Here we systematically decode endogenous lncRNAs located in unannotated regions of human genome and decipher a distinctive functional regime of lncRNAs hidden in massive RNAseq data. LncRNAs divergently distribute across chromosomes, independent of protein-coding regions. Their transcriptions barely initiate on promoters through polymerase II, but mostly on enhancers. Yet conventional enhancer activators(e.g. H3K4me1) only account for a small proportion of lncRNA activation, suggesting alternatively unknown mechanisms initiating the majority of lncRNAs. Meanwhile, lncRNA-self regulation also notably contributes to lncRNA activation. LncRNAs trans-regulate broad bioprocesses, including transcription and RNA processing, cell cycle, respiration, response to stress, chromatin organization, post-translational modification, and development. Overall lncRNAs govern their owned regime distinctive from protein’s.
Cannabinoids modulate food preference and consumption inDrosophila melanogaster
Authors: He Jianzheng , Xien Tan Alice Mei , Ng Si Yun , Yu Fengwei
Published: December 2020

Cannabinoids have an important role in regulating feeding behaviors via cannabinoid receptors in mammals. Cannabinoids also exhibit potential therapeutic functions inDrosophila melanogaster, or fruit fly that lacks cannabinoid receptors. However, it remains unclear whether cannabinoids affect food consumption and metabolism in a cannabinoid receptors-independent manner in flies. In this study, we systematically investigated pharmacological functions of various cannabinoids in modulating food preference and consumption in flies. We show that flies display preferences for consuming cannabinoids, independent of their sensory functions. Interestingly, phyto- and endo- cannabinoids exhibit an inhibitory effect on food intake. Unexpectedly, the non-selective CB1 receptor antagonist AM251 attenuates the suppression of food intake by endocannabinoids. Moreover, the endocannabinoid anandamide (AEA) and its metabolite inhibit food intake and promote resistance to starvation, possibly through reduced lipid metabolism. Thus, this study has provided insights into a pharmacological role of cannabinoids in feeding behaviors using an adultDrosophilamodel.
Mechanistic basis for ubiquitin modulation of a protein energy landscape
Authors: Carroll Emma C. , Latorraca Naomi R. , Lindner Johanna M. , Maguire Brendan C. , Pelton Jeff G. , Marqusee Susan
Published: December 2020

Ubiquitin is a common posttranslational modification canonically associated with targeting proteins to the 26S proteasome for degradation and also plays a role in numerous other non-degradative cellular processes. Ubiquitination at certain sites destabilizes the substrate protein, with consequences for proteasomal processing, while ubiquitination at other sites has little energetic effect. How this site specificity—and, by extension, the myriad effects of ubiquitination on substrate proteins—arises remains unknown. Here, we systematically characterize the atomic-level effects of ubiquitination at various sites on a model protein, barstar, using a combination of NMR, hydrogen-deuterium exchange mass spectrometry, and molecular dynamics simulation. We find that, regardless of the site of modification, ubiquitination does not induce large structural rearrangements in the substrate. Destabilizing modifications, however, increase fluctuations from the native state resulting in exposure of the substrate’s C terminus. Both of the sites occur in regions of barstar with relatively high conformational flexibility. Destabilization, however, appears to occur through different thermodynamic mechanisms, involving a reduction in entropy in one case and a loss in enthalpy in another. By contrast, ubiquitination at a non-destabilizing site protects the substrate C terminus through intermittent formation of a structural motif with the last three residues of ubiquitin. Thus, the biophysical effects of ubiquitination at a given site depend greatly on local context. Taken together, our results reveal how a single post-translational modification can generate a broad array of distinct effects, providing a framework to guide the design of proteins and therapeutics with desired degradation and quality-control properties. (248 words)

Significance Statement

Fluctuations on a protein energy landscapes encode the mechanistic basis for vital biological processes not always evident from static structures alone. Ubiquitination, a key posttranslational modification, can affect a protein’s energy landscape with consequences for proteasomal degradation, but the molecular mechanisms driving ubiquitin-induced energetic changes remain elusive. Here, we systematically characterize the energetic effects of ubiquitination at three sites on a model protein. We find that distinct thermodynamic mechanisms can produce the same outcome of ubiquitin-induced destabilization at sensitive sites. At a non-sensitive site, we observe formation of a substrate–ubiquitin interaction that may serve to protect against destabilization. This work will enable development of predictive models of the effect of ubiquitin at any given site on a protein with implications for understanding and engineering regulated ubiquitin signaling and protein quality control in vivo.
Basis for the phototaxis sign reversal in the green algaChlamydomonas reinhardtiistudied by high-speed observation
Authors: Nakajima Masako , Iizuka Kosuke , Takahashi Rina , Ueki Noriko , Isu Atsuko , Yoshimura Kenjiro , Nakagaki Toshiyuki , Hisabori Toru , Sato Katsuhiko , Wakabayashi Ken-ichi
Published: December 2020

