bluuebunny/crossref_metadata_2025 huggingface dataset:
Dual Plasma Sampling Method to Determine the Hepatic and Renal Clearance of the 2 Diastereoisomers of Gd-EOB-DTPA
Authors: Yuan Xiaodong , Wang Su , Shi Wenwei , Cai Yu , Chen Yingxin , Chen Ming , Peng Yanghong , Ao Guokun
Published: January 2020

Objectives

The aim of this study was to develop a method to determine hepatic and renal clearance of the 2 diastereoisomers (Gd-A, Gd-B) of Gd-EOB-DTPA separately.

Materials and Methods

Between July 2017 and February 2018, 41 patients with hepatic disease were prospectively included. For each patient, 1 mL of iopromide (to determine glomerular filtration rate [GFR]) was coadministered with Gd-EOB-DTPA (Gd-A and Gd-B; 65:35 wt/wt). The plasma clearances of Gd-A (PCL-GdA) and Gd-B (PCL-GdB) as well as the iopromide (GFR) were generated by using dual plasma sampling method. Meanwhile, the patient's urine was collected for measurement of renal clearance of Gd-A (RCL-GdA) and Gd-B (RCL-GdB) to confirm its agreement with GFR. Hepatic clearances of Gd-A (HCL-GdA) and Gd-B (HCL-GdB) were calculated by subtracting the GFR from PCL-GdA and PCL-GdB, respectively, and were correlated with indocyanine green (ICG) 15 minutes retention rate (ICG R15). Pharmacokinetic parameters were compared between the two isomers and between Child–Pugh classifications using student’s t test.

Results

Within the group of 41 patients evaluated, both RCL-GdA and RCL-GdB demonstrated a good correlation and agreement to GFR (statistics shown in the main body). HCL-GdA demonstrated a strong negative correlation (r = −0.86, P < 0.001) with ICG R15 and was much higher than HCL-GdB (116.18 ± 75.48 vs 19.74 ± 14.24 mL/min, P < 0.001). HCL-GdB demonstrated a weak correlation (r = −0.26, P = 0.102) with ICG R15. HCL-GdA of noncirrhosis and Child-Pugh class A (151.74 ± 68.28 mL/min, n = 26) was higher than that of Child-Pugh class B (54.54 ± 39.13 mL/min, n = 15; P = 0.001).

Conclusions

A practical method was established for the determination of hepatic and renal clearance of the 2 isomers of Gd-EOB-DTPA. The 2 isomers have equal renal clearance and different hepatic clearance. The HCL-GdA may serve as a novel marker to reflect liver function reserve.
Access to Multidisciplinary Care for Patients With 22q11.2 Deletion Syndrome: Identifying Breakdowns in the Screening Process
Authors: Abu-Ghname Amjed , Perdanasari Aurelia T. , Raj Sarth , Seema Jain , Wilson Kristina D. , Maricevich Renata S.
Published: January 2020

The 22q11.2 deletion syndrome affects approximately 1 in 4000 live births and involves cardiac defects, immunodeficiency, and endocrine disruption. The complexity of diagnosis and multifaceted care often leads to fragmented management in the short and long term. With the purpose of developing an effective multidisciplinary program, the authors aimed to identify the deficiencies in current screening and referral processes among the teams required in the care for patients with 22q11.2 deletion syndrome. A retrospective chart review was conducted at our institution between 2001 and 2016. Patients with confirmed 22q11.2 deletion diagnoses between the ages of 0 and 28 were included. A list of 15 relevant specialties that should evaluate patients with 22q11.2 deletion syndrome was created according to established guidelines. Patient medical and demographic information were collected and analyzed. A total of 270 patients were included. Mean age at diagnosis was 3.3 years. On average, patients visited 6 of 15 departments (1–14). Only 8.8% of patients visited >10 specialties. The majority were seen by Cardiology, Allergy and Immunology, Genetics, and Speech (57.4–87.8%). A minority were seen by Hematology and Oncology, Sleep Therapy, and Physical Therapy (13.3–16.3%). Only 34.1% encountered plastic surgery. Negative correlation (−0.128; P = 0.035) was demonstrated between patients’ age at diagnosis and number of specialty teams encountered. This study highlights the current underutilization of services required to manage patients with 22q11.2 deletion syndrome. While screening guidelines have been established, implementation can be challenging as it requires efficient care coordination between teams. Moving forward, the authors believe that a multidisciplinary clinical approach to streamline patient care is necessary.
Effect of light and darkness on the growth and development of downy mildew pathogenHyaloperonospora arabidopsidis
Authors: Telli Osman , Jimenez-Quiros Catherine , McDowell John M. , Tör Mahmut
Published: January 2020

