bluuebunny/crossref_metadata_2025 huggingface dataset:
KinasePhos 3.0: Redesign and Expansion of the Prediction on Kinase-specific Phosphorylation Sites
Authors: Ma Renfei , Li Shangfu , Li Wenshuo , Yao Lantian , Huang Hsien-Da , Lee Tzong-Yi
Published: November 2021

The purpose of this work is to enhance KinasePhos, a machine-learning-based kinase-specific phosphorylation site prediction tool. Experimentally verified kinase-specific phosphorylation data were collected from PhosphoSitePlus, UniProt, GPS 5.0, and Phospho.ELM. In total, 41,421 experimentally verified kinase-specific phosphorylation sites were identified. A total of 1380 unique kinases were identified, including 753 with existing classification information from KinBase and the remaining 627 annotated by building a phylogenetic tree. Based on this kinase classification, a total of 771 predictive models were built at the individual, family, and group levels, using at least 15 experimentally verified substrate sites in positive training datasets. The improved models were observed to be more effective than other prediction tools. For example, the prediction of sites phosphorylated by the Akt, CKT, and PKA families had accuracies of 94.5%, 92.5%, and 90.0%, respectively. The average prediction accuracy for all 771 models was 87.2%. For enhancing interpretability, the Shapley additive explanations (SHAP) method was employed to assess feature importance. The web interface of KinasePhos 3.0 has been redesigned with the goal of providing comprehensive annotations of kinase-specific phosphorylation sites on multiple proteins. Additionally, considering the large scale of phosphoproteomic data, a downloadable prediction tool is available at https://awi.cuhk.edu.cn/KinasePhos/index.html or https://github.com/tom-209/KinasePhos-3.0-executable-file.
NASP maintains histone H3–H4 homeostasis through two distinct H3 binding modes
Authors: Bao Hongyu , Carraro Massimo , Flury Valentin , Liu Yanhong , Luo Min , Chen Liu , Groth Anja , Huang Hongda
Published: November 2021

Histone chaperones regulate all aspects of histone metabolism. NASP is a major histone chaperone for H3–H4 dimers critical for preventing histone degradation. Here, we identify two distinct histone binding modes of NASP and reveal how they cooperate to ensure histone H3–H4 supply. We determine the structures of a sNASP dimer, a complex of sNASP with an H3 α3 peptide, and the sNASP–H3–H4–ASF1b co-chaperone complex. This captures distinct functionalities of NASP and identifies two distinct binding modes involving the H3 α3 helix and the H3 αN region, respectively. Functional studies demonstrate the H3 αN-interaction represents the major binding mode of NASP in cells and shielding of the H3 αN region by NASP is essential in maintaining the H3–H4 histone soluble pool. In conclusion, our studies uncover the molecular basis of NASP as a major H3–H4 chaperone in guarding histone homeostasis.
Inhibition of BET family proteins suppresses African swine fever virus infection
Authors: Zhao Yaru , Niu Qingli , Yang Saixia , Yang Jifei , Zhang Zhonghui , Geng Shuxian , Fan Jie , Liu Zhijie , Guan Guiquan , Liu Zhiqing , Zhou Jia , Hu Haitao , Luo Jianxun , Yin Hong
Published: November 2021

African swine fever (ASF), an acute, severe, highly contagious disease caused by African swine fever virus (ASFV) infection in domestic pigs and boars, has a mortality rate of up to 100%. Because effective vaccines and treatments for ASF are lacking, effective control of the spread of ASF remains a great challenge for the pig industry. Host epigenetic regulation is essential for the viral gene transcription. Bromodomain and extraterminal (BET) family proteins, including BRD2, BRD3, BRD4, and BRDT, are epigenetic “readers” critical for gene transcription regulation. Among these proteins, BRD4 recognizes acetylated histones via its two bromodomains (BD1 and BD2) and recruits transcription factors, thereby playing a pivotal role in transcriptional regulation and chromatin remodeling during viral infection. However, how BET/BRD4 regulates ASFV replication and gene transcription is unknown. Here, we randomly selected 12 representative BET family inhibitors and compared their effects on ASFV infection in pig’s primary alveolar macrophages (PAMs). They were found to inhibit viral infection by interfering with the different stages of viral life cycle (attachment, internalization, desencapsidation and formation of viral factories). The four most effective inhibitors (ARV-825, ZL0580, I-BET-762 and PLX51107) were selected for further antiviral activity analysis. These BET/BRD4 inhibitors dose-dependently decreased the ASFV titer, viral RNA transcription and protein production in PAMs. Collectively,our study reported novel activity of BET/BRD4 inhibitors in inducing suppression of ASFV infection, providing insights into role of BET/BRD4 in epigenetic regulation of ASFV and potential new strategies for ASF prevention and control.

IMPORTANCE

Since the continuing spread of the ASFV in the world, and lack of commercial vaccines, the development of improved control strategies including antiviral drugs are urgently needed. BRD4 is an important epigenetic factor and has been commonly used for drug development for tumor treatment. Furthermore, the latest research showed that BET/BRD4 inhibition could suppress replication of virus. In this study, we first showed the inhibitory effect of agents targeting BET/BRD4 on ASFV infection with no significant host cytotoxicity. Then, we found 4 BET/BRD4 inhibitors which can inhibit ASFV replication, RNA transcription and protein synthesis. Finally, we analyzed 4 inhibitors’ biological effect on BRD4 according to the structure of BRD4, and docking analysis of BET-762, PLX51107, ARV-825 and ZL0580 binding to BD1 and BD2 domains of BRD4 was performed. Our findings support the hypothesis that BET/BRD4 can be considered as attractive host targets in antiviral drug discovery against ASFV.
Dense granule protein, GRA64 interacts with host cell ESCRT proteins duringToxoplasma gondiiinfection
Authors: Mayoral Joshua , Guevara Rebekah B. , Rivera-Cuevas Yolanda , Tu Vincent , Tomita Tadakimi , Romano Julia , Gunther-Cummins Leslie , Sidoli Simone , Coppens Isabelle , Carruthers Vernon B. , Weiss Louis M.
Published: November 2021

