bluuebunny/crossref_metadata_2025 huggingface dataset:
An in silico genome-wide screen for circadian clock strength in human samples
Authors: Wu Gang , Ruben Marc D. , Francey Lauren J. , Lee Yin Yeng , Anafi Ron C. , Hogenesch John B.
Published: May 2022

Years of time-series gene expression studies have built a strong understanding of clock-controlled pathways across species. However, comparatively little is known about how ‘non-clock’ pathways influence clock function. We developed a new computational approach to explore candidate pathways coupled to the clock in human tissues. This method, termed LTM, is an in silico screen to infer genetic influences on circadian clock function. LTM uses natural variation in gene expression in human data and directly links gene expression variation to clock strength independent of longitudinal data. We applied LTM to three human skin and one melanoma datasets and found that the cell cycle is the top candidate clock-coupled pathway in healthy skin. In addition, we applied LTM to thousands of tumor samples from 11 cancer types in the TCGA database and found that extracellular matrix organization-related pathways are tightly associated with the clock strength in humans. Further analysis shows that clock strength in tumor samples are correlated with the proportion of cancer-associated fibroblasts and endothelial cells. Therefore, we show both the power of LTM in predicting clock-coupled pathways and classify factors associated with clock strength in human tissues. LTM is available on GitHub to facilitate its use.
Age-related reorganization of functional network architecture in semantic cognition
Authors: Martin Sandra , Williams Kathleen A. , Saur Dorothee , Hartwigsen Gesa
Published: May 2022

Semantic memory is a fundamental human ability which is central to communication. Although it is usually well preserved in healthy aging, memory problems in verbal communication due to slowed access and retrieval processes are a common complaint with increasing age. So far, the neural bases of this paradox remain poorly understood. The current neuroimaging study investigated age differences in the functional network architecture during semantic word retrieval in young and older adults. Using group spatial independent component analysis, we defined functional networks for verbal semantic fluency. Combining task-based functional connectivity, graph theory and cognitive measures of fluid and crystallized intelligence, our findings show age-accompanied large-scale network reorganization even when older adults have intact word retrieval abilities. In particular, functional networks of older adults were characterized by reduced decoupling between systems, reduced segregation and efficiency, and a larger number of hub regions relative to young adults. Exploring the predictive utility of these age-related changes in network topology revealed high, albeit less efficient, performance for older adults whose brain graphs showed stronger dedifferentiation and reduced distinctiveness. Our results extend theoretical accounts on neurocognitive aging by revealing the compensational potential of the commonly reported pattern of network dedifferentiation when older adults can rely on their prior knowledge for successful task processing. However, we also demonstrate the limitations of such compensatory reorganization processes and demonstrate that a youth-like network architecture in terms of balanced integration and segregation is associated with more economical processing.

Significance Statement

Cognitive aging is associated with widespread neural reorganization processes in the human brain. However, the behavioral impact of such reorganization is not well understood. Here, we used taskbased fMRI to demonstrate a large-scale reorganization of brain networks in older adults even when their semantic abilities are intact. In particular, functional networks of older adults were characterized by increased coupling between different systems, reduced segregation and efficiency, and a larger number of hub regions relative to young adults. Associating these changes with behavior revealed high, albeit less efficient, performance for networks in older adults showing stronger dedifferentiation and reduced distinctiveness. Our results highlight the compensatory potential of network reconfiguration with age, but also reveal the limitations of such reorganization processes.
Genetic architecture of the white matter connectome of the human brain
Authors: Sha Zhiqiang , Schijven Dick , Fisher Simon E. , Francks Clyde
Published: May 2022

White matter tracts form the structural basis of large-scale functional networks in the human brain. We applied brain-wide tractography to diffusion images from 30,810 adult participants (UK Biobank), and found significant heritability for 90 regional connectivity measures and 851 tract-wise connectivity measures. Multivariate genome-wide association analyses identified 355 independently associated lead SNPs across the genome, of which 77% had not been previously associated with human brain metrics. Enrichment analyses implicated neurodevelopmental processes including neurogenesis, neural differentiation, neural migration, neural projection guidance, and axon development, as well as prenatal brain expression especially in stem cells, astrocytes, microglia and neurons. We used the multivariate association profiles of lead SNPs to identify 26 genomic loci implicated in structural connectivity between core regions of the left-hemisphere language network, and also identified 6 loci associated with hemispheric left-right asymmetry of structural connectivity. Polygenic scores for schizophrenia, bipolar disorder, autism spectrum disorder, attention-deficit hyperactivity disorder, left-handedness, Alzheimer’s disease, amyotrophic lateral sclerosis, and epilepsy showed significant multivariate associations with structural connectivity, each implicating distinct sets of brain regions with trait-relevant functional profiles. This large-scale mapping study revealed common genetic contributions to the structural connectome of the human brain in the general adult population, highlighting links with polygenic disposition to brain disorders and behavioural traits.

