bluuebunny/crossref_metadata_2025 huggingface dataset:
CaviDB: a database of cavities and their features in the structural and conformational space of proteins
Authors: Rueda Ana Julia Velez , Bulgarelli Franco Leonardo , Palopoli Nicolás , Parisi Gustavo
Published: August 2022

Proteins are the functional and evolutionary units of cells. On their surface, proteins are sculpted into numerous concavities and bulges, offering unique microenvironments for ligand binding or catalysis. The dynamics, size, and chemical features of these cavities are essential for the mechanistic understanding of protein function.

Here we present CaviDB (https://www.cavidb.org), a novel database of cavities and their features in known protein structures, which integrates the results from commonly used software for cavities detection with protein features obtained from sequence, structure, and function analyses. Additionally, each protein in CaviDB is associated with its corresponding conformers which help to analyze conformational changes in cavities as well. We were able to characterize a total number of 16,533,339 cavities, 62,0431 of them predicted as druggable targets. CaviDB contains 276,432 different proteins, with information about all their conformers. It also offers the capability to compare cavities and their features from different conformational states of the protein. Furthermore, we have recently added the available models from the AlphaFold database versions 2 and 3, which allow further cavity explorations and comparisons.

Each entry information is organized in sections, highlighting the general cavities descriptors, including the inter-cavities contacts, activated residues per cavity, the information about druggable cavities, and the global protein descriptors. The data retrieved by the user can be downloaded in a format that is easy to parse and integrate with custom pipelines for protein analysis.

CaviDB aims to offer a comprehensive database for use not only in different aspects of drug design and discovery but also to better understand the basis of the protein structure-function relationship better. With its unique approach, CaviDB provides an essential resource for the wide community of bioinformaticians in particular and biologists in general.
Homeostatic regulation through strengthening of neuronal network-correlated synaptic inputs
Authors: Barnes Samuel J. , Keller Georg B. , Keck Tara
Published: August 2022

Homeostatic regulation is essential for neuronal function. Several synaptic mechanisms of homeostatic plasticity have been described, but the functional properties of synapses involved in homeostasis are unknown. We used longitudinal two-photon functional imaging of dendritic spine calcium signals in visual and retrosplenial cortices of awake mice to quantify the sensory deprivation-induced changes in the responses of functionally identified spines. We found that spines whose activity selectively correlated with intrinsic network activity underwent TNF-α dependent homeostatic increases in their response amplitudes, but spines identified as responsive to sensory stimulation did not. We observed an increase in the global sensory-evoked responses following sensory deprivation, despite the fact that the identified sensory inputs did not strengthen. Instead, global sensory-evoked responses correlated with the strength of network-correlated inputs. Our results suggest that homeostatic regulation of global responses is mediated through changes to intrinsic network-correlated inputs rather than changes to identified sensory inputs thought to drive sensory processing.
A Reagent and Virus Benchmarking Panel for a Uniform Analytical Performance Assessment of N Antigen–Based Diagnostic Tests for COVID-19
Authors: Golden Allison , Cantera Jason L. , Lillis Lorraine , Phan Thanh T. , Slater Hannah , Webb Edwin J. , Peck Roger B. , Domingo Gonzalo J. , Boyle David S.
Published: August 2022

Rapid diagnostic tests (RDTs) that detect antigen indicative of SARS-CoV-2 infection can help in making quick health care decisions and regularly monitoring groups at risk of infection. With many RDT products entering the market, it is important to rapidly evaluate their relative performance. Comparison of clinical evaluation study results is challenged by protocol design variations and study populations. Laboratory assays were developed to quantify nucleocapsid (N) and spike (S) SARS-CoV-2 antigens. Quantification of the two antigens in nasal eluates confirmed higher abundance of N than S antigen. The median concentration of N antigen was 10 times greater than S per genome equivalent. The N antigen assay was used in combination with quantitative RT-PCR to qualify a panel composed of recombinant antigens, inactivated virus, and clinical specimen pools. This benchmarking panel was applied to evaluate the analytical performance of the SD Biosensor STANDARD Q COVID-19 Ag test, Abbott Panbio COVID-19 Ag Rapid Test, Abbott BinaxNOW COVID-19 Ag test, and the LumiraDx SARS-CoV-2 Ag Test. The four tests displayed different sensitivities toward the different panel members, but all performed best with the clinical specimen pool. The concentration for a 90% probability of detection across the four tests ranged from 21 pg/mL to 102 pg/mL of N antigen in the extracted sample. Benchmarking panels provide a quick way to verify the baseline performance of a diagnostic and enable direct comparison between diagnostic tests.
Human activities impact microbial communities of Amazon Mangroves
Authors: da Costa Gleyciane Machado , Costa Sávio Souza , Barauna Rafael Azevedo , Castilho Bruno Pureza , Pinheiro Izabel Cruz , Silva Artur , Schaan Ana Paula , Ribeiro-dos-Santos Ândrea Kely Campos , das Graças Diego Assis
Published: August 2022