The intracellular parasiteToxoplasma gondiiadapts to diverse host cell environments within a replicative compartment that is heavily decorated by secreted proteins. In attempts to identify novel parasite secreted proteins that influence host cell activity, we identified and characterized a trans-membrane dense granule protein dubbed GRA64 (TGME49_202620). We found that GRA64 is on the parasitophorous vacuolar membrane (PVM) and is partially exposed to the host cell cytoplasm in both tachyzoite and bradyzoite parasitophorous vacuoles. Using co-immunoprecipitation and proximity-based biotinylation approaches, we demonstrate that GRA64 appears to interact with certain components of the host Endosomal Sorting Complexes Required for Transport (ESCRT). Genetic disruption of GRA64 does not affect acuteToxoplasmavirulence in mice nor encystation as observed via tissue cyst burdens in mice during chronic infection. However, ultrastructural analysis of Δgra64tissue cysts using electron tomography revealed enlarged vesicular structures underneath the cyst membrane, suggesting a role for GRA64 in organizing the recruitment of ESCRT proteins and subsequent intracystic vesicle formation. This study uncovers a novel host-parasite interaction that contributes to an emerging paradigm in which specific host ESCRT proteins are recruited to the limiting membranes (PVMs) of tachyzoite and bradyzoite vacuoles formed during acute and chronicToxoplasmainfection.

IMPORTANCE

Toxoplasma gondiiis a widespread foodborne parasite that causes congenital disease and life-threatening complications in immune compromised individuals. Part of this parasite’s success lies in its ability to infect diverse organisms and host cells, as well as to persist as a latent infection within parasite constructed structures called tissue cysts. In this study, we characterized a protein secreted byT. gondiiinto its parasitophorous vacuole during intracellular infection, which we dub GRA64. On the vacuole, this protein is exposed to the host cell and interacts with specific host ESCRT proteins. Parasites lacking the GRA64 protein exhibit ultrastructural changes in tissue cysts during chronic infection. This study lays the foundation for future studies on the mechanics and consequences of host ESCRT-parasite protein interactions.
Metabolome-defined obesity and the risk of future diabetes and mortality
Authors: Ottosson Filip , Smith Einar , Ericson Ulrika , Di Somma Salvatore , Antonini Paola , Nilsson Peter M , Fernandez Céline , Melander Olle
Published: November 2021

Background

Obesity is a key risk factor for type 2 diabetes, however, up to 20% of patients are normal weight. Our aim was to identify metabolite patterns reproducibly predictive of BMI, and subsequently to test if lean individuals who carry an obese metabolome are at hidden high risk of obesity related diseases, such as diabetes.

Methods

We measured 109 metabolites in fasted plasma samples of 7663 individuals from two Swedish and one Italian population-based cohort. Ridge regression models were used to predict BMI using the plasma metabolites. Individuals with a predicted BMI either more than 5 kg/m2higher (overestimated) or lower (underestimated) than their actual BMI were characterized as outliers and further investigated for obesity related risk factors and future risk of diabetes and mortality.

Results

The plasma metabolome could predict BMI in all cohorts (r2= 0.48, 0.26 and 0.19). The overestimated group had a BMI similar to individuals correctly predicted as normal weight, similar waist circumference, were not more likely to change weight over time but had a 2 times higher risk of future diabetes and an 80 % increased risk of all-cause mortality. These associations remained after adjustments for obesity-related risk factors and lifestyle parameters.

Conclusions

We found that lean individuals with an obese metabolome, have an increased risk for diabetes and all-cause mortality compared to lean individuals with a healthy metabolome. Metabolomics may be used to identify hidden high-risk individuals, in order to initiate lifestyle and pharmacological interventions.
Phase-locked transcranial electrical brain stimulation for tremor suppression in dystonic tremor syndromes
Authors: Nieuwhof Freek , Toni Ivan , Buijink Arthur W.G. , van Rootselaar Anne-Fleur , van de Warrenburg Bart P.C. , Helmich Rick C.
Published: November 2021

Background

Tremor is a common and burdensome symptom in patients with dystonia, which is clinically heterogeneous and often resistant to treatment. The pathophysiology is suggested to involve abnormal activity in the cerebellum and motor cortex, but the causal role of these brain regions remains to be established. Transcranial alternating current stimulaton (TACS) can suppress rhytmic cerebral activity in other tremor disorders when phase-locked to the ongoing arm tremor, but the effect on dystonic tremor syndromes is unknown.

Objective/Hypothesis

We aimed to establish the causal role of the cerebellum and motor cortex in dystonic tremor syndromes, and explore the therapeutic efficacy of phase-locked TACS.

Methods

We applied phase-locked TACS over the ipsilateral cerebellum (N=14) and contralateral motor cortex (N=17) in dystonic tremor syndrome patients, while patients assumed a tremor-evoking posture. We measured tremor power using accelerometery during 30s stimulation periods at 10 different phase-lags (36-degrees increments) between tremor and TACS for each target. Post-hoc, TACS-effects were related to a key clinical feature: the jerkiness (regularity) of tremor.