One sentence summary

Variability in white matter fiber tracts of the human brain is associated with hundreds of newly discovered genomic loci that especially implicate stem, neural and glial cells during prenatal development, and is also associated with polygenic dispositions to various brain disorders and behavioural traits.
Identification of cyclic-di-GMP-modulating protein residues by bi-directionally evolving a social trait in Pseudomonas fluorescens
Authors: Kessler Collin , Kim Wook
Published: May 2022

Modulation of the intracellular cyclic di-guanosine monophosphate (c-di-GMP) pool is central to the formation of structured bacterial communities. Genome annotations predict the presence of dozens of conserved c-di-GMP catalytic enzymes in many bacterial species, but the functionality and regulatory control of the vast majority remain underexplored. Here, we begin to fill this gap by utilizing an experimental evolution system in Pseudomonas fluorescens Pf0-1, which repeatedly produces a unique social trait through bidirectional transitions between two distinct phenotypes converging on c-di-GMP modulation. Parallel evolution of 33 lineages captured 147 unique mutations among 191 evolved isolates in genes that are empirically demonstrated, bioinformatically predicted, or previously unknown to impact the intracellular pool of c-di-GMP. Quantitative chemistry confirmed that each mutation causing the phenotypic shift predictably amplifies or reduces c-di-GMP production. We integrate our mutation, phenotype, and quantification data with current models of known regulatory and catalytic systems, describe a previously unknown relationship between a regulatory component of branched-chain amino acid biosynthesis and c-di-GMP production, and predict functions of unexpected proteins that clearly impact c-di-GMP production. Sequential mutations that continuously disrupt or recover c-di-GMP production across discrete functional elements suggest a complex and underappreciated interconnectivity within the c-di-GMP regulome of P. fluorescens.

Importance

Microbial communities comprise densely packed cells where competition for space and resources is fierce. In our model system, mutant cells with a dry (D) phenotype are selected from a population with a mucoid (M) phenotype, and vice versa, because M and D cells physically work together to spread away from the overcrowded colony. D cells produce high levels of c-di-GMP and M cells produce low levels, so each mutation impacts c-di-GMP production. C-di-GMP is a second messenger which regulates diverse bacterial phenotypes that cause tremendous clinical and environmental problems. Many bacteria possess dozens of enzymes that are predicted to produce c-di-GMP, but most are considered to be non-functional. Here, we take advantage of the bi-directional selection of M and D phenotypes to identify key residues that could force these enzymes to turn on or off. Several unexpected proteins also participate in this process, but very little is known about them.
Haplotype-resolved assembly of a tetraploid potato genome using long reads and low-depth offspring data
Authors: Serra Mari Rebecca , Schrinner Sven , Finkers Richard , Arens Paul , Schmidt Maximilian H.-W. , Usadel Björn , Klau Gunnar W. , Marschall Tobias
Published: May 2022

Potato is one of the world’s major staple crops and like many important crop plants it has a polyploid genome. Polyploid haplotype assembly poses a major computational challenge, hindering the use of genomic data in breeding strategies. Here, we introduce a novel strategy for the assembly of polyploid genomes and present an assembly of the autotetraploid potato cultivar Altus. Our method uses low-depth sequencing data from an offspring population, which is available in many plant breeding settings, to achieve chromosomal clustering and haplotype phasing directly on the assembly graph. This involves a novel strategy for the analysis of k-mers unique to specific graph nodes. Our approach generates assemblies of individual chromosomes with phased haplotig N50 values of up to 13 Mb and haplotig lengths of up to 31 Mb. This major advance provides high-quality assemblies with haplotype-specific sequence resolution of whole chromosome arms and can be applied in common breeding scenarios where collections of offspring are available.
Biosynthetic proteins targeting the SARS-CoV-2 spike as anti-virals
Authors: Thébault Stéphanie , Lejal Nathalie , Dogliani Alexis , Donchet Amélie , Urvoas Agathe , Valerio-Lepiniec Marie , Lavie Muriel , Baronti Cécile , Touret Franck , da Costa Bruno , Bourgon Clara , Fraysse Audrey , Saint-Albin-Deliot Audrey , Morel Jessica , Klonjkowski Bernard , de Lamballerie Xavier , Dubuisson Jean , Roussel Alain , Minard Philippe , Le Poder Sophie , Meunier Nicolas , Delmas Bernard
Published: May 2022

The binding of the SARS-CoV-2 spike to angiotensin-converting enzyme 2 (ACE2) promotes virus entry into the cell. Targeting this interaction represents a promising strategy to generate antivirals. By screening a phage-display library of biosynthetic protein sequences build on a rigid alpha-helicoidal HEAT-like scaffold (named αReps), we selected candidates recognizing the spike receptor binding domain (RBD). Two of them (F9 and C2) bind the RBD with affinities in the nM range, displaying neutralisation activity in vitro and recognizing distinct sites, F9 overlapping the ACE2 binding motif. The F9-C2 fusion protein and a trivalent αRep form (C2-foldon) display 0.1 nM affinities and EC50 of 8-18 nM for neutralization of SARS-CoV-2. In hamsters, F9-C2 instillation in the nasal cavity before or during infections effectively reduced the replication of a SARS-CoV-2 strain harbouring the D614G mutation in the nasal epithelium. Furthermore, F9-C2 and/or C2-foldon effectively neutralized SARS-CoV-2 variants (including delta and omicron variants) with EC50 values ranging from 13 to 32 nM. With their high stability and their high potency against SARS-CoV-2 variants, αReps provide a promising tool for SARS-CoV-2 therapeutics to target the nasal cavity and mitigate virus dissemination in the proximal environment.