Disease development in plants requires a susceptible host, a virulent pathogen, and a favourable environment. Oomycete pathogens cause many important diseases and have evolved sophisticated molecular mechanisms to manipulate their hosts. Day length has been shown to impact plant-oomycete interactions but a need exists for a tractable reference system to understand the mechanistic interplay between light regulation, oomycete pathogen virulence, and plant host immunity. Here we present data demonstrating that light is a critical factor in the interaction betweenArabidopsis thalianaand its naturally occurring downy mildew pathogenHyaloperonospora arabidopsidis(Hpa). We investigated the role of light on spore germination, mycelium development, sporulation and oospore formation ofHpa, along with defence responses in the host. We observed abundantHpasporulation on compatible Arabidopsis under day lengths ranging from 10 to 14 hours. Contrastingly, exposure to constant light or constant dark suppressed sporulation. Exposure to constant dark suppressed spore germination, mycelial development and oospore formation. Interestingly, exposure to constant light stimulated spore germination, mycelial development and oospore formation. A biomarker of plant immune system activation was induced under both constant light and constant dark. Altogether, these findings demonstrate thatHpahas the molecular mechanisms to perceive and respond to light and that both the host and pathogen responses are influenced by the light regime. Therefore, this pathosystem can be used for investigations to understand the molecular mechanisms through which oomycete pathogens likeHpaperceive and integrate light signals, and how light influences pathogen virulence and host immunity during their interactions.
RECAS9: Recombining wild species introgression via mitotic gene editing in barley
Authors: Lazar Shelly , Prusty Manas Ranjan , Bishara Khaled , Sherman Amir , Fridman Eyal
Published: January 2020

Genetic loci underlying variation in traits with agronomic importance or genetic risk factors in human diseases have been identified by linkage analysis and genome-wide association studies. However, narrowing down the mapping to the individual causal genes and variations within these is much more challenging, and so is the ability to break linkage drag between beneficial and unfavourable loci in crop breeding. We developed RECAS9 as a transgene-free approach for precisely targeting recombination events by delivering CRISPR/Cas9 ribonucleotide protein (RNP) complex into heterozygous mitotic cells for the barley (Hordeum vulgare)Heat3.1locus. A wild species (H. spontaneum) introgression in this region carries the agronomical unfavourable toughrachisphenotype (non-brittle) allele linked with a circadian clock accelerating QTL near GIGANTEA gene. We delivered RNP, which was targeted between two single nucleotide polymorphism (SNPs), to mitotic calli cells by particle bombardment. We estimated recombination events by next generation sequencing (NGS) and droplet digital PCR (ddPCR). While NGS analysis grieved from confounding effects of PCR recombination, ddPCR analysis allowed us to associate RNP treatment on heterozygous individuals with significant increase of homologous directed repair (HDR) between cultivated and wild alleles, with recombination rate ranging between zero to 57%. These results show for the first time in plants a directed and transgene free mitotic recombination driven by Cas9 RNP, and provide a starting point for precise breeding and fine scale mapping of beneficial alleles from crop wild relatives.
Macroevolutionary analysis of discrete character evolution using parsimony-informed likelihood
Authors: Grundler Michael C. , Rabosky Daniel L.
Published: January 2020

Abstract

Rates of character evolution in macroevolutionary datasets are typically estimated by maximizing the likelihood function of a continuous-time Markov chain (CTMC) model of character evolution over all possible histories of character state change, a technique known as maximum average likelihood. An alternative approach is to estimate ancestral character states independently of rates using parsimony and to then condition likelihood-based estimates of transition rates on the resulting ancestor-descendant reconstructions. We use maximum parsimony reconstructions of possible pathways of evolution to implement this alternative approach for single-character datasets simulated on empirical phylogenies using a two-state CTMC. We find that transition rates estimated using parsimonious ancestor-descendant reconstructions have lower mean squared error than transition rates estimated by maximum average likelihood. Although we use a binary state character for exposition, the approach remains valid for an arbitrary number of states. Finally, we show how this method can be used to rapidly and easily detect phylogenetic variation in tempo and mode of character evolution with two empirical examples from squamates. These results highlight the mutually informative roles of parsimony and likelihood when testing hypotheses of character evolution in macroevolution.
Role of DNA Methylation in Persister Formation in UropathogenicE. coli
Authors: Xu Yuanyuan , Liu Shuang , Zhang Ying , Zhang Wenhong
Published: January 2020

UropathogenicEscherichia coli(UPEC) persister bacteria play crucial roles in clinical treatment failure and relapse. DNA methylation is known to regulate gene expression in bacteria, but its role in persister formation has not been investigated. Here, we created adenine methylation deletion mutant (Δdam) and cytosine methylation mutant (Δdcm) from UPEC strain UTI89 and found that theΔdammutant but notΔdcmmutant had significant defect in persister formation during exposure to various antibiotics (gentamicin, fluroquinolones and cephalosporin) and stresses (acid pH and hyperosmosis), and that complementation of thedammutant restored its persister defect phenotype. PacBio sequencing of epigenetic genomewide methylation signature and RNA sequencing of theΔdammutant were performed to define, for the first time, the role of adenine methylation in persister formation. Methylome data analysis showed that 99.73% of m6A modifications on GATC were demethylated in theΔdammutant, and demethylation nucleotide site related genes suggested an overwhelming effect on transcription and metabolic processes. Transcriptome analysis of theΔdammutant in comparison to wild type showed that flagella biosynthesis, galactitol transport/utilization, and signaling related genes were upregulated while pilus, fimbriae, virulence, glycerol, nitrogen metabolism pathways and transcriptional regulators were downregulated. The comparative COG analysis of methylome and transcriptome enriched pathways identified translation, ribosomal structure and biogenesis, and cell motility were upregulated, whereas DNA repair, secondary metabolite biosynthesis and diverse transport systems, some of which are known to be involved in persister formation, were downregulated in theΔdammutant. These findings provide new insights about the molecular basis of how DNA adenine methylation may be involved in persister formation and offer novel therapeutic targets for combating persister bacteria.
Development and validation of a potent and specific inhibitor for the CLC-2 chloride channel
Authors: Koster Anna K. , Reese Austin L. , Kuryshev Yuri , Wen Xianlan , McKiernan Keri A. , Gray Erin E. , Wu Caiyun , Huguenard John R. , Maduke Merritt , Bois J. Du
Published: January 2020