Results

Cerebellar TACS modulated tremor amplitude in a phase-dependent manner, such that tremor amplitude was suppressed or enhanced at opposite sides of the phase-cycle. This effect was specific for patients with non-jerky (sinusoidal) tremor (n=10), but absent in patients with jerky (irregular) tremor (n=4). Phase-locked stimulation over the motor cortex did not modulate tremor amplitude.

Conclusions

This study indicates that the cerebellum plays a causal role in the generation of (non-jerky) dystonic tremor syndrome. Our findings suggest pathophysiologic heterogeneity between patients with dystonic tremor syndrome, which mirrors clinical variability.
Fidaxomicin as first line: What will it cost in the USA and Canada?
Authors: Patel Devangi , Senecal Julien , Spellberg Brad , Morris Andrew M. , Saxinger Lynora , Footer Brent W. , McDonald Emily G. , Lee Todd C.
Published: November 2021

Importance

Recent changes in the Infectious Diseases and Healthcare Epidemiology Societies of America (IDSA-SHEA) guidelines for managing Clostridioides difficile infections (CDI) have placed fidaxomicin as first-line treatment.

Objective

To estimate the net cost of first line fidaxomicin compared to vancomycin in the American and Canadian healthcare systems and to estimate the price points at which fidaxomicin would become cost saving. In Canada, net costs were from the public payer perspective. In the US, costs were from a healthcare and payer perspective.

Data sources

We identified all randomized controlled trials comparing fidaxomicin with vancomycin through the 2021 IDSA-SHEA guideline update. Medication costs were obtained from internet prices (US) and the Quebec drug formulary (Canada). The average cost of a CDI recurrence was established through a systematic review for each country.

Study selection

For fidaxomicin efficacy, we included double-blind and placebo-controlled trials. For the systematic review of recurrence costs, studies were included if they were primary research articles, had a cost-analysis of CDI, included cases of recurrent CDI, and were calculated with cost parameters from American or Canadian healthcare systems. Studies were excluded if the population was solely pediatric or hospitalized.

Data extraction and Synthesis

For the efficacy meta-analysis, data was pooled using a restricted maximal likelihood random effects model. For the cost review, the mean cost across identified studies was adjusted to reflect July 2022 dollars.

Main Outcomes and Measures

The primary outcome of the meta-analysis was CDI recurrence at Day 40. The primary outcome of the systematic review was the average cost of a CDI recurrence in the American and Canadian healthcare systems. The objective was to estimate the net cost per recurrence prevented and the price point below which fidaxomicin would be cost saving to either the public payer (Canada) or the insurer (US).

Results

The estimated mean system cost of a CDI recurrence was $15147USD and $8806CAD, respectively. At current drug pricing, to prevent one recurrence by using first line fidaxomicin over vancomycin would cost $43904USD (95%CI $35123-$65856) and $13,760CAD (95%CI $11,008-$20,640), respectively. The likelihood of fidaxomicin offering cost savings varies by country, with a 95% probability of fidaxomicin being cost saving if priced below $1180USD or $860CAD respectively.

Conclusions and Relevance

An increased drug expenditure on fidaxomicin will not be offset through recurrence prevention unless the fidaxomicin price is negotiated.
Structural insights into human excitatory amino acid transporter EAAT2
Authors: Kato Takafumi , Kusakizako Tsukasa , Jin Chunhuan , Quan LiLi , Ohgaki Ryuichi , Okuda Suguru , Kobayashi Kan , Yamashita Keitaro , Nishizawa Tomohiro , Kanai Yoshikatsu , Nureki Osamu
Published: November 2021

Glutamate is a pivotal excitatory neurotransmitter in mammalian brains, but excessive glutamate causes numerous neural disorders. Almost all extracellular glutamate is retrieved by the glial transporter, Excitatory Amino Acid Transporter 2 (EAAT2), belonging to the SLC1A family. However, in some cancers, EAAT2 expression is enhanced and causes resistance to therapies by metabolic disturbance. Despite its crucial roles, the detailed structural information about EAAT2 has not been available. Here, we report cryo-EM structures of human EAAT2 in substrate-free and selective inhibitor WAY213613-bound states. EAAT2 forms a trimer, with each protomer consisting of transport and scaffold domains. Along with a glutamate-binding site, the transport domain possesses a cavity, that could be disrupted during the transport cycle. WAY213613 occupies both the glutamate-binding site and cavity of EAAT2 to interfere with its alternating access, where the sensitivity is defined by the inner environment of the cavity. This is the first characterization of molecular features of EAAT2 and the selective inhibition mechanism, underlying structure-based drug design for EAAT2.
Mis6/CENP-I maintains CENP-A nucleosomes against centromeric non-coding transcription during mitosis
Authors: Hirai Hayato , Shogaki Yuki , Sato Masamitsu
Published: November 2021

Centromeres are established by nucleosomes containing the histone H3 variant CENP-A. CENP-A is recruited to centromeres by the Mis18–HJURP machinery. During mitosis, CENP-A recruitment ceases, implying the necessity of CENP-A maintenance at centromeres, although the exact underlying mechanism remains elusive. Herein, we show that the kinetochore protein Mis6 (CENP-I) retains CENP-A during mitosis in fission yeast. Eliminating Mis6 during mitosis caused immediate loss of pre-existing CENP-A at centromeres. CENP-A loss occurred due to the transcriptional upregulation of non-coding RNAs at the central core region of centromeres, as confirmed by the observation RNA polymerase II inhibition preventing CENP-A loss from centromeres in the mis6 mutant. Thus, we concluded that Mis6 blocks the indiscriminate transcription of non-coding RNAs at the core centromere, thereby retaining the epigenetic inheritance of CENP-A during mitosis.
Visual category representations in the infant brain
Authors: Xie Siying , Hoehl Stefanie , Moeskops Merle , Kayhan Ezgi , Kliesch Christian , Turtleton Bert , Köster Moritz , Cichy Radoslaw M.
Published: November 2021