Author Summary

The entry of SARS-CoV-2 in permissive cells is mediated by the binding of its spike to angiotensin-converting enzyme 2 (ACE2) on the cell surface. To select ligands able to block this interaction, we screened a library of phages encoding artificial proteins (named αReps) for binding to its receptor binding domain (RBD). Two of them were able to bind the RBD with high affinity and block efficiently the virus entry in cultured cells. Assembled αReps through covalent or non-covalent linkages blocked virus entry at lower concentration than their precursors (with around 20-fold activity increase for a trimeric αRep). These αReps derivates neutralize efficiently SARS-CoV-2 β, γ, δ and Omicron virus variants. Instillation of an αRep dimer in the nasal cavity effectively reduced virus replication in the hamster model of SARS-CoV-2 and pathogenicity.
FastIntegration: a versatile R package for accessing and integrating large-scale single-cell RNA-seq data
Authors: Li Mengwei , Zhang Xiaomeng , Ang Kok Siong , Chen Jinmiao
Published: May 2022

Constructing atlas-scale comprehensive cell maps from publicly available data enables extensive data mining to achieve novel biological insights. Data integration to construct such maps require both harmonized datasets and an atlas-scale capable data integration tool. The first requirement is met by DISCO, a comprehensive repository of harmonized publicly available single cell data with standardized annotation. To meet the second requirement, the tool must have the capacity to integrate hundreds if not thousands of samples within an acceptable time frame. Moreover, it should output batch-corrected gene expression values to facilitate downstream analyses. Here, we present FastIntegration, a package which allows users to access and integrate public data in a convenient way. FastIntegration provides a fast and high-capacity version of Seurat Integration which can integrate more than 4 million cells within 2 days. It outputs batch corrected values for all genes that we can use for downstream analyses. For the first time, we demonstrated that using batch corrected values can improve the performance of downstream analyses. In particular, we found more accurately identified differentially expressed genes for cell types that are not shared between batches. Moreover, we also showed that FastIntegration outperforms existing methods for both homogeneous and heterogeneous data integration. FastIntegration also provides an API for programmatic access to data hosted on DISCO, a single-cell RNA-seq database that contains more than 5200 single-cell datasets. Users can filter for data at the sample level by tissue, disease and platform etc., and at cell level by specifying the expressed and unexpressed genes.
Aberrant regulation of serine metabolism drives extracellular vesicle release and cancer progression
Authors: Yamamoto Tomofumi , Nakayama Jun , Urabe Fumihiko , Ito Kagenori , Nishida-Aoki Nao , Kitagawa Masami , Yokoi Akira , Kuroda Masahiko , Hattori Yutaka , Yamamoto Yusuke , Ochiya Takahiro
Published: May 2022

Cancer cells secrete extracellular vesicles (EVs) to regulate cells in the tumor microenvironment to benefit their own growth and survive in the patient’s body. Although emerging evidence has demonstrated the molecular mechanisms of EV release, regulating cancer-specific EV secretion remains challenging. In this study, we applied a microRNA library to reveal the universal mechanisms of EV secretion from cancer cells. Here, we identified miR-891b and its direct target gene, phosphoserine aminotransferase 1 (PSAT1), which promotes EV secretion through the serine-ceramide synthesis pathway. Inhibition of PSAT1 affected EV secretion in multiple types of cancer, suggesting that the miR-891b/PSAT1 axis shares a common mechanism of EV secretion from cancer cells. Interestingly, aberrant PSAT1 expression also regulated cancer metastasis via EV secretion. Our data link between the PSAT1-controlled EV secretion mechanism and cancer metastasis and show the potential of this mechanism as a therapeutic target in multiple types of cancer.
Pax6 mutant cerebral organoids partially recapitulate phenotypes of Pax6 mutant mouse strains
Authors: Ferdaos Nurfarhana , Lowell Sally , Mason John O.
Published: May 2022

Cerebral organoids show great promise as tools to unravel the complex mechanisms by which the mammalian brain develops during embryogenesis. We generated mouse cerebral organoids harbouring constitutive or conditional mutations in Pax6, which encodes a transcription factor with multiple important roles in brain development. By comparing the phenotypes of mutant organoids with the well-described phenotypes of Pax6 mutant mouse embryos, we evaluated the extent to which cerebral organoids reproduce phenotypes previously described in vivo. Organoids lacking Pax6 showed multiple phenotypes associated with its activity in mice, including precocious neural differentiation, altered cell cycle and an increase in abventricular mitoses. Neural progenitors in both Pax6 mutant and wild type control organoids cycled more slowly than their in vivo counterparts, but nonetheless we were able to identify clear changes to cell cycle attributable to the absence of Pax6. Our findings support the value of cerebral organoids as tools to explore mechanisms of brain development, complementing the use of mouse models.
Comparing DNA metabarcoding with morphology in the assessment of macrozoobenthos in Portuguese transitional waters in the scope of the Water Framework Directive monitoring
Authors: Duarte Sofia , Vieira Pedro E. , Leite Barbara R. , Teixeira Marcos A.L. , Neto João M. , Costa Filipe O.
Published: May 2022