For organisms that respond to environmental stimuli using taxes, reversal of the tactic sign should be tightly regulated for survival. The biciliate green algaChlamydomonas reinhardtiiis an excellent model for studying reversal between positive and negative phototaxis.C. reinhardtiicells change swimming direction by modulating the balance of beating forces between their two cilia after photoreception at the eyespot; however, it remains unknown how they reverse phototactic sign. In this study, we observed cells undergoing phototactic turns with a high-speed camera and found that two key factors determine the phototactic sign: which of the two cilia beats more strongly for phototactic turning and when the strong beating starts. The timing of the strong ciliary beating is suggested to be regulated by ROS-regulated switching between the light-on and light-off responses at the eyespot, which leads to the switching between positive and negative phototaxis. This idea is supported by a mathematical model that introduces the timing of the strong ciliary beating after photoreception.
Ketamine Modulates the Neural Correlates of Reward Processing in Unmedicated Patients in Remission from Depression
Authors: Kotoula Vasileia , Stringaris Argyris , Mackes Nuria , Mazibuko Ndabezinhle , Hawkins Peter.C.T , Furey Maura , Curran H Valerie , Mehta Mitul.A.
Published: December 2020

Ketamine as an antidepressant improves anhedonia, a pernicious symptom of depression as early as 2h post-infusion. The effects of ketamine on anhedonia are thought to be exerted via actions on reward-related brain areas—yet, these actions remain largely unknown. This study examines ketamine’s effects during the anticipation and receipt of an expected reward, after the psychotomimetic effects of ketamine have passed, when early antidepressant effects are reported. In order to identify brain areas that are modulated by the drug per se and are not linked to symptom changes, we have recruited 37 participants who remitted from depression and were free from symptoms and antidepressant treatments at the time of the scan. Participants were scanned while performing a monetary reward task and we examined ketamine’s effects on pre-defined brain areas that are part of the reward circuit. An overall effect of ketamine was observed during the anticipation and feedback phases of win and no-win trials. The drug effects were particularly prominent in the nucleus accumbens and putamen, upon the receipt of smaller rewards and the levels of (2R,6R)-HNK, 2h post-infusion, significantly correlated with the activation observed in the ventral tegmental area (VTA) for that contrast. These findings demonstrate that ketamine can produce detectable changes in reward-related brain areas, 2h after infusion, which occur without symptom changes and support the idea that ketamine might improve reward-related symptoms via modulation of response to feedback.
Distinct impact of IgG subclass and Fc-FcγR interaction on autoantibody pathogenicity in different IgG4-mediated diseases
Authors: Bi Yanxia , Su Jian , Zhou Shengru , Zhao Yingjie , Zhang Yan , Zhang Huihui , Liu Mingdong , Zhou Aiwu , Pan Meng , Zhao Yiming , Li Fubin
Published: December 2020

IgG4 is the least potent human IgG subclass for the FcγR-mediated antibody effector function and is considered anti-inflammatory in the context of prolonged inflammation and allergic responses. Paradoxically, IgG4 is also the dominant IgG subclass of pathogenic autoantibodies in IgG4-mediated diseases. Here we show that the IgG subclass and Fc-FcγR interaction have a distinct impact on the pathogenic function of autoantibodies in different IgG4-mediated diseases. While IgG4 and its weak Fc-FcγR interaction have an ameliorative role in the pathogenicity of anti-ADAMTS13 autoantibodies isolated from thrombotic thrombocytopenic purpura (TTP) patients, they have an unexpected exacerbating effect on anti-Dsg1 autoantibody pathogenicity in pemphigus foliaceus (PF) models. Strikingly, a non-pathogenic anti-Dsg1 antibody variant optimized for FcγR-mediated effector function can attenuate the skin lesions induced by pathogenic anti-Dsg1 antibodies by promoting the clearance of dead keratinocytes. These studies suggest that IgG effector function contributes to the clearance of autoantibody-Ag complexes, which is harmful in TTP, but beneficial in PF and may provide new therapeutic opportunity.
Investigating the effects of accelerated partner therapy (APT) on chlamydia transmission in Britain: a mathematical modelling study
Authors: Althaus Christian L , Mercer Catherine H , Cassell Jackie A , Estcourt Claudia S , Low Nicola
Published: December 2020

Understanding the effects of partner notification (PN) on the transmission of chlamydia, the most prevalent bacterial sexual transmitted infection worldwide, is critical for implementing optimal control strategies. Accelerated partner therapy (APT) aims to increase the numbers of partners treated and reduce the time to partner treatment. Our objective was to study the effects of APT interventions on partner treatment and chlamydia transmission in order to better understand the results of LUSTRUM, an APT cross-over cluster randomised controlled trial in the UK. We developed a novel deterministic, population-based chlamydia transmission model including the process of PN. We considered a population aged 16-34 years and calibrated the model to sexual behaviour data between people of the opposite-sex and chlamydia prevalence data reported by 3,671 participants in Britain's third National Survey of Sexual Attitudes and Lifestyles (Natsal-3, 2010-2012) using Approximate Bayesian Computation (ABC). We investigated the potential effects of APT on chlamydia transmission by increasing the number of treated partners and reducing the time to partner treatment compared to standard PN. The median prevalence of chlamydia in the model was 1.84% (95% credible interval, CrI: 1.60%-2.62%) in women and 1.78% (95% CrI: 1.13%-2.14%) in men. Chlamydia positivity was highest in partners of symptomatic index cases with low sexual activity. Infected partners were typically asymptomatic and belonged to the high sexual activity group, i.e., are naturally those infected individuals that will contribute most to onward transmission. Reducing the time to partner treatment without achieving higher numbers of partners treated had only minor effects on reducing chlamydia prevalence. In contrast, the model predicts that a potential increase in the number of partners treated from current levels in Britain (0.51, 95% CrI: 0.21-0.80) by 25% would reduce chlamydia prevalence by 18% (95% CrI: 5%-44%) in both women in men within 5 years. These results suggest that APT, through a potential increase in the proportion of partners treated, would be an effective method to reduce ongoing chlamydia transmission in Britain.
A generic, scalable, and rapid TR-FRET –based assay for SARS-CoV-2 antigen detection
Authors: Rusanen Juuso , Kareinen Lauri , Szirovicza Leonora , Uğurlu Hasan , Levanov Lev , Jääskeläinen Anu , Ahava Maarit , Kurkela Satu , Saksela Kalle , Hedman Klaus , Vapalahti Olli , Hepojoki Jussi
Published: December 2020