Visual categorization is a human core cognitive capacity1,2that depends on the development of visual category representations in the infant brain3–7. However, the exact nature of infant visual category representations and their relationship to the corresponding adult form remains unknown8. Our results clarify the nature of visual category representations from electroencephalography (EEG) data in 6- to 8-month-old infants and their developmental trajectory towards adult maturity in the key characteristics of temporal dynamics2,9, representational format10–12, and spectral properties13,14. Temporal dynamics change from slowly emerging, developing representations in infants to quickly emerging, complex representations in adults. Despite those differences, infants and adults already partly share visual category representations. The format of infants’ representations is visual features of low to intermediate complexity, whereas adults’ representations also encode high complexity features. Theta band activity contributes to visual category representations in infants, and these representations are shifted to the alpha/beta band in adults. Together, we reveal the developmental neural basis of visual categorization in humans, show how information transmission channels change in development, and demonstrate the power of advanced multivariate analysis techniques in infant EEG research for theory building in developmental cognitive science.
Wnt1-Cre mediated deletion of BMP7 suggests a role for neural crest-derived BMP7 in retina development and function
Authors: Kung Tiffany FC , Baddam Pranidhi , Liu Ruocun , Maripuri Devi Priyanka , Dimopoulos Ioannis S , MacDonald Ian M , Sauve Yves , Graf Daniel
Published: November 2021

Neural crest (NC) contributes to various structures of the eye including cornea, ciliary body and retina. The association of NC-derived cells with hyaloid vessels in the form of pericytes is established. Similarly, persistence of NC-derived cells in the inner retina layer of the mature retina has been suggested. To date, no specific function has been attributed to them. NC-derived Bone morphogenetic protein 7 (BMP7) controls neurogenic properties in the brain and regulates glia differentiation. Here, we assessed the role of NC-derived BMP7 in the adult retina.

BMP7 expression was determined using Bmp7LacZ reporter mice. BMP7 was expressed in GCL, IPL, OPL, and photoreceptors in P0, P14 and P30 retinas. Lineage tracing confirmed the presence of NC-derived cells in the GCL, INL, and ONL. Some but not all cells associated with vasculature. To test the function of NC-derived Bmp7, Bmp7fl/flWnt1cre (Bmp7ncko) mice were assessed by histological and functional methods. Loss of NC-derived cells in the GCL and INL and mild structural abnormalities were observed in the Bmp7ncko retina. Electroretinography revealed reduced a wave under photopic conditions and b wave under both scotopic and photopic conditions. The neuronal circuitry in the inner retina appeared affected, evidenced by decreased Calbindin in the GCL, IPL and INL. In the outer retina, S-opsin was increased. BMP7 expression in the mutant retina was strongly decreased at birth, but increased expression from cells other than NC was observed in the adult retina. This was associated with an increase in IBA1, suggestive that loss of NC-derived BMP7 predisposes to development of gliosis-like changes in the adult retina. Overall, our data reveal an important contribution of NC-derived BMP7 for the development and function of the inner and outer retina.
Biobank-scale inference of ancestral recombination graphs enables genealogy-based mixed model association of complex traits
Authors: Zhang Brian C. , Biddanda Arjun , Palamara Pier Francesco
Published: November 2021

Accurate inference of gene genealogies from genetic data has the potential to facilitate a wide range of analyses. We introduce a method for accurately inferring biobank-scale genome-wide genealogies from sequencing or genotyping array data, as well as strategies to utilize genealogies within linear mixed models to perform association and other complex trait analyses. We use these new methods to build genome-wide genealogies using genotyping data for 337,464 UK Biobank individuals and to detect associations in 7 complex traits. Genealogy-based association detects more rare and ultra-rare signals (N= 133, frequency range 0.0004% - 0.1%) than genotype imputation from ∼65,000 sequenced haplotypes (N= 65). In a subset of 138,039 exome sequencing samples, these associations strongly tag (averager= 0.72) underlying sequencing variants, which are enriched for missense (2.3×) and loss-of-function (4.5×) variation. Inferred genealogies also capture additional association signals in higher frequency variants. These results demonstrate that large-scale inference of gene genealogies may be leveraged in the analysis of complex traits, complementing approaches that require the availability of large, population-specific sequencing panels.
PGC-1α in the myofibers regulates the balance between myogenic and adipogenic progenitors affecting muscle regeneration
Authors: Beltrà Marc , Pin Fabrizio , Costamagna Domiziana , Duelen Robin , Renzini Alessandra , Ballarò Riccardo , Garcia-Castillo Lorena , Iannuzzi Ambra , Moresi Viviana , Coletti Dario , Sampaolesi Maurilio , Penna Fabio , Costelli Paola
Published: November 2021