Despite the growing use and potential of DNA metabarcoding to improve and expedite macrozoobenthos monitoring, its employment in Water Framework Directive (WFD) monitoring of transitional ecosystems still remains largely unexplored and pending proof-of-concept studies. In the current study, we addressed this research gap by building upon the biomonitoring network program of the Portuguese Environmental Agency (APA) to benchmark metabarcoding against the morphology-based approach for characterizing macrozoobenthic communities. We assessed the ecological condition of 20 sites from four major transitional ecosystems in the west coast of Portugal, namely Minho, Lima, Vouga and Mondego estuaries. A total of 154 marine invertebrate species were detected with both methodologies, distributed by 11 phyla. In the majority of the sites, metabarcoding returned a higher number of species and phyla than the morphology-based approach (up to 2.5 times higher). In parallel, the proportion of species detected concurrently by both methods was low (35 species, 23%). The use of a multi-locus strategy increased recovered diversity through metabarcoding, since 37 species were detected exclusively with COI and 46 with 18S. For about 61% of the species recovered through morphology, metabarcoding failed detection, among which 20% was due to the lack of reference sequences in genetic databases. For the remaining, we did not find any plausible reason for only 10%, which could be due either to inefficient DNA extraction or PCR failure. Although morphological and metabarcoding-derived biotic indices did not match completely, similar responses to the environmental gradient were obtained in morphology and metabarcoding based-datasets. We anticipate that metabarcoding can increase the throughput and quality of the assessments, allowing faster assessments with greater spatial-temporal density and robust identifications of all specimens in a sample including larval stages, juveniles, and cryptic lineages, as well as smaller taxonomic groups that cannot be identified to species level using the traditional approach.
Genome sequence assembly evaluation using long-range sequencing data
Authors: Guan Dengfeng , McCarthy Shane A. , Wood Jonathan M. D. , Sims Ying , Chow William , Ning Zemin , Howe Kerstin , Wang Guohua , Wang Yadong , Durbin Richard
Published: May 2022

Genome sequences are computationally assembled from millions of much shorter sequencing reads. Although this process can be impressively accurate with long reads, it is still subject to a variety of types of errors, including large structural misassembly errors in addition to localised base pair substitutions. Recent advances in long single molecule sequencing in combination with other long-range technologies such as synthetic long read clouds and Hi-C have dramatically increased the contiguity of assembly. This makes it all the more important to be able to validate the structural integrity of the chromosomal scale assemblies now being generated. Here we describe a novel assembly evaluation tool, Asset, which evaluates the consistency of a proposed genome assembly with multiple primary long-range data sets, identifying both supported regions and putative structural misassemblies. We present tests on three de novo assemblies from a human, a goat and a fish species, demonstrating that Asset can identify structural misassemblies accurately by combining regionally supported evidence from long read and other raw sequencing data. Not only can Asset be used to assess overall assembly confidence, and discover specific problematic regions for downstream genome curation, a process that leads to improvement in genome quality, but it can also provide feedback to automated assembly pipelines.
The N-terminal domain of MX1 proteins is essential for their antiviral activity against different families of RNA viruses
Authors: McKellar Joe , Arnaud-Arnould Mary , Chaloin Laurent , Tauziet Marine , Arpin-André Charlotte , Pourcelot Oriane , Blaise Mickael , Moncorgé Olivier , Goujon Caroline
Published: May 2022

Myxovirus resistance protein 1 (MX1) and MX2, are homologous, dynamin-like large GTPases, induced upon interferon (IFN) exposure. Human MX1 (HsMX1) is known to inhibit many viruses, including influenza A virus (IAV), by likely acting at various steps of their life cycles. Despite decades of studies, the mechanism(s) of action with which MX1 proteins manage to inhibit target viruses is not fully understood. MX1 proteins are mechano-enzymes and share a similar organization to dynamin, with an amino-terminal GTPase domain and a carboxy-terminal stalk domain, connected by a Bundle Signalling Element (BSE). These three elements are known to be essential for antiviral activity. HsMX1 has two unstructured regions, the L4 loop, also essential for antiviral activity, and a short amino (N)-terminal region, which greatly varies between MX1 proteins of different species. The role of this N-terminal domain in antiviral activity is not known. Herein, using mutagenesis, imaging and biochemical approaches, we demonstrate that the N-terminal domain of HsMX1 is essential for antiviral activity against IAV and Vesicular Stomatitis Virus (VSV), and for the ability to aggregate Orthobunyavirus nucleoproteins. Furthermore, we pinpoint a highly conserved leucine within this region, which is absolutely crucial for human, mouse and bat MX1 protein antiviral activity. Importantly, mutation of this leucine does not compromise GTPase activity or oligomerization capabilities, but does modify MX1 protein subcellular localisation. The discovery of this essential and highly conserved residue defines this region as key and may reveal insights as to the mechanism of action of MX1 proteins.
Macrophages inhibit Coxiella burnetii by the ACOD1-itaconate pathway for containment of Q fever
Authors: Kohl Lisa , Alam Siddique Md. Nur A , Bodendorfer Barbara , Berger Raffaela , Preikschat Annica , Daniel Christoph , Ölke Martha , Mauermeir Michael , Yang Kai-Ting , Hayek Inaya , Szperlinski Manuela , Schulze-Luehrmann Jan , Schleicher Ulrike , Bozec Aline , Krönke Gerhard , Murray Peter J. , Wirtz Stefan , Yamamoto Masahiro , Schatz Valentin , Jantsch Jonathan , Oefner Peter , Degrandi Daniel , Pfeffer Klaus , Rauber Simon , Bogdan Christian , Dettmer Katja , Lührmann Anja , Lang Roland
Published: May 2022