The ongoing COVID-19 pandemic has seen an unprecedented increase in the demand for rapid and reliable diagnostic tools, leaving many laboratories scrambling for resources. We present a fast and simple method for the detection of SARS-CoV-2 in nasopharyngeal swabs. The method is based on the detection of SARS-CoV-2 nucleoprotein (NP) and S protein (SP) via time-resolved Förster resonance energy transfer (TR-FRET) with donor- and acceptor-labeled polyclonal anti-NP and -SP antibodies. Using recombinant proteins and cell culture-grown SARS-CoV-2 the limits of detection were established as 25 pg of NP or 20 infectious viral units (i.u.), and 875 pg of SP or 625 i.u. of SARS-CoV-2. Testing RT-PCR positive (n=48, with cycle threshold [Ct] values from 11 to 30) or negative (n=96) nasopharyngeal swabs, we showed that the assay yields positive results for all samples with Ct values of <25 and a single RT-PCR negative sample. We determined the presence of infectious virus in the RT-PCR-positive nasopharyngeal swabs by virus isolation, and observed a strong association between the presence of infectious virus and a positive antigen test result. The NP-based assay showed 97.4% (37/38) sensitivity and 100% (10/10) specificity in comparison with virus isolation, and 77.1% (37/48) and 99.0% (95/96) in comparison with SARS-CoV-2 RT-PCR. The assay is performed in a buffer that neutralizes SARS-CoV-2 infectivity and is relatively simple to set up as an “in-house” test. The assay principle as such is applicable to other viral infections, and could also be readily adapted to a massively high throughput testing format.
Integrative analyses identify susceptibility genes underlying COVID-19 hospitalization
Authors: Pathak Gita A , Singh Kritika , Miller-Fleming Tyne W , Wendt Frank , Ehsan Nava , Hou Kangcheng , Johnson Ruth , Lu Zeyun , Gopalan Shyamalika , Dimbou Loic Yengo , Mohammadi Pejman , Pasaniuc Bogdan , Polimanti Renato , Davis Lea K , Mancuso Nicholas
Published: December 2020

Despite rapid progress in characterizing the role of host genetics in SARS-Cov-2 infection, there is limited understanding of genes and pathways that contribute to COVID-19. Here, we integrated a genome-wide association study of COVID-19 hospitalization (7,885 cases and 961,804 controls from COVID-19 Host Genetics Initiative) with mRNA expression, splicing, and protein levels (n=18,502). We identified 27 genes related to inflammation and coagulation pathways whose genetically predicted expression was associated with COVID-19 hospitalization. We functionally characterized the 27 genes using phenome- and laboratory-wide association scans in Vanderbilt Biobank (BioVU; n=85,460) and identified coagulation-related clinical symptoms, immunologic, and blood-cell-related biomarkers. We replicated these findings across trans-ethnic studies and observed consistent effects in individuals of diverse ancestral backgrounds in BioVU, pan-UK Biobank, and Biobank Japan. Our study highlights putative causal genes impacting COVID-19 severity and symptomology through the host inflammatory response.
Test-retest reliability of hepatic venous pressure gradient and impact on trial design: a study in 289 patients from the control groups of 20 randomized trials
Authors: Bai Wayne , Al-Kharagouli Mustafa , Stach Jesse , Matheson Granville J , Abraldes Juan G
Published: December 2020

Abstract
Background and Aims: Portal hypertension (PH) is a major driver for cirrhosis complications. Portal pressure is estimated in practice by the hepatic venous pressure gradient (HVPG). The assessment of HVPG changes has been used for drug development in PH. This study aimed at quantifying the test-retest reliability and consistency of HVPG in the specific context of RCTs for the treatment of PH in cirrhosis and its impact on power calculations for trial design.
Method: We conducted a search of published RCTs in patients with cirrhosis reporting individual patient-level data of HVPG at baseline and after an intervention, and that included a placebo or untreated control arm. Baseline and follow-up HVPG in the control groups were extracted after digitizing the plots. We assessed different reliability parameters and the potential impact of study characteristics.
Results: We retrieved a total of 289 before-after HVPG measurements in the placebo/untreated groups from 20 RCTs. Time range between the two HVPGs measurements was 20 min to 730 days. Test-retest reliability was higher in studies including only compensated patients and, therefore, modelled sample size calculations for trials in compensated cirrhosis were lower than for decompensated cirrhosis. Higher proportion of alcohol-related cirrhosis and unicentric trials were associated with lower differences between baseline and follow-up measurements. Smallest detectable difference in an individual was 24% and 32% in compensated and decompensated patients respectively
Conclusion: The test-retest reliability of HVPG is overall excellent, but higher in studies limited to compensated cirrhosis. These findings should be taking into account when powering trials based in the effects on HVPG or when consider HVPG as a tool to guide therapy of portal hypertension
Excess deaths associated with the Iranian COVID-19 epidemic: a province-level analysis
Authors: Ghafari Mahan , Kadivar Alireza , Katzourakis Aris
Published: December 2020

Background

The number of publicly reported deaths from COVID-19 may underestimate the true death toll from the epidemic as they rely on provisional data that are often incomplete or omit undocumented deaths from COVID-19. In addition, these reports may be subject to significant under-reporting due to a limited testing capacity of a country to identify suspect cases. This study estimated the number of seasonal excess deaths attributable to the COVID-19 epidemic in 31 provinces of Iran.