Mangroves provide a unique ecological environment for complex microbial communities. They are particularly important in controlling the chemical environment of the ecosystem. Microbial diversity analyses of these ecosystems help us understand the environmental dynamics and the changes due to impacts. Amazonian mangroves are distributed across Amapá, Pará and Maranhão states occupying an area of 9,000 km2 and correspond to 70% of the mangroves in Brazil, in which studies on microbial biodiversity are extremely rare. The present study aimed to determine changes in microbial community structure along the PA-458 highway that divided the mangrove. Mangrove samples were collected from three zones, (i) under anthropogenic actions, (ii) in the process of recovery and (ii) preserved. Total DNA was extracted, submitted for 16S rDNA amplification and sequenced on the Illumina MiSeq platform. Reads were analyzed using USEARCH tools and packages available in R Platform. The most abundant phyla were Proteobacteria, followed by Firmicutes and Bacteroidetes in all three mangrove locations. Genera such as Desulfuromonas, Desulfatiglans, Collinsella, Dorea, Dialister were not found in the impacted mangrove. The impacted mangrove proved to be ecologically different from other mangroves. Our results show that human impact in the mangrove areas has resulted in loss of biodiversity, caused by the construction of the PA-458 highway, reducing the support and maintenance of habitat life and the ecosystem’s resilience to disturbance.
Combinatorial perturbation sequencing on single cells using microwell-based droplet random pairing
Authors: Xie Run , Liu Yang , Wang Shiyu , Shi Xuyang , Zhao Zhantao , Liu Longqi , Liu Ya , Li Zida
Published: August 2022

Combinatorial drug therapy reduces drug resistance and disease relapse, but informed drug combinations are lacking due to the high scale of possible combinations and the relatively simple phenotyping strategies. Here we report combinatorial perturbation sequencing (CP-seq) on single cells using microwell-base droplet random pairing. CP-seq uses oligonucleotides to barcode drugs, encapsulates drugs and cells in separate droplets, and pairs cell droplets with two drug droplets randomly on a microwell array chip to complete combinatorial drug treatment and barcode-tagging on cells. The subsequent single-cell RNA sequencing simultaneously detects the single-cell transcriptomes and drug barcodes to demultiplex the corresponding drug treatment. The microfluidic droplet operations had robust performance, with overall success rate among the microwells being up to 83%. We then progressively validated the CP-seq by performing single drug treatment and then combinatorial drug treatment. Leveraging the advantage of droplet microfluidics in massive multiplexing, the CP-seq can test thousands of drug combinations in a single experiment and represents a great technology for combinatorial perturbation screening with high throughput and comprehensive profiling.
Visualizing proteins by expansion microscopy
Authors: Shaib Ali H. , Chouaib Abed Alrahman , Chowdhury Rajdeep , Mihaylov Daniel , Zhang Chi , Imani Vanessa , Georgiev Svilen Veselinov , Mougios Nikolaos , Monga Mehar , Reshetniak Sofiia , Mimoso Tiago , Chen Han , Fatehbasharzad Parisa , Crzan Dagmar , Saal Kim-Ann , Alawar Nadia , Eilts Janna , Kang Jinyoung , Alvarez Luis , Trenkwalder Claudia , Mollenhauer Brit , Outeiro Tiago F. , Köster Sarah , Preobraschenski Julia , Becherer Ute , Moser Tobias , Boyden Edward S. , Aricescu A Radu , Sauer Markus , Opazo Felipe , Rizzoli Silvio O.
Published: August 2022

Fluorescence imaging is one of the most versatile and widely-used tools in biology1. Although techniques to overcome the diffraction barrier were introduced more than two decades ago, and the nominal attainable resolution kept improving2, 3, fluorescence microscopy still fails to image the morphology of single proteins or small molecular complexes, either purified or in a cellular context4, 5. Here we report a solution to this problem, in the form ofone-stepnanoscaleexpansion (ONE) microscopy. We combined the 10-fold axial expansion of the specimen (1000-fold by volume) with a fluorescence fluctuation analysis6, 7to enable the description of cultured cells, tissues, viral particles, molecular complexes and single proteins. At the cellular level, using immunostaining, our technology revealed detailed nanoscale arrangements of synaptic proteins, including a quasi-regular organisation of PSD95 clusters. At the single molecule level, upon main chain fluorescent labelling, we could visualise the shape of individual membrane and soluble proteins. Moreover, conformational changes undergone by the ∼17 kDa protein calmodulin upon Ca2+binding were readily observable. We also imaged and classified molecular aggregates in cerebrospinal fluid samples from Parkinson’s Disease (PD) patients, which represents a promising new development towards improved PD diagnosis. ONE microscopy is compatible with conventional microscopes and can be performed with the software we provide here as a free, open-source package. This technology bridges the gap between high-resolution structural biology techniques and light microscopy, and provides a new avenue for discoveries in biology and medicine.
Single-molecule fluorescence microscopy demonstrates fast dynamics of the variant surface glycoprotein coat on living trypanosomes
Authors: Schwebs Marie , Paul Torsten , Glogger Marius , Forster Leonard , Morriswood Brooke , Teßmar Jörg , Groll Jürgen , Engstler Markus , Kollmannsberger Philip , Fenz Susanne
Published: August 2022