CLC-2 is a voltage-gated chloride channel that is widely expressed in many mammalian tissues. In the central nervous system (CNS), CLC-2 is expressed in neurons and glia. Studies to define how this channel contributes to normal and pathophysiological function in the CNS have been controversial, in part due to the absence of precise pharmacological tools for modulating CLC-2 activity. Herein, we describe the development and optimization of AK-42, a specific small-molecule inhibitor of CLC-2 with nanomolar potency (IC50 = 17 ± 1 nM). AK-42 displays unprecedented selectivity (>1000-fold) over CLC-1, the closest CLC-2 homolog, and exhibits no off-target engagement against a panel of 58 common channels, receptors, and transporters expressed in brain tissue. Computational docking, validated by mutagenesis and kinetic studies, indicates that AK-42 binds to an extracellular vestibule above the channel pore. In electrophysiological recordings of mouse CA1 hippocampal pyramidal neurons, AK-42 acutely and reversibly inhibits CLC-2 currents; no effect on current is observed on brain slices taken from CLC-2 knockout mice. These results establish AK-42 as a powerful new tool for investigating CLC-2 neurophysiology.

Significance Statement

The CLC-2 ion channel facilitates selective passage of Cl− ions across cell membranes. In the central nervous system (CNS), CLC-2 is expressed in both neurons and glia and is proposed to regulate electrical excitability and ion homeostasis. CLC-2 has been implicated in various CNS disorders, including certain types of epilepsy and leukodystrophy. Establishing a causative role for CLC-2 in neuropathologies, however, has been limited by the absence of selective reagents that enable acute and specific channel modulation. Our studies have resulted in the identification of a highly potent, small-molecule inhibitor that enables specific block of CLC-2 Cl− currents in hippocampal brain slices. This precise molecular tool should enable future efforts to identify and treat CLC-2-related disease.
Receptor compaction and GTPase movements drive cotranslational protein translocation
Authors: Lee Jae Ho , Chung SangYoon , Fu Yu-Hsien Hwang , Qian Ruilin , Sun Xuemeng , Weiss Shimon , Shan Shu-ou
Published: January 2020

Signal recognition particle (SRP) is a universally conserved targeting machine that couples the synthesis of ~30% of the proteome to their proper membrane localization1,2. In eukaryotic cells, SRP recognizes translating ribosomes bearing hydrophobic signal sequences and, through interaction with SRP receptor (SR), delivers them to the Sec61p translocase on the endoplasmic reticulum (ER) membrane1,2. How SRP ensures efficient and productive initiation of protein translocation at the ER is not well understood. Here, single molecule fluorescence spectroscopy demonstrates that cargo-loaded SRP induces a global compaction of SR, driving a >90 Å movement of the SRP•SR GTPase complex from the vicinity of the ribosome exit, where it initially assembles, to the distal site of SRP. These rearrangements bring translating ribosomes near the membrane, expose conserved Sec61p docking sites on the ribosome and weaken SRP’s interaction with the signal sequence on the nascent polypeptide, thus priming the translating ribosome for engaging the translocation machinery. Disruption of these rearrangements severely impairs cotranslational protein translocation and is the cause of failure in an SRP54 mutant linked to severe congenital neutropenia. Our results demonstrate that multiple largescale molecular motions in the SRP•SR complex are required to drive the transition from protein targeting to translocation; these post-targeting rearrangements provide potential new points for biological regulation as well as disease intervention.
Carbamazepine restores neuronal signaling, protein synthesis and cognitive function in a mouse model of fragile X syndrome
Authors: Ding Qi , Zhang Fan , Feng Yue , Wang Hongbing
Published: January 2020

Fragile X syndrome (FXS) is a leading genetic disorder of intellectual disability caused by the loss of the functional fragile X mental retardation protein (FMRP). To date, there is no efficacious mechanism-based medication for FXS. With regard to potential disease mechanisms in FXS, it is widely accepted that the lack of FMRP causes elevated protein synthesis and deregulation of neuronal signaling. Abnormal enhancement of the ERK½ (extracellular signal-regulated kinase ½) and PI3K-Akt (Phosphoinositide 3 kinase-protein kinase B) signaling pathways has been identified in both FXS patients and FXS mouse models. In this study, we show that carbamazepine, which is an FDA-approved drug and has been mainly used to treat seizure and neuropathic pain, corrects cognitive deficits including passive avoidance and object location memory in FXS mice. Carbamazepine also rescues hyper locomotion and social deficits. At the cellular level, carbamazepine dampens the elevated level of ERK½ and Akt signaling as well as protein synthesis in FXS mouse neurons. Together, these results advocate repurposing carbamazepine for FXS treatment.
Post-transcriptional Regulation is the Major Driver of microRNA Expression Variation
Authors: Zhang Dingyao , Tian Jingru , Roden Christine , Lu Jun
Published: January 2020