Methods

We gathered the nationwide and provincial time series of the seasonal all-cause mortality data from spring 2015 to summer 2020 (21 March 2015 to 21 September 2020), in accordance with the Solar Hijri (SH) calendar, from the National Organization for Civil Registration (NOCR). We estimated the expected number of seasonal deaths for each province using a piecewise linear regression model which we established based on the mortality figures for the previous years and considered any significant deviations from the expectation during winter, spring, and summer of 2020 to be directly associated with COVID-19.

Results

Our analysis shows that from the start of winter to the end of summer (from 22 December 2019 to 21 September 2020), there were a total of 58.9K (95%CI: 46.9K - 69.5K) excess deaths across all 31 provinces with 27% (95%CI: 20% - 34%) estimated nationwide exposure to SARS-CoV-2. In particular, 2 provinces in the central and northern Iran, namely Qom and Golestan, had the highest level of exposure with 57% (95%CI: 44% - 69%) and 56% (95%CI: 44% - 69%), respectively, while another 27 provinces had significant levels of excess mortality in at least one season with >20% population-level exposure to the virus. We also detected unexpectedly high levels of excess mortality during fall 2019 (from 23 September to 21 December 2019) across 18 provinces. Our findings suggest that this spike cannot be a result of an early cryptic transmission of COVID-19 across the country and is also inconsistent with the molecular phylogenetics estimates for the start of the pandemic and its arrival to Iran. However, in the absence of appropriate surveillance data for detecting severe acute respiratory infections we were unable to make a determination as to what caused the spike in fall 2019.

Conflict of Interest

None.
Light-dependent induction ofEdn2expression and attenuation of retinal pathology by endothelin receptor antagonists inProminin-1- deficient mice
Authors: Kobayashi Yuka , Watanabe Shizuka , Shirai Manabu , Yamashiro Chiemi , Ogata Tadahiko , Higashijima Fumiaki , Yoshimoto Takuya , Hayano Takahide , Asai Yoshiyuki , Sasai Noriaki , Kimura Kazuhiro
Published: December 2020

Retinitis pigmentosa (RP) and macular dystrophy (MD) are prevalent retinal degenerative diseases associated with gradual photoreceptor death. These diseases are often caused by genetic mutations that result in degeneration of the retina postnatally after it has fully developed. TheProminin-1gene (Prom1) is a causative gene for RP and MD, andProm1- knockout (KO) mice recapitulate key features of these diseases including light-dependent retinal degeneration and stenosis of retinal blood vessels. The mechanisms underlying progression of such degeneration have remained unknown, however. We here analysed early events associated with retinal degeneration inProm1-KO mice. We found that photoreceptor cell death and glial cell activation occur between 2 and 3 weeks after birth. High-throughput analysis revealed that expression of the endothelin-2 gene (Edn2) was markedly up-regulated in the Prom1-deficient retina during this period. Expression ofEdn2was also induced by light stimulation inProm1-KO mice that had been reared in the dark. Finally, treatment with endothelin receptor antagonists attenuated photoreceptor cell death, gliosis, and retinal vessel stenosis inProm1-KO mice. Our findings suggest that inhibitors of endothelin signalling may delay the progression of RP and MD and therefore warrant further study as potential therapeutic agents for these diseases.
Population genomics of the emerging forest pathogenNeonectria neomacrospora
Authors: Nielsen Knud Nor , Gopalakrishnan Shyam , Korneliussen Thorfinn Sand , Skovrind Mikkel , Sirén Kimmo , Petersen Bent , Sicheritz-Pontén Thomas , Thomsen Iben M. , Gilbert M. Thomas P. , Hansen Ole Kim
Published: December 2020

The fungal pathogenNeonectria neomacrosporais of increasing concern in Europe where, within the last decade, it has caused substantial damage to forest stands and ornamental trees of the genusAbies(Mill.). Using whole-genome sequencing of a comprehensive collection of isolates, we show the extent of three major clades withinN. neomacrospora, which most likely diverged around the end of the last Ice Age. We find it likely that the current European epidemic ofN. neomacrosporawas founded from a population belonging to the east North American clade. All European isolates (1957-2019) had a common evolutionary history, but substantial and asymmetrical gene flow from the larger American source population could be detected. The European population shows multiple signs of having gone through a bottleneck and subsequent population expansion.
DNA metabarcoding provides insights into the diverse diet of a dominant suspension feeder, the giant plumose anemoneMetridium farcimen
Authors: Wells Christopher D. , Paulay Gustav , Nguyen Bryan N. , Leray Matthieu
Published: December 2020