The fluidity of Trypanosoma brucei’s dense coat of GPI-anchored variant surface glycoproteins (VSGs) is fundamental for the survival of the parasite. In order to maintain the integrity of the coat, it is recycled on the time scale of a few minutes. This is surprisingly fast as endo- and exocytosis take place in the same small membrane invagination called the flagellar pocket. Here, we present measurements of VSG dynamics on the single-molecule level in living trypanosomes. A large number of short protein trajectories sampling the parasite’s surface were analysed in two distinct scenarios: diffusion and directed motion. To this end, we employed a previously published algorithm and implemented two extensions to consider rim effects as well as localisations errors inherent to single-mole tracking. Neglect of the latter can have a significant distortive effect on the measured diffusion coefficient; in our case resulting in an underestimation by 20 %. We found large heterogeneity in the local diffusion coefficients and velocities with a surprisingly high average value of and , respectively. To decide on the locally dominant motion mode, we present a guideline based on random walk simulations. We find that VSG dynamics is indeed dominated by diffusion. Complementary simulations on long time scales not accessible in the experiment showed that passive VSG randomisation is fast enough to prevent re-endocytosis newly exocytosed VSGs and to accomplish turnover of the full VSG coat within a few minutes.

Author summary

Single-molecule tracking in biological systems often suffers from trajectories being too short to obtain statistically robust decisions on the present motion mode. We faced this issue when investigating the dynamics of the protein surface coat of African trypanosomes. To address the question whether diffusion or directed motion governs coat dynamics, we have adopted an algorithm based on temporal decomposition and spatial binning of an ensemble of single-molecule trajectories. We introduced several extensions to the original approach, including the consideration of localisation errors inherent to single-molecule tracking. This improvement alone already prevented the diffusion coefficient from being underestimated by 20 %. We analysed the coat dynamics in two scenarios, diffusion and directed motion, and offer a decision guideline to identify the locally dominating motion mode. Our extended algorithm is available to the scientific community via GitHub. For the trypanosome surface coat we found that the motion is indeed mainly characterised by diffusion with a surprisingly high diffusion coefficient. This finding solves a long-standing question how the parasite maintains its protein coat.
Novel algorithms for improved detection and analysis of fluorescent signal fluctuations
Authors: Stopper Gebhard , Caudal Laura C. , Rieder Phillip , Gobbo Davide , Felix Lisa , Everaerts Katharina , Bai Xianshu , Stopper Laura , Rose Christine R. , Scheller Anja , Kirchhoff Frank
Published: August 2022

Fluorescent dyes and genetically encoded fluorescence indicators (GEFI) are common tools for visualizing concentration changes of specific ions and messenger molecules during intra-as well as intercellular communication. Using advanced imaging technologies, fluorescence indicators are a prerequisite for the analysis of physiological molecular signaling. Automated detection and avnalysis of fluorescence signals requires to overcome several challenges, including correct estimation of fluorescence fluctuations at basal concentrations of messenger molecules, detection and extraction of events themselves as well as proper segmentation of neighboring events. Moreover, event detection algorithms need to be sensitive enough to accurately capture localized and low amplitude events exhibiting a limited spatial extent. Here, we present two algorithms (PBasE and CoRoDe) for accurate baseline estimation of fluorescent detection of messenger molecules and automated detection of fluorescence fluctuations.

Author summary

Analyzing molecular signalling is crucial in understanding intra- and intercellular communication. These signals are visualized using fluorescent dyes or genetically encoded fluorescence indicators. In the brain, Ca2+ signals of glial cells are essential in deciphering complex regulatory functions in health and disease. Due to signal heterogeneity, detection and analysis are highly challenging. They can be stationary, with low amplitude and localized in cell processes, occur as prominent somatic signals or propagate as waves across cellular networks.

We have developed two algorithms to analyze fluorescence transients, each tackling a specific problem. PBasE performs automatic and adaptive background correction, removing basal fluorescence fluctuations. CoRoDe automatically extracts regions of interest, explicitly including temporal information to obtain a precise segmentation, which is essential for accurate transient extraction. Combined, these algorithms are able to detect regions exhibiting low amplitude transients with small spatial extent as well as large, high amplitude signals. Extracted transients are categorized based on their peak amplitude, allowing detailed analyses by comparing changes of specific properties. In order to make these algorithms accessible, an interactive application, called Msparkles, has been designed.
Competitive HIV budding suggests that a self-packaging gRNA:Gag-Pol complex directs HIV assembly and enforces infectivity
Authors: Durden Haley , Saha Ipsita , Preece Benjamin , MacArthur Brian , Petersen Abby , Peppel Wiley , Gallegos Rodrigo , Saffarian Saveez
Published: August 2022

To resolve the assembly mechanism of infectious HIV virions, we tested the ability of HIV to assemble infectious virions in the presence of a titrated mix of infectious/ non-infectious proviral genomes. The analysis of our assembly competitions shows that during translation, 15 ± 5 Gag-Pols bind back to their parental gRNA creating a gRNA1: Gag-Pol15 complex. This complex initiates the infectious virion assembly through interactions mediated by cis packaged Gag/Gag-pols and the plasma membrane. Our analysis also shows the number of Gag-Pol and Env proteins packaged in an infectious HIV virion and the minimum functional units of these proteins required for viral infectivity. We suggest that aside from orchestrating the infectious virion assembly the gRNA1: Gag-Pol15 complex plays a major role in HIV evolution and likely hampers effectiveness of antiviral therapies.