Infection with the intracellular bacterium Coxiella (C.) burnetii can cause chronic Q fever with severe complications and limited treatment options. Here, we identify the enzyme cis- aconitate decarboxylase 1 (ACOD1 or IRG1) and its product itaconate as protective host immune pathway in Q fever. Infection of mice with C. burnetii induced expression of several anti-microbial candidate genes, including Acod1. In macrophages, Acod1 was essential for restricting C. burnetii replication, while other antimicrobial pathways were dispensable. Intratracheal or intraperitoneal infection of Acod1-/- mice caused increased C. burnetii burden, significant weight loss and stronger inflammatory gene expression. Exogenously added itaconate restored pathogen control in Acod1-/- mouse macrophages and blocked replication in human macrophages. In axenic cultures, itaconate directly inhibited growth of C. burnetii. Finally, treatment of infected Acod1-/-mice with itaconate efficiently reduced the tissue pathogen load. Thus, ACOD1-derived itaconate is a key factor in the macrophage-mediated defense against C. burnetii and may be exploited for novel therapeutic approaches in chronic Q fever.
Selectively expressed RNA molecules: a new dimension in functionalized cell targeting
Authors: Hoffmann Marco , Schüle Sylvie , Hoffmann Christina , Rastfeld Frederik , Gerlach Sven , Hersch N. , Walter Helene L. , Wiedermann Dirk , Fink Gereon R. , Merkel Rudolf , Bohlen Heribert , Rueger Maria A. , Hoffmann Bernd
Published: May 2022

Exact targeting of specific mammalian cell types or diseased cells is one of the most urgently needed prerequisites for a new generation of potent pharmaceuticals. Different approaches have been pursued, failing mainly due to a lack of specific surface markers in most cases. Developing a completely novel RNA-based methodology, we can now ensure exact cell targeting and simultaneously combine this with selective expression of effector proteins, thereby functionalization of the target cell for therapy, diagnostics or cell steering. The specific combination of the molecular properties of antisense technology and mRNA therapy with functional RNA secondary structures allowed us to develop selectively expressed RNA molecules for medical applications. These so-called seRNAs remain inactive in non-target cells and are only activated by partial degradation to induce translation in preselected cell types of interest. Cell type specificity and type of functionalization are easily adaptable based on a simple modular system. In proof of concept in vitro and in vivo studies we used seRNAs as a highly selective platform technology for powerful glioblastoma cancer cell targeting and significantly reduce brain tumors of mice without detectable side effects with just a single treatment within days. Our data open up new potential avenues for the efficient treatment of various cancers and other human diseases.
Molecular Mechanisms of Steric Pressure Generation and Membrane Remodeling by Intrinsically Disordered Proteins
Authors: Houser Justin R. , Cho Hyun Woo , Hayden Carl C. , Yang Noel X. , Wang Liping , Lafer Eileen M. , Thirumalai D. , Stachowiak Jeanne C.
Published: May 2022

Cellular membranes, which are densely crowded by proteins, take on an elaborate array of highly curved shapes. Steric pressure generated by protein crowding plays a significant role in shaping membrane surfaces. It is increasingly clear that many proteins involved in membrane remodeling contain substantial regions of intrinsic disorder. These domains have large hydrodynamic radii, suggesting that they may contribute significantly to steric congestion on membrane surfaces. However, it has been unclear to what extent they are capable of generating steric pressure, owing to their conformational flexibility. To address this gap, we use a recently developed sensor based on Förster resonance energy transfer to measure steric pressure generated at membrane surfaces by the intrinsically disordered domain of the endocytic protein, AP180. We find that disordered domains generate substantial steric pressure that arises from both entropic and electrostatic components. Interestingly, this steric pressure is largely invariant with the molecular weight of the disordered domain, provided that coverage of the membrane surface is held constant. Moreover, equivalent levels of steric pressure result in equivalent degrees of membrane remodeling, regardless of protein molecular weight. This result, which is consistent with classical polymer scaling relationships for semi-dilute solutions, helps to explain the molecular and physical origins of steric pressure generation by intrinsically disordered domains. From a physiological perspective, these findings suggest that a broad range of membrane-associated disordered domains are likely to play a significant and previously unknown role in controlling membrane shape.