MicroRNA (miRNA) expression patterns are highly variable across human tissues and across cancer specimens. The intuitive assumption is that transcription is the main contributor to mature miRNA expression patterns, with post-transcriptional processes further modifying miRNA expression levels. Here we report the surprising model that, on the global level, post-transcriptional regulation dominates over transcriptional regulation in determining mature miRNA expression patterns in both normal tissues and cancer. Taking advantage of large genomic datasets in which the expression of both mature miRNAs and their host genes have been quantified, we establish and validate transcriptional and post-transcriptional metrics, with miRNA host gene expression estimating transcriptional regulation and mature miRNA to host gene ratio estimating post-transcriptional regulation. On average, the post-transcriptional metric contributes 2.8-fold more than the transcriptional metric to the variance of mature miRNA expression. The variation of the balance between the two mature miRNAs (5p and 3p miRNAs) produced from the same precursor hairpin is a non-negligible contributor to miRNA expression, explaining ∼27% of the variance of miRNAs’ post-transcriptional metric. Data of normal tissues yield similar results as cancer specimens. Additionally, the post-transcriptional metric is superior to the transcriptional metric in classifying cancer types. We further demonstrate that the post-transcriptional metric separates miRNAs into distinct groups, suggesting that there are groups of miRNAs that are co-regulated on the post-transcriptional level. Our data support a model in which the post-transcriptional regulation is the major driver of miRNA expression variation, and paves a way toward better mechanistic understanding of post-transcriptional regulation of mature miRNA expression.
cAMP-Fyn signaling in the dorsomedial striatum direct pathway drives excessive alcohol use
Authors: Ehinger Yann , Morisot Nadege , Phamluong Khanhky , Sakhai Samuel A. , Soneja Drishti , Adrover Martin F. , Alvarez Veronica A. , Ron Dorit
Published: January 2020

Fyn kinase in the dorsomedial striatum (DMS) of rodents plays a central role in mechanisms underlying excessive alcohol intake. The DMS is comprised of medium spiny neurons (MSNs) that project directly (dMSNs) or indirectly (iMSNs) to the substantia nigra. Here, we examined the cell-type specificity of Fyn’s actions on alcohol use. First, we knocked down Fyn selectively in DMS dMSNs or iMSNs of mice and measured the level of alcohol consumption. We found that downregulation of Fyn in dMSNs, but not in iMSNs, reduces excessive alcohol but not saccharin intake. D1Rs are coupled to Gαs/olf, which activate cAMP signaling. To examine whether Fyn’s actions are mediated through cAMP signaling, DMS dMSNs were infected with GαsDREADD, and the activation of Fyn signaling was measured following CNO treatment. We found that remote stimulation of cAMP signaling in DMS dMSNs activates Fyn and promotes the phosphorylation of the Fyn substrate, GluN2B. In contract, remote activation of GαsDREADD in DLS dMSNs did not alter Fyn signaling. We then tested whether activation of GαsDREADD in DMS dMSNs or iMSNs alters alcohol intake and observed that CNO-dependent activation of GαsDREADD in DMS dMSNs but not iMSNs increases alcohol but not saccharin intake. Finally, we examined the contribution of Fyn to GαsDREADD-dependent increase in alcohol intake, and found that systemic administration of the Fyn inhibitor, AZD0503 blocks GαsDREADD-dependent increase in alcohol consumption. Our results suggest that the cAMP-Fyn axis in the DMS dMSNs is a molecular transducer of mechanisms underlying the development of excessive alcohol consumption.
Reconstruction of Gene Regulatory Networks by integrating biological model and a recommendation system
Authors: Wang Yijie , Fear Justin M , Berger Isabelle , Lee Hangnoh , Oliver Brian , Przytycka Teresa M
Published: January 2020

Gene Regulatory Networks (GRNs) control many aspects of cellular processes including cell differentiation, maintenance of cell type specific states, signal transduction, and response to stress. Since GRNs provide information that is essential for understanding cell function, the inference of these networks is one of the key challenges in systems biology. Leading algorithms to reconstruct GRN utilize, in addition to gene expression data, prior knowledge such as Transcription Factor (TF) DNA binding motifs or results of DNA binding experiments. However, such prior knowledge is typically incomplete hence resulting in missing values. Therefore, the integration of such incomplete prior knowledge with gene expression to elucidate the underlying GRNs remains difficult.

To address this challenge we introduce NetREX-CF – RegulatoryNetworkReconstruction usingEXpression andCollaborativeFiltering – a GRN reconstruction approach that brings together a modern machine learning strategy (Collaborative Filtering model) and a biologically justified model of gene expression (sparse Network Component Analysis based model). The Collaborative Filtering (CF) model is able to overcome the incompleteness of the prior knowledge and make edge recommends for building the GRN. Complementing CF, the sparse Network Component Analysis (NCA) model can use gene expression data to validate the recommended edges. Here we combine these two approaches using a novel data integration method and show that the new approach outperforms the currently leading GRN reconstruction methods.