One-Sentence Summary

Competitive HIV budding reveals gRNA1: Gag-Pol15 complex orchestrating viral assembly.
CaMKII activity spreads by inter-holoenzyme phosphorylation
Authors: Lučić Iva , Héluin Léonie , Jiang Pin-Lian , Castro Scalise Alejandro G. , Wang Cong , Franz Andreas , Wahl Markus , Liu Fan , Plested Andrew J.R.
Published: August 2022

The dodecameric protein kinase CaMKII is expressed throughout the body. The alpha isoform is responsible for synaptic plasticity and participates in memory through its phosphorylation of synaptic proteins. Its elaborate subunit organization and propensity for autophosphorylation allow it to preserve neuronal plasticity across space and time. The prevailing hypothesis for the spread of CaMKII activity, involving shuffling of subunits between activated and naïve holoenzymes, is broadly termed subunit exchange. In contrast to the expectations of previous work, we found little evidence for subunit exchange upon activation, and no effect of restraining subunits to their parent holoenzymes. Rather, mass photometry, crosslinking mass spectrometry, single molecule TIRF microscopy and biochemical assays identify inter-holoenzyme phosphorylation (IHP) as the mechanism for spreading phosphorylation. The transient, activity-dependent formation of groups of holoenzymes is well suited to the speed of neuronal activity. Our results place fundamental limits on the activation mechanism of this kinase.
The cost of sympatry: spatio-temporal patterns in leopard dietary and physiological responses to tiger competition in Rajaji Tiger Reserve, India
Authors: Patel Shiv Kumari , Ruhela Sourabh , Biswas Suvankar , Bhatt Supriya , Pandav Bivash , Mondol Samrat
Published: August 2022

1.

Apex predators have critical roles in maintaining the structure of ecosystem functioning by controlling intraguild subordinate populations. Such dominant-subordinate interactions involve agonistic interactions including direct (death/displacement) or indirect (physiological and/or health implications) impacts on the subordinates. As these indirect effects are often mediated through physiological processes, it is important to quantify such responses for better understanding of population parameters.

We used a well-known sympatric large carnivore intraguild system involving tiger (Panthera tigris) and leopard (Panthera pardus) to understand the dietary and physiological responses under a spatio-temporal gradient of tiger competition pressures in Rajaji Tiger Reserve (RTR), a major source tiger population of the western Terai-Arc Landscape, India between 2015-2020. The park provided a natural experimental set-up for tiger competition with the eastern part (ERTR) having high tiger density and the western (WRTR) part with functionally no competition from tigers.

We conducted systematic faecal sampling in the winters of 2015 and 2020 from ERTR and WRTR to assess diet and physiological measures. Analyses of leopard-confirmed faeces suggest a dietary-niche separation as a consequence of tiger competition. In 2020, we found increased occurrence of large-bodied prey species without tiger competition in WRTR. Physiological measures followed the dietary responses where leopards with large-sized prey in diet showed higher fT3M and lower fGCM measures in WRTR. In contrast, ERTR leopards showed lower levels of fT3M as well as fGCM in 2020, possibly due to intense competition from tigers. Overall, these pattens strongly indicate a physiological cost of sympatry where competition with dominant tigers resulted in elevated nutritional stress.

The combination of the natural habitat providing unique experimental setup, spatio-temporal sampling strategy and multidisciplinary approaches provide critical conservation perspectives for leopards, particularly in the context of recent increase in tiger numbers across India. We recommend expansion of leopard monitoring and population estimation efforts to buffers, developing appropriate plans for human-leopard conflict mitigation and intensive efforts to understand leopard population dynamics patterns to ensure their persistence during the ongoing Anthropocene.
Faster detection of “darks” than “brights” by monkey superior colliculus neurons
Authors: Malevich Tatiana , Zhang Tong , Baumann Matthias P. , Bogadhi Amarender R. , Hafed Ziad M.
Published: August 2022

Visual processing is segregated into ON and OFF channels as early as in the retina, and the superficial (output) layers of the primary visual cortex are dominated by neurons preferring dark stimuli. However, it is not clear how the timing of neural processing differs between “darks” and “brights” in general, especially in light of psychophysical evidence; it is also equally not clear how subcortical visual pathways that are critical for active orienting represent stimuli of positive (luminance increments) and negative (luminance decrements) contrast polarity. Here, we recorded from all visually-responsive neuron types in the superior colliculus (SC) of two male rhesus macaque monkeys. We presented a disc (0.51 deg radius) within the response fields (RF’s) of neurons, and we varied, across trials, stimulus Weber contrast relative to a gray background. We also varied contrast polarity. There was a large diversity of preferences for darks and brights across the population. However, regardless of individual neural sensitivity, most neurons responded significantly earlier to dark than bright stimuli. This resulted in a dissociation between neural preference and visual response onset latency: a neuron could exhibit a weaker response to a dark stimulus than to a bright stimulus of the same contrast, but it would still have an earlier response to the dark stimulus. Our results highlight an additional candidate visual neural pathway for explaining behavioral differences between the processing of darks and brights, and they demonstrate the importance of temporal aspects in the visual neural code for orienting eye movements.