Skeletal muscle repair is accomplished by satellite cells (MuSC) in cooperation with interstitial stromal cells (ISCs). So far, the relationship between the function of these cells and the metabolic state of myofibers remains unclear. The present study reports alterations in the proportion of both MuSCs and adipogenesis regulators (Aregs) induced by overexpression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) in the myofibers (MCK-PGC-1α mice). Although PGC-1α–driven increase of MuSCs does not accelerate muscle regeneration, myogenic progenitors isolated from MCK-PGC-1α mice and transplanted into intact and regenerating muscles are more prone to fuse with recipient myofibers than those derived from WT donors. Moreover, both young and aged MCK-PGC-1α animals show reduced perilipin-positive areas when challenged with an adipogenic stimulus, demonstrating low propensity to accumulate adipocytes within the muscle. These results provide new insights on the role played by PGC-1α in promoting myogenesis and hindering adipogenesis in the skeletal muscle.
Increasing the throughput of sensitive proteomics by plexDIA
Authors: Derks Jason , Leduc Andrew , Wallmann Georg , Huffman R. Gray , Willetts Matthew , Khan Saad , Specht Harrison , Ralser Markus , Demichev Vadim , Slavov Nikolai
Published: November 2021

Current mass-spectrometry methods enable high-throughput proteomics of large sample amounts, but proteomics of low sample amounts remains limited in depth and throughput. To increase the throughput of sensitive proteomics, we developed an experimental and computational framework, plexDIA, for simultaneously multiplexing the analysis of both peptides and samples. Multiplexed analysis with plexDIA increases throughput multiplicatively with the number of labels without reducing proteome coverage or quantitative accuracy. By using 3-plex nonisobaric mass tags, plexDIA enables quantifying 3-fold more protein ratios among nanogram-level samples. Using 1 hour active gradients and first-generation Q Exactive, plexDIA quantified about 8,000 proteins in each sample of labeled 3-plex sets. plexDIA also increases data completeness, reducing missing data over 2-fold across samples. We applied plexDIA to quantify proteome dynamics during the cell division cycle in cells isolated based on their DNA content; plexDIA detected many classical cell cycle proteins and discovered new ones. When applied to single human cells, plexDIA quantified about 1,000 proteins per cell and achieved 98 % data completeness within a plexDIA set while using about 5 min of active chromatography per cell. These results establish a general framework for increasing the throughput of sensitive and quantitative protein analysis.
A multi-species co-occurrence index to avoid type II errors in null model testing
Authors: Lagat Vitalis K. , Latombe Guillaume , Hui Cang
Published: November 2021

Community structure is determined by the interplay among different processes, including biotic interactions, abiotic filtering and dispersal. Their effects can be detected by comparing observed patterns of co-occurrence between different species (e.g. C-score and the natural metric) to patterns generated by null models based on permutations of species-by-site matrices under constraints on row or column sums. These comparisons enable us to detect significant signals of species association or dissociation, from which the type of biotic interactions between species (e.g. facilitative or antagonistic) can be inferred. Commonly used patterns are based on the levels of co-occurrence between randomly paired species. The level of co-occurrence for three or more species is rarely considered, ignoring the potential existence of functional guilds or motifs composed of multiple species within the community. Null model tests that do not consider multi-species co-occurrence could therefore generate false negatives (Type II error) in detecting non-random forces at play that would only be apparent for such guilds. Here, we propose a multi-species co-occurrence index (hereafter, joint occupancy) that measures the number of sites jointly occupied by multiple species simultaneously, of which the pairwise metric of co-occurrence is a special case. Using this joint occupancy index along with standard permutation algorithms for null model testing, we illustrate nine archetypes of multi-species co-occurrence and explore how frequent they are in the seminal database of 289 species-by-site community matrices published by Atmar and Patterson in 1995. We show that null model testing using pairwise co-occurrence metrics could indeed lead to severe Type II errors in one specific archetype, accounting for 2.4% of the tested community matrices.
A novel hybrid protein promotes Aβ clearance and reduces inflammatory response through MerTK
Authors: Samentar Lorena P. , Salazar Arnold , Pan Pei-Pei , Etebar Kayvon , Choy Kelly , Uddin Durin , Eliseeff Pauline , Bugayong Adrienne Marrie , Garrido Jose Antonio Ma. G. , Emini Aurora , Rock Nicole , Caberoy Nora Blanca
Published: November 2021

Alzheimer’s disease (AD) is the world’s leading cause of dementia and the most common neurodegenerative disorder. Its major pathological features are amyloid beta (Aβ) plaques, tau tangles, and neuroinflammation that eventually leads to massive death of nerve cells. Even with the multifactorial aspect of AD, the most accepted theory is that Aβ is the driving force of AD pathogenesis. We engineered a novel hybrid protein that facilitates the phagocytosis of Aβ and redirect its clearance to the noninflammatory Mer tyrosine kinase (MerTK) pathway. The novel hybrid protein facilitates robust uptake and clearance of Aβ in BV2 microglia through MerTK receptor with reduced production of inflammatory factors and oxidative products. In APP/PS1 transgenic AD mouse model, intraperitoneal administration of the hybrid protein for two months results in significant reduction of Aβ burden in the brain and protection of nerve cells from dying. Taken together, our results suggest that the novel hybrid may have the potential for AD treatment by targeting both Aβ clearance and reduction of inflammation.
Meta-Analysis of cortical inhibitory interneurons markers landscape and their performances in scRNA-seq studies
Authors: Martini Lorenzo , Bardini Roberta , Di Carlo Stefano
Published: November 2021