Furthermore, our mathematical formalization of the model has lead to a complex optimization problem of a type that has not been attempted before. Specifically, the formulation containsℓ0norm that can not be separated from other variables. To fill this gap, we introduce here a new method Generalized PALM (GPALM) that allows us to solve a broad class of non-convex optimization problems and prove its convergence.
In depth analysis of kinase cross screening data to identify CAMKK2 inhibitory scaffolds
Authors: O’Byrne Sean N. , Scott John W. , Pilotte Joseph R. , Santiago André de S. , Langendorf Christopher G. , Oakhill Jonathan S. , Eduful Benjamin J. , Couñago Rafael M. , Wells Carrow I. , Zuercher William J. , Willson Timothy M. , Drewry David H.
Published: January 2020

The calcium/calmodulin-dependent protein kinase kinase 2 (CAMKK2) plays a central role in many cell signaling pathways. CAMKK2 activates CAMK1, CAMK4, AMPK, and AKT leading to numerous physiological responses. Deregulation of CAMKK2 is linked to several diseases, suggesting utility of CAMKK2 inhibitors for oncological, metabolic and inflammatory indications. In this work we review the role of CAMKK2 in biology and disease. Through analysis of literature and public databases we have identified starting points for CAMKK2 inhibitor medicinal chemistry campaigns. These starting points provide an opportunity for the development of selective CAMKK2 inhibitors and will lead to tools that delineate the roles of this kinase in disease biology.
Life histories and niche dynamics in late Quaternary proboscideans from Midwestern North America: evidence from stable isotope analyses
Authors: Widga Chris , Hodgins Greg , Kolis Kayla , Lengyel Stacey , Saunders Jeff , Walker J. Douglas , Wanamaker Alan D.
Published: January 2020

Stable isotopes of mammoths and mastodons have the potential to illuminate ecological changes in late Pleistocene landscapes and megafaunal populations as these species approached extinction. The ecological factors at play in this extinction remain unresolved, but isotopes of bone collagen (δ13C, δ15N) and tooth enamel (δ13C, δ18O,87Sr/86Sr) from the Midwest, USA are leveraged to examine ecological and behavioral changes that occurred during the last interglacial-glacial cycle. Both species had significant C3 contributions to their diets and experienced increasing levels of niche overlap as they approached extinction. A subset of mastodons after the last glacial maximum (LGM) exhibit low δ15N values that may represent expansion into a novel ecological niche, perhaps densely occupied by other herbivores. Stable isotopes from serial and micro-sampled enamel show increasing seasonality and decreasing temperatures as mammoths transitioned from Marine Isotope Stage (MIS) 5e to glacial conditions (MIS 4, MIS 3, MIS 2). Isotopic variability in enamel suggests mobility patterns and life histories have potentially large impacts on the interpretation of their stable isotope ecology. This study further refines the ecology of midwestern mammoths and mastodons demonstrating increasing seasonality and niche overlap as they responded to landscape changes in the final millennia before extinction.
Neuronal dynamics of signal selective motor plan cancellation in the macaque dorsal premotor cortex
Authors: Franco Giarrocco , Giampiero Bardella , Margherita Giamundo , Francesco Fabbrini , Emiliano Brunamonti , Pierpaolo Pani , Stefano Ferraina
Published: January 2020

Primates adopt various strategies to interact with the environment. Yet, no study has examined the effects of behavioral strategies with regard to how movement inhibition is implemented at the neuronal level. We modified a classical approach to study movement control (stop-task) by adding an extra signal – termed the Ignore signal – which influenced movement inhibition only under a specific strategy. We simultaneously recorded multisite neuronal activity from the dorsal premotor (PMd) cortex of macaque monkeys during a task and applied a state-space approach. As a result, we found that movement generation is characterized by neuronal dynamics that evolve between subspaces. When the movement is halted, this evolution is arrested and inverted. Conversely, when the Ignore signal is presented, inversion of the evolution is observed briefly and only when a specific behavioral strategy is adopted. Moreover, neuronal signatures during the inhibitory process were predictive of how PMd processes inhibitory signals, allowing the classification of the resulting behavioral strategy. Our data corroborate the PMd as a critical node in movement inhibition.
Over-Confluence of expanded bone marrow mesenchymal stem cells ameliorates their chondrogenic capacity in 3D cartilage tissue engineering
Authors: Tucker Damien , Still Karen , Blom Ashley , Hollander Anthony P. , Kafienah Wael
Published: January 2020

Cartilage tissue engineering using bone marrow-derived mesenchymal stem cells (BM-MSCs) is a growing technology for the repair of joint defects. Culturing BM-MSCs to over confluence has historically been avoided due to perceived risk to cell viability, growth inhibition and differentiation potential. Here we show that a simple change in culture practice, based on mimicking the condensation phase during embryonic cartilage development, results in biochemically and histologically superior cartilage tissue engineered constructs. Whole transcriptome analysis of the condensing cells revealed a phenotype associated with early commitment to chondrogenic precursors. This simple adjustment to the common stem cell culture technique would impact the quality of all cartilage tissue engineering modalities utilising these cells.
Regulation of canonical Wnt signalling by the ciliopathy protein MKS1 and the E2 ubiquitin-conjugating enzyme UBE2E1
Authors: Szymanska Katarzyna , Boldt Karsten , Logan Clare V. , Adams Matthew , Robinson Philip A. , Ueffing Marius , Zeqiraj Elton , Wheway Gabrielle , Johnson Colin A.
Published: January 2020