Significance statement

Objects in our environment, such as birds flying across a bright sky, often project shadows (or images darker than the surround) on our retina. We studied how primate superior colliculus (SC) neurons visually process such dark stimuli. We found that the overall population of SC neurons represented both dark and bright stimuli equally well, as evidenced by a relatively equal distribution of neurons that were either more or less sensitive to darks. However, independent of sensitivity, the great majority of neurons detected dark stimuli earlier than bright stimuli, evidenced by a smaller response latency for the dark stimuli. Thus, SC neural response latency can be dissociated from response sensitivity, and it favors the faster detection of dark image contrasts.
Sequencing the orthologs of human autosomal forensic short tandem repeats provides individual- and species-level identification in African great apes
Authors: Fedele Ettore , Wetton Jon H. , Jobling Mark A.
Published: August 2022

Background

Great apes are a global conservation concern, with anthropogenic pressures threatening their survival. Genetic analysis can be used to assess the effects of reduced population sizes and the effectiveness of conservation measures. In humans, autosomal short tandem repeats (aSTRs) are widely used in population genetics and for forensic individual identification and kinship testing. Traditionally, genotyping is length-based via capillary electrophoresis (CE), but there is an increasing move to direct analysis by massively parallel sequencing (MPS). Here we assess in African great ape DNAs the human-based ForenSeq DNA Sequencing Prep Kit, which amplifies multiple loci including 27 aSTRs, prior to sequencing via Illumina technology. We ask whether cross-species genotyping of the orthologs of these loci can provide both individual and (sub)species identification.

Results

The Forenseq kit was used to amplify and sequence aSTRs in 52 individuals (14 chimpanzees; 4 bonobos; 16 western lowland, 6 eastern lowland, and 12 mountain gorillas). The orthologs of 24/27 human aSTRs amplified across species, and a core set of thirteen loci could be genotyped in all individuals. Genotypes were individually and (sub)species identifying. Both allelic diversity and the power to discriminate (sub)species were greater when considering STR sequences rather than allele lengths. Comparing human and African great-ape STR sequences with an orangutan outgroup showed general conservation of repeat types and allele size ranges, but variation in repeat array structures and little relationship with the known phylogeny, suggesting stochastic origins of mutations giving rise to diverse imperfect repeat arrays. Interruptions within long repeat arrays in African great apes do not appear to reduce allelic diversity, indicating a possible mutational difference to humans.

Conclusions

Despite some variability in amplification success, orthologs of most human aSTRs in the ForenSeq DNA Sequencing Prep Kit can be analysed in African great apes. MPS of the orthologs of human loci provides better resolution for both individual and (sub)species identification in great apes than standard CE-based approaches, and has the further advantage that there is no need to limit the number and size ranges of analysed loci.
Reliability and accuracy of single-molecule FRET studies for characterization of structural dynamics and distances in proteins
Authors: Agam Ganesh , Gebhardt Christian , Popara Milana , Mächtel Rebecca , Folz Julian , Ambrose Benjamin , Chamachi Neharika , Chung Sang Yoon , Craggs Timothy D. , de Boer Marijn , Grohmann Dina , Ha Taekjip , Hartmann Andreas , Hendrix Jelle , Hirschfeld Verena , Hübner Christian G. , Hugel Thorsten , Kammerer Dominik , Kang Hyun-Seo , Kapanidis Achillefs N. , Krainer Georg , Kramm Kevin , Lemke Edward , Lerner Eitan , Margeat Emmanuel , Martens Kristen , Michaelis Jens , Mitra Jaba , Moya Muñoz Gustavo G. , Quast Robert , Robb Nicole B. , Sattler Michael , Schlierf Michael , Schneider Jonathan , Schröder Tim , Sefer Anna , Tan Piau Siong , Thurn Johann , Tinnefeld Philip , van Noort John , Weiss Shimon , Wendler Nicolas , Zijlstra Niels , Barth Anders , Seidel Claus A. M. , Lamb Don C. , Cordes Thorben
Published: August 2022

Single-molecule FRET (smFRET) has become an established tool to study biomolecular structure and dynamics in vitro and in live cells. We performed a worldwide blind study involving 19 labs to assess the uncertainty of FRET experiments for proteins with respect to the measured FRET efficiency histograms, determination of distances, and the detection and quantification of structural dynamics. Using two protein systems that undergo distinct conformational changes, we obtained an uncertainty of the FRET efficiency of less than ± 0.06, corresponding to an interdye distance precision of ≤ 0.2 nm and accuracy of ≤ 0.5 nm. We further discuss the limits for detecting distance fluctuations with sensitivity down to ≲ 10% of the Förster distance and provide guidelines on how to detect potential dye perturbations. The ability of smFRET experiments to simultaneously measure distances and avoid averaging of conformational dynamics slower than the fluorescence lifetime is unique for dynamic structural biology.
Cassette recombination dynamics within chromosomal integrons are regulated by toxin–antitoxin systems
Authors: Richard Egill , Darracq Baptiste , Littner Eloi , Vit Claire , Whiteway Clémence , Bos Julia , Fournes Florian , Garriss Geneviève , Conte Valentin , Lapaillerie Delphine , Parissi Vincent , Rousset François , Skovgaard Ole , Bikard David , Rocha Eduardo P. C. , Mazel Didier , Loot Céline
Published: August 2022