Benthic suspension feeders have significant impacts on plankton communities by depleting plankton or modifying composition of the plankton through prey selectivity. Quantifying diets of planktivorous animals can be difficult because plankton are frequently microscopic, may lack diagnostic characters, and are digested at variable rates. With DNA metabarcoding, the identification of gut contents has become faster and more accurate, and the technique allows for higher taxonomic resolution-n while also identifying rare and highly degraded items that would otherwise not be detected. We used DNA metabarcoding to examine the diet of the giant plumose anemoneMetridium farcimen,a large, abundant, competitively-dominant anemone on subtidal rock surfaces and floating docks in the northeast Pacific Ocean. Gut contents of 12 individuals were compared to 80- and 330-μm filtered plankton samples collected one hour prior between 0.02 and 1.5 km from the anemones. The objectives of this study were to determine ifM. farcimenhas a selective diet and compare our findings with traditional gut content analyses.Metridium farcimencaptured a wider range of prey than previously suspected and metabarcoding found many more taxa than traditional sampling techniques. Gut contents were less diverse than 80-μm filtered plankton samples, but more diverse than 330-μm filtered plankton samples. The diet of the anemones was 52% arthropods with a surprisingly high relative abundance of an ant (10%) that has mating flights in August when this study was conducted. The gut contents ofM. farcimenlikely include all prey that it can detect and that cannot escape. There were no overrepresented taxa in the gut contents compared to the plankton but there were underrepresented taxa. This study highlights the usefulness of the metabarcoding method in identifying prey within the gut of planktivorous animals and the significant terrestrial input into marine food webs.
Ambiguous support for extra-tropical accelerated diversification of rosids
Authors: Gamisch Alexander
Published: December 2020

Sun et al.1used a comprehensive phylogenetic and a locality based climatic dataset to examine how past climates have driven diversification across 17 orders of rosids. They concluded that tropical rosids diversify slower than the (younger) non-tropical counterparts in part due to a strong negative relationship between paleo-temperature and diversification (i.e. higher diversification rates under lower temperatures). Their conclusions are based on tip rates (derived from BAMM2; DR3) and correlations with current temperature (e-SIM4) or binary tropicality data sets (BAMM/STRAPP5; FiSSE6) as well as tree-wide estimates of diversification with paleo-temperature (BAMM; RPANDA7) or tropicality (BiSSE8; HiSSE9). Here, I highlight several inconsistencies in their diversification analyses as well as a systematic error pertaining to the RPANDA model selection procedure, which, together with several minor technical issues, weaken the support for Sun et al.’s1conclusions. A re-analysis of their BiSSE/HiSSE and RPANDA analyses are performed.
A massively multi-scale approach to characterising tissue architecture by synchrotron micro-CT applied to the human placenta
Authors: Tun W. M. , Poologasundarampillai G. , Bischof H. , Nye G. , King O. N. F. , Basham M. , Tokudome Y. , Lewis R. M. , Johnstone E. D. , Brownbill P. , Darrow M. , Chernyavsky I. L.
Published: December 2020

Multi-scale structural assessment of biological soft tissue is challenging but essential to gain insight into structure-function relationships of tissue/organ. Using the human placenta as an example, this study brings together sophisticated sample preparation protocols, advanced imaging, and robust, validated machine-learning segmentation techniques to provide the first massively multi-scale and multi-domain information that enables detailed morphological and functional analyses of both maternal and fetal placental domains. Finally, we quantify the scale-dependent error in morphological metrics of heterogeneous placental tissue, estimating the minimal tissue scale needed in extracting meaningful biological data. The developed protocol is beneficial for high-throughput investigation of structure-function relationships in both normal and diseased placentas, allowing us to optimise therapeutic approaches for pathological pregnancies. In addition, the methodology presented is applicable in characterisation of tissue architecture and physiological behaviours of other complex organs with similarity to the placenta, where an exchange barrier possesses circulating vascular and avascular fluid spaces.
High-resolution radioluminescence microscopy of FDG uptake in an engineered 3D tumor-stoma model
Authors: Khan Syamantak , Kim Sungwoo , Yang Yunzhi Peter , Pratx Guillem
Published: December 2020

PURPOSE

The increased glucose metabolism of cancer cells is the basis for18F-fluorodeoxyglucose positron emission tomography (FDG-PET). However, due to its coarse image resolution, PET is unable to resolve the metabolic role of cancer-associated stroma, which often influences the metabolic reprogramming of a tumor. This study investigates the feasibility of imaging engineered 3D tumor models with high resolution using FDG.

METHOD

Multicellular tumor spheroids (A549 lung adenocarcinoma) were co-cultured with GFP-expressing human umbilical vein endothelial cells (HUVECs) within an artificial extracellular matrix to mimic a tumor and its surrounding stroma. The tumor model was constructed as a thin 3D layer over a transparent CdWO4 scintillator plate to allow high-resolution imaging of the cultured cells. The radioluminescence signal was collected by a highly sensitive microscope and camera. Fluorescence microscopy was performed in the same instrument to localize endothelial and tumor cells.

RESULTS

Simultaneous brightfield and fluorescence imaging, co-localized with radioluminescence signal, provided high-resolution information on FDG accumulation in the engineered tissue. The microvascular stromal compartment took up a large fraction of FDG, comparable to the tumor spheroids. In vitro gamma counting also revealed that both A549 and HUVEC cells were highly glycolytic and characterized by rapid FDG-uptake kinetics.