A functional primary cilium is essential for normal and regulated signalling. Primary ciliary defects cause a group of developmental conditions known as ciliopathies, but the precise mechanisms of signal regulation by the cilium remain unclear. Previous studies have implicated the ubiquitin proteasome system (UPS) in regulation of Wnt signalling at the ciliary basal body. Here, we provide mechanistic insight into ciliary ubiquitin processing in cells and for a ciliopathy mouse model lacking the ciliary protein Mks1.In vivoloss of Mks1 sensitizes cells to proteasomal disruption, leading to abnormal accumulation of ubiquitinated proteins. To substantiate a direct link between MKS1 and the UPS, we identified UBE2E1, an E2 ubiquitin-conjugating enzyme that polyubiquitinates β-catenin, and RNF34, an E3 ligase, as novel interactants of MKS1. UBE2E1 and MKS1 colocalized, particularly during conditions of ciliary resorption, and loss of UBE2E1 recapitulates the ciliary and Wnt signalling phenotypes observed during loss of MKS1. Levels of UBE2E1 and MKS1 are co-dependent and UBE2E1 mediates both regulatory and degradative ubiquitination of MKS1. Furthermore, we demonstrate that processing of phosphorylated β-catenin occurs at the ciliary base through the functional interaction between UBE2E1 and MKS1. These observations suggest that correct β-catenin levels are tightly regulated at the primary cilium by a ciliary-specific E2 (UBE2E1) and a regulatory substrate-adaptor (MKS1), confirming the fundamental role of UPS defects in the molecular pathogenesis of ciliopathies.
Modeling the immunological pre-adaptation of HIV-1
Authors: van Dorp Christiaan H. , van Boven Michiel , de Boer Rob J.
Published: January 2020

It is becoming increasingly evident that the evolution of HIV-1 is to a large extent determined by the immunological background of the host. On the population-level this results in associations between specific human leukocyte antigen (HLA) alleles and polymorphic loci of the virus. Furthermore, some HLA alleles that were previously associated with slow progression to AIDS have been shown to lose their protective effect, because HLA-specific immunological escape variants have spread through the population. This phenomenon is known as immunological pre-adaptation. Apart from adapting to human immune responses, the set-point virus load (SPVL) of HIV-1 is thought to have evolved to values that optimize the population-level fitness of the virus. This suggestion is supported by considerable heritability of the SPVL. Previous modeling studies show that whether or not SPVL optimization is expected to occur depends sensitively on the underlying assumptions with respect to the extent of within-versus between-host selection. Here we use a detailed and semi-realistic multi-level HIV-1 model in which immunological pre-adaptation and SPVL evolution can emerge from the underlying interactions of the virus with the immune system of the host. This enables us to study the effect of immunological escape on disease progression, and how disease progression may be molded by SPVL evolution. We find that the time to AIDS could decrease significantly (0.5-1.0 years) in a HLA-dependent manner by immunological pre-adaptation over the long-term course of the epidemic (> 100 years). We find that SPVL is not expected to evolve to optimize the population-level fitness of HIV-1, even though high heritability of the SPVL emerges from continual selection of immune-escape mutations.
Inferring Disease-Associated Piwi-Interacting RNAs via Graph Attention Networks
Authors: Zheng Kai , You Zhu-Hong , Wang Lei , Wong Leon , Chen Zhan-Heng , Jiang Han-Jing
Published: January 2020

Motivation

PIWI proteins and Piwi-Interacting RNAs (piRNAs) are commonly detected in human cancers, especially in germline and somatic tissues, and correlates with poorer clinical outcomes, suggesting that they play a functional role in cancer. As the problem of combinatorial explosions between ncRNA and disease exposes out gradually, new bioinformatics methods for large-scale identification and prioritization of potential associations are therefore of interest. However, in the real world, the network of interactions between molecules is enormously intricate and noisy, which poses a problem for efficient graph mining. This study aims to make preliminary attempts on bionetwork based graph mining.

Results

In this study, we present a method based on graph attention network to identify potential and biologically significant piRNA-disease associations (PDAs), called GAPDA. The attention mechanism can calculate a hidden representation of an association in the network based on neighbor nodes and assign weights to the input to make decisions. In particular, we introduced the attention-based Graph Neural Networks to the field of bio-association prediction for the first time, and proposed an abstract network topology suitable for small samples. Specifically, we combined piRNA sequence information and disease semantic similarity with piRNA-disease association network to construct a new attribute network. In the experiment, GAPDA performed excellently in five-fold cross-validation with the AUC of 0.9038. Not only that, but it still has superior performance compared to methods based on collaborative filtering and attribute features. The experimental results show that GAPDA ensures the prospect of the graph neural network on such problems and can be an excellent supplement for future biomedical research.

Contact

zhuhongyou@ms.xjb.ac.cn;leiwang@ms.xjb.ac.cn

Supplementary information

Supplementary data are available at Bioinformatics online.
Convergent Met and voltage-gated Ca2+channel signaling on Ras-Erk MAPK drives migratory activation of dendritic cells parasitized byToxoplasma gondii
Authors: Ólafsson Einar B. , ten Hoeve Arne L. , Wang Xiaoze Li , Westermark Linda , Varas-Godoy Manuel , Barragan Antonio
Published: January 2020

Ras-Erk MAPK signaling controls many of the principal pathways involved in metazoan cell motility, drives metastasis of multiple cancer types and is targeted in chemotherapy. Yet, its putative roles in immune cell functions or in infections have remained elusive. Here, using primary dendritic cells (DCs) in an infection model with the protozoanToxoplasma gondii, we show that two pathways activated by infection converge on Ras-Erk MAPK signaling to promote migration of parasitized DCs. We identify signaling through the receptor tyrosine kinase Met (also known as HGFR) as a driver ofT. gondii-induced DC hypermotility. Further, we show that voltage-gated Ca2+channel (VGCC, subtype CaV1.3) signaling impacts the migratory activation of DCs via calmodulin-calmodulin kinase II. We report that VGCC and Met signaling converge on Ras GTPase to drive Erk1/2 phosphorylation and migratory activation ofT. gondii-infected DCs. The data provide a molecular basis for the hypermigratory mesenchymal-to-amoeboid transition (MAT) of parasitized DCs. The emerging concept suggests that parasitized DCs acquire metastasis-like migratory properties to promote infection-related dissemination.
Does seed mass drive interspecies variation in the effect of management practices on weed demography?
Authors: Elena Kazakou , Guillaume Fried , Pierre-Olivier Cheptou , Olivier Gimenez
Published: January 2020