Significance

With nearly half their surfaces covered by proteins, biological membranes are highly crowded. Rapid diffusion and collision of membrane-bound proteins generates substantial steric pressure that is capable of shaping membrane surfaces. Many proteins involved in membrane remodeling, are intrinsically disordered. Having large hydrodynamic radii, disordered domains could contribute substantially to membrane crowding. However, it is unclear to what extent they are capable of generating steric pressure, owing to their conformational flexibility. Toward resolving this uncertainty, we have measured steric pressure at membrane surfaces during dynamic membrane remodeling events. Our data indicate that disordered domains generate significant steric pressure through entropic and electrostatic mechanisms, suggesting that they may constitute a critical, yet previously neglected class of membrane remodeling proteins.
SpatialDEG: Identification of differentially expressed genes by leveraging spatial information in spatially resolved transcriptomic studies
Authors: Yang Yi , Lim Jeffrey ChunTatt , Ng Cedric Chuan Young , Lee Jing Yi , Yeong Joe , Sun Lei , Liu Jin
Published: May 2022

Motivation

Spatially resolved transcriptomics (SRT) technologies have been developed to simultaneously profile gene expression while retaining physical information. To explore differentially expressed genes using SRT in the context of various conditions, statistical methods are needed to perform spatial differential expression analysis.

Results

We propose that a new probabilistic framework, spatialDEG, can perform differential expression analysis by leveraging spatial information on gene expression with spatial information. SpatialDEG utilizes the average information algorithm and can be scalable to tens of thousands of genes. Comprehensive simulations demonstrated that spatialDEG can identify genes differentially expressed in tissues across different conditions with a controlled type-I error rate. We further applied spatialDEG to analyze datasets for human dorsolateral prefrontal cortex and mouse whole liver.

Availability

The R package spatialDEG can be downloaded from https://github.com/Shufeyangyi2015310117/spatialDEG.
Optogenetics and electron tomography for structure-function analysis of cochlear ribbon synapses
Authors: Chakrabarti Rituparna , Jaime Tobon Lina Maria , Slitin Loujin , Redondo-Canales Magdalena , Hoch Gerhard , Slashcheva Marina , Fritsch Elisabeth , Bodensiek Kai , Özçete Özge Demet , Gültas Mehmet , Michanski Susann , Opazo Felipe , Neef Jakob , Pangrsic Tina , Moser Tobias , Wichmann Carolin
Published: May 2022

Ribbon synapses of cochlear inner hair cells (IHCs) are specialized to indefatigably transmit sound information at high rates. To understand the underlying mechanisms, structure-function analysis of the active zone (AZ) of these synapses is essential. Previous electron microscopy studies of synaptic vesicle (SV) dynamics at the IHC AZ used potassium stimulation, which limited the temporal resolution to minutes. Here, we established optogenetic IHC stimulation followed by quick freezing within milliseconds and electron tomography to study the ultrastructure of functional synapse states with good temporal resolution. We characterized optogenetic IHC stimulation by patch-clamp recordings from IHCs and postsynaptic boutons revealing robust IHC depolarization and transmitter release. Ultrastructurally, the number of docked SVs increased and distances to the presynaptic density decreased upon short (17-25 ms) and long (48-76 ms) light stimulation paradigms. We did not observe enlarged SVs or other morphological correlates of homotypic fusion events. Our results suggest a rapid replenishment of docked SVs at IHC ribbon synapses and argue against synchronized multiquantal release under our experimental conditions.
A neural code for spatiotemporal context
Authors: Schonhaut Daniel R. , Aghajan Zahra M. , Kahana Michael J. , Fried Itzhak
Published: May 2022

Time and space are principle organizing dimensions of human experience. Whereas separate lines of study have identified neural correlates of time and space, little is known about how these representations converge during self-guided experience. Here we asked how neurons in the human brain represent time and space concurrently. Subjects fitted with intracranial microelectrodes played a timed navigation game where they alternated between searching for and retrieving objects in a virtual environment. Significant proportions of both time- and place-selective neurons were present during navigation, and distinct time-selective neurons appeared during task-free delays absent movement. We find that temporal and spatial codes are dissociable, with time cells remapping between search and retrieval tasks while place cells maintained stable firing fields. Other neurons tracked the context unique to each task phase, independent of time or space. Together these neuronal classes comprise a biological basis for the cognitive map of spatiotemporal context.
“Human platelet lysate derived extracellular vesicles enhance angiogenesis through miR-126”
Authors: Bordin Antonella , Chirivì Maila , Pagano Francesca , Milan Marika , Iuliano Marco , Scaccia Eleonora , Fortunato Orazio , Mangino Giorgio , Dhori Xhulio , De Marinis Elisabetta , D’Amico Alessandra , Miglietta Selenia , Picchio Vittorio , Rizzi Roberto , Romeo Giovanna , Pulcinelli Fabio , Chimenti Isotta , Frati Giacomo , De Falco Elena
Published: May 2022

Objectives

extracellular vesicles (EVs) are key biological mediators of several physiological functions within the cell microenvironment. Platelets are the most abundant source of EVs in the blood. Similarly, platelet lysate (PL), the best platelet derivative and angiogenic performer for regenerative purposes, is enriched of EVs, but their role is still too poorly discovered to be suitably exploited. Here we explored the contribution of the EVs in PL, by investigating the angiogenic features extrapolated from that possessed by PL.