CONCLUSION

Our study demonstrates the feasibility of imaging FDG distribution in tumor and stromal components separately with high spatial resolution in engineeredin vitrotumor models. Our imaging method is safe, simple, rapid, and can be easily used for otherin vitrocancer models, surgical tissue slices, and tumor biopsies to interrogate PET radiotracer uptake at high resolution.
Structural basis for piRNA-targeting
Authors: Anzelon Todd A. , Chowdhury Saikat , Hughes Siobhan M. , Xiao Yao , Lander Gabriel C. , MacRae Ian J.
Published: December 2020

Piwi proteins use PIWI-interacting RNAs (piRNAs) to identify and silence the transposable elements (TEs) pervasively found in animal genomes. The Piwi targeting mechanism is proposed to be similar to targeting by Argonaute proteins, which employ microRNA (miRNA) guides to repress cellular mRNAs, but has not been characterized in detail. We present cryo-EM structures of a Piwi-piRNA complex with and without target RNAs and analysis of target recognition. Resembling Argonaute, Piwi identifies targets using the piRNA seed-region. However, Piwi creates a much weaker seed so that prolonged target association requires further piRNA-target pairing. Beyond the seed, Piwi creates wide central cleft wide for unencumbered piRNA-target pairing, enabling long-lived Piwi-piRNA-target interactions that are tolerant of mismatches. Piwi ensures targeting fidelity by blocking propagation of the piRNA-target duplex in the absence of faithful seed pairing, and by requiring extended piRNA-target pairing to reach an endonucleolytically active conformation. This mechanism allows Piwi to minimize off-targeting cellular mRNAs and adapt piRNA sequences to evolving genomic threats.
Paradoxical effects of cigarette smoke and COPD on SARS-CoV2 infection and disease
Authors: Tomchaney M. , Contoli M. , Mayo J. , Baraldo S. , Shuaizhi L. , Cabel CR. , Bull DA. , Lick S. , Malo J. , Knoper S. , Kim SS. , Tram J. , Rojas-Quintero J. , Kraft M. , Ledford J. , Tesfaigzi Y. , Martinez F.D. , Thorne C. A. , Kheradmand F. , Campos S. K. , Papi A. , Polverino F.
Published: December 2020

Introduction

How cigarette smoke (CS) and chronic obstructive pulmonary disease (COPD) affect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and severity is controversial. We investigated the protein and mRNA expression of SARS-CoV-2 entry receptor ACE2 and proteinase TMPRSS2 in lungs from COPD patients and controls, and lung tissue from mice exposed acutely and chronically to CS. Also, we investigated the effects of CS exposure on SARS-CoV-2 infection in human bronchial epithelial cells.

Methods

In Cohort 1, ACE2-positive cells were quantified by immunostaining in FFPE sections from both central and peripheral airways. In Cohort 2, we quantified pulmonary ACE2 protein levels by immunostaining and ELISA, and both ACE2 and TMPRSS2 mRNA levels by RT-qPCR. In C57BL/6 WT mice exposed to air or CS for up to 6 months, pulmonary ACE2 protein levels were quantified by triple immunofluorescence staining and ELISA. The effects of CS exposure on SARS-CoV-2 infection were evaluated after 72hr in vitro infection of Calu-3 cells. After SARS-CoV-2 infection, the cells were fixed for IF staining with dsRNA-specific J2 monoclonal Ab, and cell lysates were harvested for WB of viral nucleocapsid (N) protein. Supernatants (SN) and cytoplasmic lysates were obtained to measure ACE2 levels by ELISA.

Results

In both human cohorts, ACE2 protein and mRNA levels were decreased in peripheral airways from COPD patients versus both smoker and NS controls, but similar in central airways. TMPRSS2 levels were similar across groups. Mice exposed to CS had decreased ACE2 protein levels in their bronchial and alveolar epithelia versus air-exposed mice exposed to 3 and 6 months of CS. In Calu3 cells in vitro, CS-treatment abrogated infection to levels below the limit of detection. Similar results were seen with WB for viral N protein, showing peak viral protein synthesis at 72hr.

Conclusions

ACE2 levels were decreased in both bronchial and alveolar epithelial cells from uninfected COPD patients versus controls, and from CS-exposed versus air-exposed mice. CS-pre-treatment did not affect ACE2 levels but potently inhibited SARS-CoV-2 replication in this in vitro model. These findings urge to further investigate the controversial effects of CS and COPD on SARS-CoV2 infection.
RIOK2 drives glioblastoma cell proliferation by modulating MYC through the RNA-binding protein IMP3
Authors: Chen Alexander S. , Marquez Jhomar , Boyd Nathaniel H. , Oh Se-Yeong , Gulbronson Riley , Hamilton Jamie A.G. , Legan Emily R. , Read Renee D.
Published: December 2020