Optimizing the effect of management practices on weed population dynamics is challenging due to the difficulties in inferring demographic parameters in seed banks and their response to disturbance. Here, we used a long-term plant survey between 2006 and 2012 in 46 French vineyards and quantified the effects of management practices (tillage, mowing and herbicide) on colonization, germination and seed survival of 30 weed species in relation to their seed mass. To do so, we used a recent statistical approach to reliably estimate demographic parameters for plant populations with a seed bank using time series of presence–absence data, which we extended to account for interspecies variation in the effects of management practices on demographic parameters. Our main finding was that when the level of disturbance increased (i.e., in plots with a higher number of herbicide, tillage or mowing treatments), colonization and survival in large-seeded species increased faster than in small-seeded species. High disturbance through tillage increased survival in the seed bank of species with high seed mass. The application of herbicides, considered as an intermediate disturbance, increased germination, survival and colonization probabilities of species with high seed mass. Mowing, representing habitats more competitive for light, increased the survival of species with high seed mass. Overall, the strong relationships between the effects of management practices and seed mass provides an indicator for predicting the dynamics of weed communities under disturbance.
Microbial diversity characterization of seawater in a pilot study using Oxford Nanopore Technologies long-read sequencing
Authors: Liem M. , Regensburg-Tuïnk A.J.G. , Henkel C.V. , Spaink H.P.
Published: January 2020

Currently the majority of non-culturable microbes in sea water are yet to be discovered, Nanopore offers a solution to overcome the challenging tasks to identify the genomes and complex composition of oceanic microbiomes. In this study we evaluate the utility of Oxford Nanopore Technologies (ONT) sequencing to characterize microbial diversity in seawater from multiple locations. We compared the microbial species diversity of retrieved environmental samples from two different locations and time points. With only three ONT flow cells we were able to identify thousands of organisms, including bacteriophages, from which a large part at species level. It was possible to assemble genomes from environmental samples with Flye. In several cases this resulted in >1 Mbp contigs and in the particular case of a Thioglobus singularis species it even produced a near complete genome. k-mer analysis reveals that a large part of the data represents species of which close relatives have not yet been deposited to the database. These results show that our approach is suitable for scalable genomic investigations such as monitoring oceanic biodiversity and provides a new platform for education in biodiversity
Dissecting the roles of GRK2 and GRK3 in μ-opioid receptor internalization and β-arrestin2 recruitment using CRISPR/Cas9-edited HEK293 cells
Authors: Møller Thor C. , Pedersen Mie F. , van Senten Jeffrey R. , Seiersen Sofie D. , Mathiesen Jesper M. , Bouvier Michel , Bräuner-Osborne Hans
Published: January 2020

Most G protein-coupled receptors (GPCRs) recruit β-arrestins and internalize upon agonist stimulation. For the μ-opioid receptor (μ-OR), this process has been linked to development of opioid tolerance. GPCR kinases (GRKs), particularly GRK2 and GRK3, have been shown to be important for μ-OR recruitment of β-arrestin and internalization. However, the contribution of GRK2 and GRK3 to β-arrestin recruitment and receptor internalization, remain to be determined in their complete absence. Using CRISPR/Cas9-mediated genome editing we established HEK293 cells with knockout of GRK2, GRK3 or both to dissect their individual contributions in β-arrestin2 recruitment and μ-OR internalization upon stimulation with four different agonists. We showed that GRK2/3 removal reduced agonist-induced μ-OR internalization and β-arrestin2 recruitment substantially and we found GRK2 to be more important for these processes than GRK3. Furthermore, we observed a sustained and GRK2/3 independent component of β-arrestin2 recruitment to the plasma membrane upon μ-OR activation. Rescue expression experiments restored GRK2/3 functions. Inhibition of GRK2/3 using the small molecule inhibitor CMPD101 showed a high similarity between the genetic and pharmacological approaches, cross-validating the specificity of both. However, off-target effects were observed at high CMPD101 concentrations. These GRK2/3 KO cell lines should prove useful for a wide range of studies on GPCR function.
Abdominal symptoms during primary Sjögren’s syndrome: a prospective study
Authors: Parreau Simon , Jacques Jérémie , Dumonteil Stéphanie , Palat Sylvain. , Geyl Sophie , Gondran Guillaume , Bezanahary Holy , Liozon Eric , Azaïs Julie , Colombie Stéphanie , Jauberteau Marie-Odile , Loustaud-Ratti Véronique , Ly Kim-Heang , Fauchais Anne-Laure
Published: January 2020

Context

Abdominal symptoms are poorly documented during primary Sjögren’s syndrome (pSS).

Objectives

To describe abdominal symptoms among pSS patients and to assess their association with characteristics of the disease.

Methods

One hundred and fifty patients followed at Hospital and University Center of Limoges were prospectively included and were evaluated using a composite global symptom score (GSS) describing abdominal symptoms and their severity. Data concerning the clinical and biological characteristics of the pSS and abdominal disorders were also collected.