Methods

we tested angiogenic ability and molecular cargo in 3D bioprinted models and by RNA sequencing analysis of PL-derived EVs.

Results

a subset of small vesicles is highly represented in PL. The EVs do not retain aggregation ability, preserving a low redox state in HUVEC and increasing the angiogenic tubularly-like structures in 3D endothelial bioprinted constructs. EVs resembled the miRNome profile of PL, mainly enriched of small RNAs and a high amount of miR-126, the most abundant angiogenic miRNA in platelets. The transfer of miR-126 by EVs in HUVEC after the in vitro inhibition of the endogenous form, restored angiogenesis, without involving VEGF as downstream target in this system.

Conclusions

PL is a biological source of available EVs with angiogenic effects involving a miRNAs-based cargo. These properties can be exploited for targeted molecular/biological manipulation of PL, by potentially developing a product exclusively manufactured of EVs.
Biochemical Characterization of Emerging SARS-CoV-2 Nsp15 Endoribonuclease Variants
Authors: Wilson Isha M. , Frazier Meredith N. , Li Jian-Liang , Randall Thomas A. , Stanley Robin E.
Published: May 2022

Global sequencing efforts from the ongoing COVID-19 pandemic, caused by the novel coronavirus SARS-CoV-2, continue to provide insight into the evolution of the viral genome. Coronaviruses encode 16 nonstructural proteins, within the first two-thirds of their genome, that facilitate viral replication and transcription as well as evasion of the host immune response. However, many of these viral proteins remain understudied. Nsp15 is a uridine-specific endoribonuclease conserved across all coronaviruses. The nuclease activity of Nsp15 helps the virus evade triggering an innate immune response. Understanding how Nsp15 has changed over the course of the pandemic, and how mutations affect its RNA processing function, will provide insight into the evolution of an oligomerization-dependent endoribonuclease and inform drug design. In combination with previous structural data, bioinformatics analyses of 1.9+ million SARS-CoV-2 sequences revealed mutations across Nsp15’s three structured domains (N-terminal, Middle, EndoU). Selected Nsp15 variants were characterized biochemically and compared to wild type Nsp15. We found that mutations to important catalytic residues decreased cleavage activity but increased the hexamer/monomer ratio of the recombinant protein. Many of the highly prevalent variants we analyzed led to decreased nuclease activity as well as an increase in the inactive, monomeric form. Overall, our work establishes how Nsp15 variants seen in patient samples affect nuclease activity and oligomerization, providing insight into the effect of these variants in vivo.
The SARS-CoV-2 Spike Protein Activates the Epidermal Growth Factor Receptor-Mediated Signaling
Authors: Palakkott Abdulrasheed , Alneyadi Aysha , Muhammad Khalid , Eid Ali H. , Amiri Khaled , Ayoub Mohammed Akli , Iratni Rabah
Published: May 2022

Objectives

The coronavirus disease-19 (COVID-19) pandemic is caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). At the molecular and cellular levels, the SARS-Cov-2 uses its envelope glycoprotein, the spike S protein, to infect the target cells in the lungs via binding with their transmembrane receptor, the angiotensin-converting enzyme 2 (ACE2). Here, we wanted to invesitgate if other molecular targets and pathways may be used by SARS-Cov-2.

Methods

We investigated the possibility for the spike 1 S protein and its receptor-binding domain (RBD) to target the epidermal growth factor receptor (EGFR) and its downstream signaling pathway in vitro using the lung cancer cell line (A549 cells). Protein expression and phosphorylation was examined upon cell treatment with the recombinant full spike 1 S protein or RBD.

Results

We demonstrate for the first time the activation of EGFR by the Spike 1 protein associated with the phosphorylation of the canonical ERK1/2 and AKT kinases and an increase of survivin expression controlling the survival pathway.

Conclusions

Our study suggests the putative implication of EGFR and its related signaling pathways in SARS-CoV-2 infectivity and Covid-19 pathology. This may open new perspectives in the treatment of Covid-19 patients by targeting EGFR.
Novel tumorigenic FOXM1-PTAFR-PTAF axis revealed by multi-omic profiling in TP53/CDKN2A-double knockout human gastroesophageal junction organoid model
Authors: Zhao Hua , Cheng Yulan , Kalra Andrew , Ma Ke , Zheng Yueyuan , Ziman Benjamin , Tressler Caitlin , Glunde Kristine , Shin Eun Ji , Ngamruengphong Saowanee , Khashab Mouen , Singh Vikesh , Anders Robert A. , Jit Simran , Wyhs Nicolas , Chen Wei , Li Xu , Lin De-Chen , Meltzer Stephen J.
Published: May 2022