Integrons are adaptive bacterial devices that rearrange promoter less gene cassettes into variable ordered arrays under stress conditions, to sample combinatorial phenotypic diversity. Chromosomal integrons often carry hundreds of silent gene cassettes, with integrase-mediated recombination leading to rampant DNA excision and integration, posing a potential threat to genome integrity. How this activity is regulated and controlled, particularly through selective pressures, to maintain such large cassette arrays is unknown. Here we show a key role of promoter-containing toxin–antitoxin (TA) cassettes as abortive systems that kill the cell when the overall cassette excision rate is too high. These results highlight the importance of TA cassettes regulating the cassette recombination dynamics and provide insight into the evolution and success of integrons in bacterial genomes.

Teaser

The accumulation of cassette functions in integrons is ensured by toxin–antitoxin systems which kill the cell when the cassette excision rate is too high.
Genome-wide single cell annotation of the human protein-coding genes
Authors: Karlsson Max , Álvez María Bueno , Shi Mengnan , Méar Loren , Schutten Rutger , Hikmet Feria , Digre Andreas , Katona Borbala , Vuu Jimmy , Bosic Martina , Sjöstedt Evelina , Edfors Fredrik , Oksvold Per , von Feilitzen Kalle , Zwahlen Martin , Forsberg Mattias , Johansson Fredric , Mulder Jan , Hökfelt Tomas , Luo Younglun , Butler Lynn , Zhong Wen , Mardinoglu Adil , Sivertsson Åsa , Ponten Fredrik , Zhang Cheng , Lindskog Cecilia , Fagerberg Linn , Uhlén Mathias
Published: August 2022

An important quest for the life science community is to deliver a complete annotation of the human building-blocks of life, the genes and the proteins. Here, we report on a genome-wide effort to annotate all protein-coding genes based on single cell transcriptomics data representing all major tissues and organs in the human body, integrated with data from bulk transcriptomics and antibody-based tissue profiling. Altogether, 25 tissues have been analyzed with single cell transcriptomics resulting in genome-wide expression in 444 single cell types using a strategy involving pooling data from individual cells to obtain genome-wide expression profiles of individual cell type. We introduce a new genome-wide classification tool based on clustering of similar expression profiles across single cell types, which can be visualized using dimensional reduction maps (UMAP). The clustering classification is integrated with a new “tau” score classification for all protein-coding genes, resulting in a measure of single cell specificity across all cell types for all individual genes. The analysis has allowed us to annotate all human protein-coding genes with regards to function and spatial distribution across individual cell types across all major tissues and organs in the human body. A new version of the open access Human Protein Atlas (www.proteinatlas.org) has been launched to enable researchers to explore the new genome-wide annotation on an individual gene level.
Genome-wide analysis of lncRNAs points to their roles in the modulation of developmental regulator expression during plant male germline development
Authors: Lohani Neeta , Golicz Agnieszka A. , Allu Annapurna D. , Bhalla Prem L. , Singh Mohan B.
Published: August 2022

LncRNAs can function in regulating of gene expression, but their roles as essential regulators of developmental processes and organismal phenotypes remain largely unclear. Especially the roles of lncRNAs in plants are largely unexplored. However, it has been proposed that plant lncRNAs act as regulators of protein-coding genes during development and that the similar roles of animal and plant lncRNAs result from convergent evolution. Since pollen development follows an established program with well-defined and characterized stages, we have used it as a model for studying plant lncRNAs and their roles in reproductive development. We investigated of lncRNA expression and function during pollen formation in field mustard (Brassica rapa). Reference-based transcriptome assembly performed to update the existing genome annotation identified novel expressed protein-coding genes and long non-coding RNAs (lncRNAs), including 4,347 long intergenic non-coding RNAs (lincRNAs, 1058 expressed) and 2,045 lncRNAs overlapping protein-coding genes on the opposite strand (lncNATs, 780 expressed). The analysis of expression profiles reveals that lncRNAs are significant and stage-specific contributors to the gene expression profile of developing pollen. Gene co-expression networks accompanied by genome location analysis identified 38 cis-acting lincRNA, 31 cis-acting lncNAT, 7 trans-acting lincRNA and 14 trans-acting lncNAT to be substantially co-expressed with target protein-coding genes involved in biological processes regulating pollen development and male lineage specification. These findings provide a foundation for future research aiming at developing strategies to employ lncRNAs as regulatory tools for gene expression control during reproductive development.
Dynamic attention signaling in V4: relation to excitatory/inhibitory cell class and population coupling
Authors: Sachse Elizabeth M. , Snyder Adam C.
Published: August 2022