Results

Ninety-five per cent of pSS patients suffered from abdominal symptoms with a median GSS of 7.5±5.5 points out of 30. More than half of the patients experienced abdominal tension (68%), upper abdominal pain (54%), abdominal discomfort (58%) and/or constipation (54%). Regarding the pSS activity ESSDAI score items, general and central nervous system involvement was associated with a high GSS. Regarding the patients’ symptoms ESSPRI score, there was a positive correlation with the GSS (p<0.01). Multivariate analysis showed a statistical association between a high GSS and seronegative status for SSA, gastroparesis and ESSPRI score (p<0.01for each one).

Conclusion

This study revealed that more than 90% of pSS patients suffered from abdominal symptoms. There is currently no therapeutic recommendation because of the lack of specific study and comprehension of the physiopathological mechanisms involved.
NvPOU4/Brain3functions as a terminal selector gene in the nervous system of the cnidarianNematostella vectensis
Authors: Tournière Océane , Dolan David , Richards Gemma Sian , Sunagar Kartik , Columbus-Shenkar Yaara Y , Moran Yehu , Rentzsch Fabian
Published: January 2020

Terminal selectors are transcription factors that control the morphological, physiological and molecular features that characterize distinct cell types. Here we use expression analyses and a transgenic reporter line to show thatNvPOU4is expressed in post-mitotic cells that give rise to a diverse set of neural cell types in the sea anemoneNematostella vectensis. We generated a loss-of-function allele by CRISPR/Cas9 and used additional transgenic reporter lines to show that the initial specification of neural cells is not affected in theNvPOU4mutants. Analyses of transcriptomes derived from the mutants and from different neural cell populations revealed thatNvPOU4is required for the execution of the terminal differentiation program of these neural cells. These findings suggest that POU4 genes have ancient functions as terminal selectors for morphologically and functionally highly disparate types of neurons and they provide experimental support for the relevance of terminal selectors for understanding the evolution of cell types.
A new long-read RNA-seq analysis approach identifies and quantifies novel transcripts of very large genes
Authors: Uapinyoying Prech , Goecks Jeremy , Knoblach Susan M. , Panchapakesan Karuna , Bonnemann Carsten G , Partridge Terence A. , Jaiswal Jyoti K , Hoffman Eric P.
Published: January 2020

RNA-seq is widely used for studying gene expression, but commonly used sequencing platforms produce short reads that only span up to two exon-junctions per read. This makes it difficult to accurately determine the composition and phasing of exons within transcripts. Although long-read sequencing improves this issue, it is not amenable to precise quantitation, which limits its utility for differential expression studies. We used long-read isoform sequencing combined with a novel analysis approach to compare alternative splicing of large, repetitive structural genes in muscles. Analysis of muscle structural genes that produce medium (Nrap−5kb), large (nebulin - 22 kb) and very-large (titin - 106 kb) transcripts in cardiac muscle, and fast and slow skeletal muscles identified unannotated exons for each of these ubiquitous muscle genes. This also identified differential exon usage and phasing for these genes between the different muscle types. By mapping the in-phase transcript structures to known annotations, we also identified and quantified previously unannotated transcripts. Results were confirmed by endpoint PCR and Sanger sequencing, which revealed muscle-type specific differential expression of these novel transcripts. The improved transcript identification and quantification demonstrated by our approach removes previous impediments to studies aimed at quantitative differential expression of ultra-long transcripts.
cGAS, a DNA Sensor, Promotes Inflammatory Responses in Huntington Disease
Authors: Sharma Manish , Rajendrarao Sumitha , Shahani Neelam , Ramĺrez-Jarquĺn Uri Nimrod , Subramaniam Srinivasa
Published: January 2020

The genetic cause of Huntington disease (HD) is attributed to the N-terminal polyglutamine expansion of huntingtin (mHTT). mHTT, which is a ubiquitously expressed protein, induces noticeable damage to the striatum, which affects motor, psychiatric, and cognitive functions in HD individuals. Although inflammatory responses apparently precede striatal damage and an overall progression of HD, the molecular mechanisms at work remain unclear (1-6). In this study, we found that cyclic GMP-AMP synthase (cGAS), a DNA sensor, which regulates inflammation, autophagy, and cellular senescence (7-9), plays a critical role in the inflammatory responses of HD. Ribosome profiling analysis reveals thatcGAS mRNAhas a high ribosome occupancy at exon 1 and codon-specific pauses at positions 171 (CCG) and 172 (CGT) in HD cells, compared to the control, indicating an altered cGAS expression. Accordingly, cGAS protein levels and activity, as measured by phosphorylation of stimulator of interferon genes (STING) or TANK-binding kinase 1 (TBK1), are increased in HD striatal cells, mouse Q175HD striatum and human postmortem HD striatum, compared to the healthy control. Furthermore, cGAS-dependent inflammatory genes such asCxcl10andCcl5show enhanced ribosome occupancy at exon 3 and exon 1, respectively and are upregulated in HD cells. Depletion of cGAS via CRISPR/Cas-9 diminishes cGAS activity and decreases expression of inflammatory genes while suppressing the autophagy upregulation in HD cells. We additionally detected the presence of numerous micronuclei, a known inducer of cGAS, in the cytoplasm of HD cells. Overall, the data indicates that cGAS is highly upregulated in HD and mediates inflammatory and autophagy responses. Thus, targeting cGAS may offer therapeutic benefits in HD.