Glioblastoma (GBM) is the most common primary malignant brain tumor and is resistant to current therapies. Here, using a combination of proteomic and genetic approaches in bothDrosophilaand human GBM models, we identify novel signaling effectors that act downstream of EGFR and PI3K signaling pathways, which drive GBM. Our results indicate that RIOK2, an atypical serine/threonine kinase, forms a complex with the RNA-binding protein, IMP3 (Imp inDrosophila), and that the dRIOK2/RIOK-Imp/IMP3 pathway is necessary for tumorigenesis. Furthermore, our results indicate that RIOK2 catalytic activity is important for neoplasia and recruits TORC2 to phosphorylate IMP3. Finally, the dRIOK2-Imp pathway regulates protein levels of dMyc, a known Imp target mRNA, and these key findings were recapitulated in human GBM models. Collectively, our data indicates that RIOK2 catalytic activity promotes the activation of TORC2 and recruitment of IMP3, which in turn, modulates MYC mRNA and protein levels to promote tumorigenesis.
Modern Japanese ancestry-derived variants revealed the formation process of the current Japanese regional gradations
Authors: Watanabe Yusuke , Ohashi Jun
Published: December 2020

Modern Japanese have two major ancestral populations: the indigenous Jomon hunter gatherers and continental East Asian farmers. To figure out the formation process of current Japanese population, we developed a reference-free detection method of variants derived from ancestral populations using a summary statistic, the ancestry-marker index (AMI). We confirmed by computer simulations thatAMIcan detect ancestry-derived variants even in an admixed population of recently diverged source populations with high accuracy, which cannot be achieved by the most widely used statistics, S*, for identifying archaic ancestry. We applied theAMIto modern Japanese samples and identified 208,648 single nucleotide polymorphisms (SNPs) that were likely derived from the Jomon people (Jomon-derived variants). The analysis of Jomon-derived variants in 10,842 modern Japanese individuals recruited from all over Japan revealed that the admixture proportions of the Jomon people varied between prefectures, probably due to the differences of population sizes of immigrants in the final Jomon to the Yayoi period. The estimated allele frequencies of genome-wide SNPs in the ancestral populations of modern Japanese suggested their phenotypic characteristics possibly for adaptation to their respective livelihoods; higher triglycerides and blood sugar for the Jomon ancestry and higher C-reactive protein and eosinophil counts for continental ancestry. According to our findings, we propose a formation model of modern Japanese population; regional variations in admixture proportions of the Jomon people and continental East Asians formed genotypic and phenotypic gradations of current Japanese archipelago populations.
Spatial organization of the transcriptome in individual neurons
Authors: Wang Guiping , Ang Cheen-Euong , Fan Jean , Wang Andrew , Moffitt Jeffrey R. , Zhuang Xiaowei
Published: December 2020

Neurons are highly polarized cells with complex neurite morphology. Spatial organization and local translation of RNAs in dendrites and axons play an important role in many neuronal functions. Here we performed super-resolution spatial profiling of RNAs inside individual neurons at the genome scale using multiplexed error-robust fluorescencein situhybridization (MERFISH), and mapped the spatial organization of up to ∼4,200 RNA species (genes) across multiple length scales, ranging from sub-micrometer to millimeters. Our data generated a quantitative intra-neuronal atlas of RNAs with distinct transcriptome compositions in somata, dendrites, and axons, and revealed diverse sub-dendritic distribution patterns of RNAs. Moreover, our spatial analysis identified distinct groups of genes exhibiting specific spatial clustering of transcripts at the sub-micrometer scale that were dependent on protein synthesis and differentially dependent on synaptic activity. Overall, these data provide a rich resource for characterizing the subcellular organization of the transcriptome in neurons with high spatial resolution.
Mutated clones driving leukemic transformation are already detectable at the single cell level in CD34-positive cells in the chronic phase of primary myelofibrosis
Authors: Parenti Sandra , Rontauroli Sebastiano , Carretta Chiara , Mallia Selene , Genovese Elena , Chiereghin Chiara , Peano Clelia , Tavernari Lara , Bianchi Elisa , Fantini Sebastian , Sartini Stefano , Romano Oriana , Bicciato Silvio , Tagliafico Enrico , Della Porta Matteo , Manfredini Rossella
Published: December 2020

Disease progression of myeloproliferative neoplasms is the result of increased genomic complexity. Since the ability to predict disease evolution is crucial for clinical decision, we studied single cell genomics and transcriptomics of CD34-positive cells from a primary myelofibrosis (PMF) patient who progressed to acute myeloid Leukemia (AML) while receiving Ruxolitinib.

Single cell genomics allowed the reconstruction of clonal hierarchy and demonstrated thatTET2was the first mutated gene whileFLT3was the last one. Disease evolution was accompanied by increased clonal heterogeneity and mutational rate, but clones carryingTP53andFLT3mutations were already present in chronic phase.

Single cell transcriptomics unraveled repression of interferon signaling suggesting an immunosuppressive effect exerted by Ruxolitinib. Moreover, AML transformation was associated with a differentiative block and immune escape.

These results suggest that single cell analysis can unmask tumor heterogeneity and provide meaningful insights about PMF progression that might guide personalized therapy.
Central metabolism as a potential origin of sex differences in morphine analgesia but not in the induction of analgesic tolerance in mice
Authors: Gabel Florian , Hovhannisyan Volodya , Andry Virginie , Goumon Yannick
Published: December 2020

In rodents, morphine analgesia is influenced by sex. However, conflicting results exist regarding the interaction between sex and morphine analgesic tolerance. Morphine is metabolized in the liver and brain into morphine-3-glucuronide (M3G). Sex differences in morphine metabolism and differential metabolic adaptations during tolerance development might explain the behavioral discrepancies. The present article investigates the differences in peripheral and central morphine metabolism after acute and chronic morphine treatment in male and female mice.