The mammalian cortex contains a great variety of neuronal cells. In particular, GABAergic interneurons, which play a major role in neuronal circuit function, exhibit an extraordinary diversity of cell types. In this regard, single-cell RNA-seq analysis is crucial to study cellular heterogeneity. To identify and analyze rare cell types, it is necessary to reliably label cells through known markers. In this way, all the related studies are dependent on the quality of the employed marker genes. Therefore, in this work, we investigate how a set of chosen inhibitory interneurons markers perform. The gene set consists of both immunohistochemistry-derived genes and single-cell RNA-seq taxonomy ones. We employed various human and mouse datasets of the brain cortex, consequently processed with the Monocle3 pipeline. We defined metrics based on the relations between unsupervised cluster results and the marker expression. Specifically, we calculated the specificity, the fraction of cells expressing, and some metrics derived from decision tree analysis like entropy gain and impurity reduction. The results highlighted the strong reliability of some markers but also the low quality of others. More interestingly, though, a correlation emerges between the general performances of the genes set and the experimental quality of the datasets. Therefore, the proposed method allows evaluating the quality of a dataset in relation to its reliability regarding the inhibitory interneurons cellular heterogeneity study.
Endothelial Rap1B mediates T-cell exclusion to promote tumor growth – a novel mechanism underlying vascular immunosuppression
Authors: Sharma Guru Prasad , Kosuru Ramoji , Lakshmikanthan Sribalaji , Zheng Shikan , Chen Yao , Burns Robert , Xin Gang , Cui Weiguo , Chrzanowska Magdalena
Published: November 2021

Overcoming vascular immunosuppression: lack of endothelial cell (EC) responsiveness to inflammatory stimuli in the proangiogenic environment of tumors, is essential for successful cancer immunotherapy. The mechanisms through which Vascular Endothelial Growth Factor (VEGF) modulates tumor EC response to exclude T cells are not well understood. The goal was to determine the role of EC Rap1B, a small GTPase that positively regulates VEGF- angiogenesis during development, in tumor growth in vivo. Using mouse models of Rap1B deficiency, Rap1B+/- and EC-specific Rap1B KO (Rap1BiΔEC) we demonstrate that EC Rap1B restricts tumor growth and angiogenesis. More importantly, EC-specific Rap1B deletion leads to an altered tumor microenvironment with increased recruitment of leukocytes and increased activity of tumor CD8+ T cells. We find that tumor growth, albeit not angiogenesis, is restored in Rap1BiΔEC mice by depleting CD8+ T cells. Mechanistically, global transcriptome analysis indicated upregulation of the tumor cytokine, TNF-α, -induced signaling and NFκB transcriptional activity in Rap1B-deficient ECs. Functionally, EC Rap1B deletion led to upregulation of NFκB activity and enhanced Cell Adhesion Molecules (CAMs) expression in TNF-α stimulated ECs. Importantly, CAM expression was upregulated also in tumor ECs from Rap1BiΔEC mice, vs. controls. Significantly, deletion of Rap1B abrogated VEGF immunosuppressive downregulation of CAM expression, demonstrating that Rap1B is essential for VEGF-suppressive signaling. Thus, our studies identify a novel endothelial-endogenous mechanism underlying VEGF-dependent desensitization of EC to pro-inflammatory stimuli. Significantly, they identify EC Rap1 as a potential novel vascular target in cancer immunotherapy.
Biofabrication of development-inspired scaffolds for regeneration of the annulus fibrosus macro- and microarchitecture
Authors: Kluser Nadine , Sprecher Christoph , Alig Gion Ursin , Häckel Sonja , Albers Christoph E. , Alini Mauro , Grad Sibylle , Eglin David , Vernengo Andrea
Published: November 2021

Annulus fibrosus (AF) tissue engineering is a promising strategy for repairing the degenerated intervertebral disc (IVD) and a research area that could benefit from improved tissue models to drive translation. AF tissue is composed of concentric layers of aligned collagen bundles arranged in an angle-ply pattern, an architecture which is challenging to recapitulate with current scaffold design strategies. In response to this need, we developed a strategy to print 3D scaffolds that induce cell and tissue organization into oriented patterns mimicking the AF. Polycaprolactone (PCL) was printed in an angle-ply macroarchitecture possessing microscale aligned topographical cues. The topography was achieved by extrusion through custom-designed printer nozzles which were either round or possessing circumferential sinusoidal peaks. Whereas the round nozzle produced extruded filaments with a slight uniaxial texture, patterned nozzles with peak heights of 60 or 120 μm produced grooves, 10.87 ± 3.09 μm or 17.77 ± 4.91 μm wide, respectively. Bone marrow derived mesenchymal stem cells (BM-MSCs) cultured on the scaffolds for four weeks exhibited similar degrees of alignment within ± 10 ° of the printing direction and upregulation of outer AF markers (COL1, COL12, SFRP, MKX, MCAM, SCX and TAGLN), with no statistically significant differences as a function of topography. Interestingly, the grooves generated by the patterned nozzles induced longitudinal end-to-end alignment of cells, capturing the arrangement of cells during fibrillogenesis. In contrast, topography produced from the round nozzle induced a continuous web of elongated cells without end-to-end alignment. Extracellular collagen I, decorin and fibromodulin were detected in patterns closely following cellular organization. Taken together, we present a single-step biofabrication strategy to induce anisotropic cellular alignments in x-, y-, and z-space, with potential application as anin vitromodel for studying AF tissue morphogenesis and growth.
Effects of chlorpyrifos on early development and anti-predator behavior of agile frogs
Authors: Mikó Zsanett , Bókony Veronika , Ujhegyi Nikolett , Nemesházi Edina , Erös Réka , Orf Stephanie , Hettyey Attila
Published: November 2021