Neurons have different roles in attention processing. These roles are determined by a neuron’s firing properties, neurotransmitter expression, and functional connectivity. Neurons in the visual cortical area, V4, are reliably engaged by selective attention but exhibit differences in firing rate and correlated variability. It remains unclear what specific neuronal properties shape these attention effects. We identified neurons as fast-spiking (FS) (putative inhibitory) and regular-spiking (RS) (putative excitatory) and investigated their role in anticipatory attention and how this related to their functional connectivity. V4 neurons exhibited a continuum of time-varying attention effects ranging from “restless-weak” neurons at one extreme to “quiet-strong” neurons at another. We found an interdependence between neural attention effects (e.g., restless-weak or quiet-strong), neuron type (FS, RS), and functional connectivity. In particular, we found neurons with restless-weak attention effects were more likely to be RS and have greater population coupling, compared to neurons with other types of attention effects. Also, quiet-strong neurons were more likely to be FS and these FS neurons exhibited higher spike synchrony. From this we propose that time-varying attention effects in a neuronal population depends on the relative involvement of neurons that drive stimulus processing and those that are engaged by intrinsic population activity. These results add important information to our understanding of visual attention circuits at the cellular level.
Differential patterns of taxonomic and functional diversity for two groups of canopy arthropods across spatial scales
Authors: Mahon Michael B. , Penn Hannah J. , Campbell Kaitlin U. , Crist Thomas O.
Published: August 2022

Aim

Arthropod diversity is often linked to variation in resource use, dispersal ability, habitat connectivity, and climate factors that differ across spatial scales. The aim of this research was to examine how species richness, functional diversity, and community composition of two taxa differing in functional roles and dispersal ability are structured across spatial scales and to identify the importance of vegetation, climate, and landscape in explaining these patterns at different scales.

Location

96 trees in 24 stands of 6 deciduous forest sites in 2 ecoregions of the eastern USA (North-Central Till Plain and Western Allegheny Plateau)

Time period

2000

Major taxa studied

Canopy dwelling ants (Hymenoptera: Formicidae) and spiders (Araneae)

Methods

Organisms were collected from tree canopies using insecticidal fogging. Ant and spider taxonomic and functional beta diversity were partitioned across four hierarchical spatial scales (individual tree, forest stand, site, and ecoregion). The contribution of climactic, landscape, and vegetation variables was determined using model selection.

Results

Ant and spider species richness, functional diversity, and community composition differed between taxa and across spatial scales. Alpha diversity (within trees) was lower than expected for both taxa and types of diversity, with host tree species supporting different species of ants and spiders. While beta components of species diversity among trees and forest stands was greater than expected for both taxa, spiders also showed significant levels of beta diversity among sites. Functional beta diversity was less scale-dependent than taxonomic beta diversity. Stand-level patterns of beta diversity were significantly predicted by variation in climate and landscape connectivity.

Main conclusions

Effects of climate and landscape fragmentation on the diversity and community structure of both taxa indicate that anthropogenic climate change and land use change will alter canopy arthropod communities. Results also suggest that patterns of diversity among fragmentation metrics is influenced by differences in dispersal ability.
Neuronal Maturation-dependent Nano-Neuro Interaction and Modulation
Authors: Gupta Prashant , Rathi Priya , Gupta Rohit , Baldi Harsh , Coquerel Quentin , Debnath Avishek , Derami Hamed Gholami , Raman Baranidharan , Singamaneni Srikanth
Published: August 2022

Nanotechnology-enabled neuromodulation, a rapidly growing technique, is a promising minimally-invasive tool in neuroscience and engineering for both fundamental studies as well as clinical applications. However, the nano-neuro interactions at different stages of maturation of a neural network and its implications on the nano-neuromodulation remain unclear. Here, we report heterogeneous to homogenous transformation of neuromodulation in a progressively maturing neural network. Utilizing plasmonic fluors as ultrabright fluorescent nanolabels, we reveal that negative surface charge of the nanoparticles renders selective nano-neuro interaction with a strong correlation between the maturation stage of the individual neurons in the neural network and the density of the nanoparticles bound on the neurons. In stark contrast to homogeneous neuromodulation in a mature neural network reported so far, the maturation-dependent density of the nanoparticles bound to neurons in a developing neural network resulted in a heterogeneous optical neuromodulation (i.e., simultaneous excitation and inhibition of neural network activity). This study advances our understanding of nano-neuro interactions and nano-neuromodulation with potential applications in minimally-invasive technologies for treating neuronal disorders in parts of mammalian brain where neurogenesis persists throughout aging.
Methanol fixation is the method of choice for droplet-based single-cell transcriptomics of neural cells
Authors: Gutiérrez-Franco Ana , Hassan Mohamed N. , Mularoni Loris , Plass Mireya
Published: August 2022