The first experiment aimed to determine whether morphine analgesia and tolerance differ between male and female mice using the tail-immersion test. The second experiment evaluated morphine and M3G metabolic kinetics in the blood using LC-MS/MS. Morphine and M3G were also quantified in several central nervous system (CNS) regions after acute and chronic morphine treatment. Finally, the blood-brain barrier permeability of M3G was assessed in male and female mice.

This study demonstrated that female mice showed weaker morphine analgesia. In addition, tolerance appeared earlier in females but the sex discrepancies observed seemed to be due to the initial differences in morphine analgesia rather than to sex-specific mechanisms involving metabolism. Additionally, compared to male mice, female mice showed higher levels of M3G in the blood and in several CNS regions, whereas lower levels of morphine were observed in these brain regions. These differences are attributable mainly to morphine central metabolism, which differed between males and females in pain-related brain regions, consistent with the weaker analgesic effect in females. However, the role of morphine metabolism in analgesic tolerance seems rather limited.
Non-autophagy role of Atg5 and NBR1 in unconventional secretion of IL-12 prevents gut dysbiosis and inflammation
Authors: Merkley Seth D. , Goodfellow Samuel M. , Guo Yan , Wilton Zoe E.R. , Byrum Janie R. , Schwalm Kurt C. , Dinwiddie Darrell L. , Gullapalli Rama R. , Deretic Vojo , Hernandez Anthony Jimenez , Bradfute Steven B. , In Julie G. , Castillo Eliseo F.
Published: December 2020

Intestinal myeloid cells play a critical role in balancing intestinal homeostasis and inflammation. Here, we report that expression of the autophagy related 5 (Atg5) protein in myeloid cells prevents dysbiosis and excessive intestinal inflammation by limiting IL-12 production. Mice with a selective genetic deletion ofAtg5in myeloid cells (Atg5ΔMye) showed signs of dysbiosis prior to colitis and exhibited severe intestinal inflammation upon colitis induction that was characterized by increased IFNγ production. This increase in IFNγ was due to excess IL-12 secretion fromAtg5-deficient myeloid cells. Atg5 functions to limit IL-12 secretion through modulation of late endosome (LE) acidity. Additionally, the autophagy cargo receptor NBR1, which accumulates in Atg5-deficient cells, played a role by delivering IL-12 to LE. Restoration of the intestinal microbiota and alleviation of intestinal inflammation was achieved by genetic deletion of IL-12 in Atg5ΔMye mice. In summary, Atg5 expression in intestinal myeloid cells acts as an anti-inflammatory brake to regulate IL-12 thus preventing dysbiosis and uncontrolled IFNγ-driven intestinal inflammation.
Corpus callosum morphology across the lifespan in baboons (Papio anubis): a cross-sectional study of relative mid-sagittal surface area and thickness
Authors: Westerhausen René , Meguerditchian Adrien
Published: December 2020

The axons forming the corpus callosum enable integration and coordination of cognitive processing between the cerebral hemispheres. In the aging human brain, these functions are affected by progressive axon and myelin deteriorations, which results in a substantial atrophy of the midsagittal corpus callosum in old age. In non-human primates, these degenerative processes are less pronounced as previous morphometric studies on capuchin monkey, rhesus monkeys, and chimpanzees do not find old-age callosal atrophy. The objective of the present study was to extend these previous findings by studying the aging trajectory of the corpus callosum of the olive baboon (Papio anubis) across the lifespan. For this purpose, total relative (to forebrain volume) midsagittal area, subsectional area, and regional thickness of the corpus callosum was assessed in 91 male and female animals using non-invasive MRI-based morphometry. The studied age range was 2.5 to 26.6 years, and the sample included 11 old-age animals (above the age of 20 years). Fitting lifespan trajectories using general additive modelling (GAM) we found that the relative area of the total corpus callosum and the anterior subsection follow a positive linear trajectory. That is, both measures increased slowly but continuously from childhood into old age, and no stagnation of growth or decline was observed in old age. Thus, comparable with all other non-human primates studied to-date, baboons do not show callosal atrophy in old age. This observation lends supports to the notion that atrophy of the corpus callosum is a unique characteristic of human brain aging.
Easyreporting simplifies the implementation of Reproducible Research Layers in R software
Authors: Righelli Dario , Angelini Claudia
Published: December 2020

During last years “irreproducibility” became a general problem in omics data analysis due to the use of sophisticated and poorly described computational procedures. For avoiding misleading results, it is necessary to inspect and reproduce the entire data analysis as a unified product. Reproducible Research (RR) provides general guidelines for public access to the analytic data and related analysis code combined with natural language documentation, allowing third-parties to reproduce the findings. We developedeasyreporting, a novel R/Bioconductor package, to facilitate the implementation of an RR layer inside reports/tools without requiring any knowledge of the R Markdown language. We describe the main functionalities and illustrate how to create an analysis report using a typical case study concerning the analysis of RNA-seq data. Then, we also show how to trace R functions automatically. Thanks to this latter feature,easyreportingresults beneficial for developers to implement procedures that automatically keep track of the analysis steps within Graphical User Interfaces (GUIs).Easyreportingcan be useful in supporting the reproducibility of any data analysis project and the implementation of GUIs. It turns out to be very helpful in bioinformatics, where the complexity of the analyses makes it extremely difficult to trace all the steps and parameters used in the study.