The widespread application of pesticides makes it important to understand the impacts of these chemicals on wildlife populations. Chlorpyrifos is an organophosphate insecticide which can affect the development and behavior of aquatic organisms and may thereby alter predator-prey interactions. To investigate how environmentally relevant, sublethal concentrations of chlorpyrifos affect anti-predator behavior and larval development of the agile frog (Rana dalmatina), we exposed tadpoles to one of three concentrations (0, 0.5 and 5 μg chlorpyrifos / L) either for a brief period of three days (acute exposure) or throughout larval development (chronic exposure). We observed tadpole activity and space use in the presence or absence of chemical cues of predatory fish. We also assessed mortality, time to metamorphosis, mass at metamorphosis, brain morphology and sex ratio. We found that tadpoles chronically exposed to 5 μg/L chlorpyrifos swam distances that were longer by more than 20 % and exhibited body masses at metamorphosis that were lower by ca. 7 % than in control individuals, but the other fitness-related traits remained unaffected. The lower concentration of chlorpyrifos applied chronically, and either one of the acute chlorpyrifos treatments did not influence any measured trait. Our results demonstrate that exposure to chlorpyrifos can induce changes in locomotor activity and may result in lowered body mass of agile frog tadpoles, but only if the insecticide is present chronically at concentrations which are rarely reached in natural waterbodies. Thus, agile frog tadpoles appear to be relatively tolerant to chlorpyrifos, but may nonetheless suffer from its presence in situations of repeated high-dose application.
Singlet oxygen-induced signalling depends on the metabolic status of the Chlamydomonas cell
Authors: Al Youssef Waeil , Feil Regina , Saint-Sorny Maureen , Johnson Xenie , Lunn John E. , Grimm Bernhard , Brzezowski Pawel
Published: November 2021

Using a novel mutant screen, we identified TREHALOSE 6-PHOSPHATE PHOSPHATASE 1 (TSPP1) as a functional enzyme dephosphorylating trehalose 6-phosphate (Tre6P) to trehalose in Chlamydomonas reinhardtii. The tspp1 knock-out results in reprogramming of the cell metabolism via significantly altered transcriptome. As a secondary effect, tspp1 also shows impairment in 1O2-induced chloroplast retrograde signalling. From transcriptomic analysis and metabolite profiling, we conclude that accumulation or deficiency of certain metabolites directly affect 1O2-signalling. 1O2-inducible GLUTATHIONE PEROXIDASE 5 (GPX5) gene expression is suppressed by increased content of fumarate, an intermediate in the tricarboxylic acid cycle (TCA cycle) in mitochondria and dicarboxylate metabolism in the cytosol, while it is promoted by another TCA cycle intermediate, aconitate. Furthermore, genes encoding known essential components of chloroplast-to-nucleus 1O2-signalling, PSBP2, MBS, and SAK1, show decreased transcript levels in the tspp1 mutant, which can be rescued by exogenous application of aconitate. We demonstrate that chloroplast retrograde signalling involving 1O2 depends on mitochondrial and cytosolic processes and that the metabolic status of the cell determines the response to 1O2.
The promotion of stress tolerant Symbiodiniaceae dominance in juveniles of two coral species due to simulated future conditions of ocean warming and acidification
Authors: Terrell Alyx P. , Marangon Emma , Webster Nicole S. , Cooke Ira , Quigley Kate M.
Published: November 2021

The symbiotic relationship between coral and its endosymbiotic algae, Symbiodiniaceae, greatly influences the hosts’ potential to withstand environmental stress. To date, the effects of climate change on this relationship has primarily focused on adult corals. Uncovering the effects of environmental stress on the establishment and development of this symbiosis in early life stages is critical for predicting how corals may respond to climate change. To determine the impacts of future climate projections on the establishment of symbionts in juvenile corals, ITS2 amplicon sequencing of single coral juveniles was applied to Goniastrea retiformis and Acropora millepora before and after exposure to three climate conditions of varying temperature and pCO2 levels (current and RCP8.5 in 2050 and 2100). Compared to ambient conditions, juvenile corals experienced shuffling in the relative abundance of Cladocopium (C1m, reduction) to Durusdinium (D1 and D1a, increase) over time. We calculated a novel risk metric incorporating functional redundancy and likelihood of impact on host physiology to identify the loss of D1a as a ‘low risk’ to the coral compared to the loss of “higher risk” taxa like D1 and C1m. Although the increase in stress tolerant Durusdinium under future warming was encouraging for A. millepora, by 2100, G. retiformis communities displayed signs of symbiosis de-regulation, suggesting this acclimatory mechanism may have species-specific thresholds. These results emphasize the need for understanding of long-term effects of climate change induced stress on coral juveniles and their potential for increased acclimation to heat tolerance through changes in symbiosis.

Originality Statement

Here we assessed changes in the uptake and establishment of Symbiodiniaceae in the early lifehistory stages of two coral species under future climate scenarios. Our study represents the first such assessment of future climate change projections (increased temperature and pCO2) influencing Symbiodiniaceae acquisition and specifically shows a community structure dominated by the stress tolerant genus Durusdinium. We also develop a novel risk metric that includes taxonomic function and redundancy to estimate the impact of symbiont taxa changes on coral physiology. Through the risk metric, we relate the stress-induced changes in symbiont community structure to the likelihood of functional loss to better understand the extent to which these changes may lead to a decrease in coral health.
Transcription factor induction of vascular blood stem cell niches in vivo
Authors: Hagedorn Elliott J. , Perlin Julie R. , Freeman Rebecca J. , Wattrus Samuel J. , Han Tianxiao , Mao Clara , Kim Ji Wook , Fernández-Maestre Inés , Daily Madeleine L. , D’Amato Christopher , Fairchild Michael J. , Riquelme Raquel , Li Brian , Ragoonanan Dana A.V.E. , Enkhbayar Khaliun , Henault Emily L. , Wang Helen G. , Redfield Shelby E. , Collins Samantha H. , Lichtig Asher , Yang Song , Zhou Yi , Kunar Balvir , Gomez-Salinero Jesus Maria , Dinh Thanh T. , Pan Junliang , Holler Karoline , Feldman Henry A. , Butcher Eugene C. , van Oudenaarden Alexander , Rafii Shahin , Philipp Junker J. , Zon Leonard I.
Published: November 2021