Single-cell transcriptomics methods have become very popular to study the cellular composition of organs and tissues and characterize the expression profiles of the individual cells that compose them. The main critical step in single-cell transcriptomics is sample preparation. Several methods have been developed to preserve cells after sample dissociation to uncouple sample handling from library preparation. Yet, the suitability of these methods depends on the types of cells to be processed. In this project, we perform a systematic comparison of preservation methods for droplet-based single-cell RNA-seq (scRNA-seq) on neural and glial cells derived from induced pluripotent stem cells (iPSCs) and highlight their strengths and weaknesses. We compared the cellular composition and expression profile of single-cell suspensions from fresh NPCs with that of NPCs preserved with Dimethyl Sulfoxide (DMSO), Methanol, vivoPHIX and Acetil-methanol (ACME). Our results show that while DMSO provides the highest cell quality in terms of RNA molecules and genes detected per cell, it strongly affects the cellular composition and the expression profile of the resulting datasets. In contrast, methanol fixed samples display a cellular composition like that of fresh samples while providing a good cell quality and smaller expression biases. Taken together, our results show that methanol fixation is the method of choice for performing droplet-based single-cell transcriptomics experiments on neural cell populations.
A Novel Hyper-Variable Variable Number Tandem Repeat in the Dopamine Transporter Gene (SLC6A3)
Authors: Apsley Abner T. , Domico Emma R. , Verbiest Max A. , Brogan Carly A. , Buck Evan R. , Burich Andrew J. , Cardone Kathleen M. , Stone Wesley J. , Anisimova Maria , Vandenbergh David J.
Published: August 2022

The dopamine transporter gene,SLC6A3, has received substantial attention in genetic association studies of various phenotypes. Although some variable number tandem repeats (VNTRs) present inSLC6A3have been tested in genetic association studies, results have not been consistent. VNTRs inSLC6A3that have not been examined genetically were characterized. Tandem Repeat Annotation Library (TRAL) was used to characterize the VNTRs of 64 unrelated long-read haplotype-phasedSLC6A3sequences. Sequence similarity of each repeat unit of the five VNTRs is reported, along with the correlations of SNP-SNP, SNP-VNTR and VNTR-VNTR alleles across the gene. One of these VNTRs is a novel hyper-VNTR (hyVNTR) in intron 8 ofSLC6A3, which contains a range of 3.4-133.4 repeat copies and has a consensus sequence length of 38bp, with 82% G+C content. The 38-base repeat was predicted to form G-quadruplexesin silicoand was confirmed by circular dichroism spectroscopy. Additionally, this hyVNTR contains multiple putative binding sites for PRDM9, which, in combination with low levels of linkage disequilibrium around the hyVNTR, suggests it might be a recombination hotspot.

Summary Blurb

This VNTR has a heterozygosity value of 0.93, forms G-tetrads, and is in low linkage disequilibrium with surrounding sequence, making it a new site for genetic analysis.
Mechanistic dissection of antibody inhibition of influenza entry yields unexpected heterogeneity
Authors: Sengar Anjali , Cervantes Marcos , Kasson Peter M.
Published: August 2022

Neutralizing antibodies against influenza have generally been classified according to their recognition sites, with antibodies against the head domain of hemagglutinin thought to inhibit attachment and antibodies against the stalk region thought to inhibit fusion. Here, we report the development of a microfluidic assay to measure neutralization of viral entry that can clearly differentiate between effects on attachment and fusion. Testing multiple broadly-neutralizing antibodies against the hemagglutinin stalk domain, we obtain a surprising result: some broadly-neutralizing antibodies inhibit fusion only, while others inhibit both fusion and viral attachment. Antibodies binding the globular head domain primarily inhibit attachment but can also reduce the fusogenic capability of viral particles that nonetheless bind receptor. These findings shed light on the unexpectedly heterogeneous mechanisms of antibody neutralization even within similar recognition sites. The assay we have developed also provides a tool to optimize vaccine design by permitting assessment of the elicited antibody response with greater mechanistic resolution.
Dynamics of transposable element accumulation in the non-recombining regions of mating-type chromosomes in anther-smut fungi
Authors: Duhamel Marine , Hood Michael E. , Rodríguez de la Vega Ricardo C. , Giraud Tatiana
Published: August 2022

Background

Transposable element (TE) activity is generally deleterious to the host fitness, such that TE copies are often purged by selection, which is facilitated when meiotic recombination reshuffles variation among loci. In the absence of recombination, the number of TE insertions usually increases, but the dynamics of such TE accumulations is unknown.

Results

In this study, we investigated the temporal dynamics of TE accumulation in the non-recombining genomic regions of 15Microbotryumspecies, leveraging on a unique dataset of 21 independent evolutionary strata of recombination cessation of different ages. We show that TEs rapidly accumulated in regions lacking recombination, but that the TE content reached a plateau at ca. 50% of occupied base pairs by 1.5 MY following recombination suppression. The same TE superfamilies have repeatedly expanded in independently evolved non-recombining regions, in particular rolling-circle replication elements (Helitrons), despite being scarce before recombination suppression. The most abundant elements, long-terminal repeat (LTR) retrotransposons of theCopiaandTy3superfamilies, expanded through transposition bursts affecting both the non-recombining regions of mating-type chromosomes and autosomes, thus suggesting that non-recombining regions constitute a reservoir of TEs that transpose to recombining regions. Based on LTR sequence divergence within and among copies, we could distinguish bursts of transposition from gene conversion.

Conclusion

Our study supports the TE reservoir hypothesis, by showing that TE accumulation in non-recombining has a genome-wide impact. TEs accumulated through bursts, and following a non-linear, decelerating dynamics, thus improving our knowledge on genome evolution, particularly in association with sex or mating-type chromosomes.