Inactivation of the tumor suppressor genes TP53 and CDKN2A occurs early during gastroesophageal junction (GEJ) tumorigenesis. However, due to a paucity of GEJ-specific disease models, cancer-promoting consequences of TP53 and CDKN2A inactivation at the GEJ have been incompletely characterized. Here we report the development of the first wild-type primary human GEJ organoid model, as well as a CRISPR-edited transformed GEJ organoid model. CRISPR/Cas9 engineering to inactivate TP53 and CDKN2A (TP53/CDKN2AKO) in GEJ organoids induced morphologic dysplasia as well as pro-neoplastic features in vitro and tumor formation in vivo. Notably, lipidomic profiling identified several Platelet-Activating Factors (PTAFs) among the most upregulated lipids in CRISPR-edited organoids; and importantly, PTAF/PTAFR abrogation by siRNA knockdown or a pharmacologic inhibitor (WEB2086) significantly blocked proliferation and other pro-neoplastic features of TP53/CDKN2AKO GEJ organoids in vitro and tumor formation in vivo. In addition, murine xenografts derived from Eso26, an established esophageal adenocarcinoma (EAC) cell line, were suppressed by WEB2086. Mechanistically, TP53/CDKN2A dual inactivation disrupted both the transcriptome and the DNA methylome, likely mediated by key transcription factors, particularly Forkhead Box M1 (FOXM1). Importantly, FOXM1 activated PTAFR transcription by binding to the PTAFR promoter, further amplifying the PTAF-PTAFR pathway. In summary, we established a robust model system for investigating early GEJ neoplastic events, identified crucial metabolic and epigenomic changes occurring during GEJ model tumorigenesis, and discovered a potential cancer-therapeutic strategy, while providing insights into pro-neoplastic mechanisms associated with TP53/CDKN2A inactivation in early GEJ neoplasia.

One Sentence Summary

Novel tumorigenic FOXM1-PTAFR-PTAF axis revealed by multi-omic profiling in TP53/CDKN2A-double knockout human gastroesophageal junction organoid model.

Graphic Abstract
Identification of transcriptional network disruptions in drug-resistant prostate cancer with TraRe
Authors: Blatti Charles , de la Fuente Jesús , Gao Huanyao , Marín Irene , Chen Zikun , Zhao Sihai. D. , Tan Winston , Weinshilbaum Richard , Kalari Krishna R. , Wang Liewei , Hernaez Mikel
Published: May 2022

Metastatic castration-resistant prostate cancer (mCRPC) presents very low survival rates due to lack of response or acquired resistance to the available therapies. To date no molecular mechanisms of resistance have been identified, pointing out their complex dynamics. To identify key genes and processes associated with phenotypically-driven regulatory differences, we developed TraRe, a computational method that provides a three-tier analysis: i) at the network level, inferring differentially regulated modules; ii) at the regulon level, identifying regulatory relationships linked to phenotypic differences; and iii) at the single gene level, identifying TFs consistently linked to rewired modules. We applied TraRe (available in Bioconductor with full documentation) to transcriptomic data from 46 mCRPC patients with Abiraterone-response clinical data and uncovered abrogated immune response regulatory modules that showed strong differential regulation in Abi-resistant patients. These modules were replicated in an independent mCRPC study. Further, we experimentally validated key rewiring predictions and their associated transcription factors. Among them, ELK3, MXD1, and MYB were found to have a differential role in cell survival for Abi-response-specific settings. Moreover, we identified the role of ELK3 in cell migration capacity, which could have direct impact on mCRPC. Collectively, these findings shed light on the underlying regulatory mechanisms driving abiraterone response.
Back from the dead: the atypical kinase activity of a pseudokinase regulator of cation fluxes during inducible immunity
Authors: Brauer Elizabeth K. , Ahsan Nagib , Popescu George V. , Thelen Jay J. , Popescu Sorina C.
Published: May 2022

While most pseudokinases lack phosphotransfer activity due to altered catalytic residues within their kinase domain, a subset of pseudokinases maintain kinase activity through atypical mechanisms. The Arabidopsis ILK1 is a pseudokinase from the Raf-like MAP3K family and is the only known plant pseudokinase with confirmed kinase activity. ILK1 activity promotes microbial pattern-induced root growth inhibition through its stabilization of the HAK5 potassium transporter with the calmodulin-like protein CML9. ILK1 also has a kinase-independent function in salt stress suggesting that it interacts with additional proteins. We determined that members of the ILK subfamily are the sole pseudokinases within the Raf-like MAP3K family and identified 179 novel putative ILK1 protein interactors. We also identified 70 novel peptide targets for ILK1, the majority of which were phosphorylated in the presence of Mn2+instead of Mg2+in line with modifications in ILK1’s DFG cofactor binding domain. Overall, the ILK1-targeted or interacting proteins included diverse protein types including transporters (HAK5, STP1), protein kinases (MEKK1, MEKK3), and a cytokinin receptor (AHK2). The expression of 31 genes encoding putative ILK1-interacting or phosphorylated proteins, including AHK2, were altered in the root and shoot in response to molecular patterns suggesting a role for these genes in immunity. We describe a potential role for ILK1 interactors in the context of cation-dependent immune signaling responses, highlighting the importance of K+in MAMP responses. This work further supports the notion that ILK1 is an atypical kinase with an unusual cofactor dependence that may interact with multiple proteins in the cell.