The hematopoietic niche is a supportive microenvironment comprised of distinct cell types, including specialized vascular endothelial cells that directly interact with hematopoietic stem and progenitor cells (HSPCs). The molecular factors that specify niche endothelial cells and orchestrate HSPC homeostasis remain largely unknown. Using multi-dimensional gene expression and chromatin accessibility analyses, we define a conserved gene expression signature and cis-regulatory landscape unique to sinusoidal endothelial cells in the HSPC niche. Using enhancer mutagenesis and transcription factor overexpression, we elucidate a transcriptional code involving members of the Ets, Sox and Nuclear Hormone Receptor families that is sufficient to induce ectopic niche endothelial cells that associate with mesenchymal stromal cells and support the recruitment, maintenance and division of HSPCs in vivo. These studies set forth an approach for generating synthetic HSPC niches, in vitro or in vivo, and for effective therapies to modulate the endogenous niche.
Leishmania amazonensis sabotages host cell SUMOylation for intracellular survival
Authors: Okuda Kendi , Costa Franco Miriam Maria Silva , Yasunaga Ari , Gazzinelli Ricardo , Rabinovitch Michel , Cherry Sara , Silverman Neal
Published: November 2021

Leishmania parasites use elaborate virulence mechanisms to invade and thrive in macrophages. These virulence mechanisms inhibit host cell defense responses and generate a specialized replicative niche, the parasitophorous vacuole. In this work, we performed a genome-wide RNAi screen in Drosophila macrophage-like cells to identify host factors necessary for Leishmania amazonensis infection. This screen identified 52 conserved genes required specifically for parasite entry, including several components of the SUMOylation machinery. Further studies in mammalian macrophages found that L. amazonensis infection inhibited SUMOylation within infected macrophages and this inhibition enhanced parasitophorous vacuole growth and parasite proliferation through modulation of multiple genes especially ATP6V0D2, which in turn effects CD36 expression and cholesterol levels. Together, these data suggest that parasites actively sabotage host SUMOylation and alter host transcription to improve their intracellular niche and enhance their replication.
Reciprocal zebrafish-medaka hybrids reveal maternal control of zygotic genome activation timing
Authors: Gert Krista R. , Quio Luis Enrique Cabrera , Novatchkova Maria , Guo Yixuan , Cairns Bradley R. , Pauli Andrea
Published: November 2021

After fertilization, the sperm and egg contribute unequally to the newly formed zygote. While the sperm contributes mainly paternal DNA, the egg provides both maternal DNA and the bulk of the future embryonic cytoplasm. Most embryonic processes (like the onset of zygotic transcription) depend on maternally-provided cytoplasmic components, and it is largely unclear whether paternal components besides the centrosome play a role in the regulation of early embryogenesis. Here we report a reciprocal zebrafish-medaka hybrid system as a powerful tool to investigate paternal vs. maternal influence during early development. By combining expression of zebrafish Bouncer on the medaka egg with artificial egg activation, we demonstrate the in vitro generation of paternal zebrafish x maternal medaka (reripes) hybrids. These hybrids complement the previously established paternal medaka x maternal zebrafish (latio) hybrids (Herberg et al., 2018). As proof of concept, we investigated maternal vs. paternal control of zygotic genome activation (ZGA) timing using this reciprocal hybrid system. RNA-seq analysis of the purebred fish species and hybrids revealed that the onset of ZGA is primarily governed by the egg. Overall, our study establishes the reciprocal zebrafish-medaka hybrid system as a versatile tool to dissect parental control mechanisms during early development.
Adar1 deletion causes degeneration of exocrine pancreas via Mavs-dependent interferon signaling
Authors: Rupani Dhwani N. , Cowan Robert W. , Thege Fredrik I. , Chandra Vidhi , Wörmann Sonja M. , Rajaei Hajar , Malaney Prerna , Le Roux Olivereen , Manning Sara L. , Hashem Jack , Bailey-Lundberg Jennifer , McAllister Florencia , Rhim Andrew D.
Published: November 2021

Adenosine deaminase acting on RNA 1 (ADAR1) is an RNA-binding protein that deaminates adenosine(A) to inosine(I). A-to-I editing alters post-transcriptional RNA processing making ADAR1 a critical regulator of gene expression. Consequently, Adar1 has been implicated in organogenesis. To determine the role of Adar1 in pancreatic development and homeostasis, we specifically deleted Adar1 from the murine pancreas (Ptf1aCre/+; Adar1Fl/Fl). The resulting mice had stunted growth likely due to malabsorption associated with exocrine pancreas insufficiency. Analyses of pancreases revealed ductal expansion, heightened interferon-stimulated gene expression and an increased influx of immune cells. In addition, we observed an increased prevalence of CD4+ T and natural killer cells in their splenic tissue. These results indicate an association between loss of pancreatic Adar1 with dysregulation of systemic immunity. Concurrent deletion of Adar1 and Mavs, a signaling protein implicated in the innate immune pathway rescued the degenerative phenotype and resulted in normal pancreatic development. Taken together, our work suggests that the primary function of Adar1 in the pancreas is to prevent aberrant activation of the Mavs-mediated innate immune pathway, thereby maintaining pancreatic homeostasis.

Summary statement

This work defines the role of Adar1 in pancreatic development and homeostasis.