bluuebunny/crossref_metadata_2025 huggingface dataset:
Auditory cortical responses to speech are shaped by statistical learning of short-term speech input regularities
Authors: Wu Yunan Charles , Viswanathan Vibha , Abel Taylor J. , Holt Lori L.
Published: December 2022

Speech perception presents an exemplary model of how neurobiological systems flexibly adjust when input departs from the norm. Dialects, accents, and even head colds can negatively impact comprehension by shifting speech from listeners’ expectations. Comprehension improves with exposure to shifted speech regularities, but there is no neurobiological model of this rapid learning. We used electroencephalography to examine human auditory cortical responses to utterances that varied only in fundamental frequency (F0, perceived as voice pitch) as we manipulated the statistical distributions of speech acoustics across listening contexts. Participants overtly categorized speech sampled across two acoustic dimensions that signal /b/ from /p/ (voice onset time [VOT] and F0) to model typical English speech regularities or an expectation-violating accent. These blocks were interleaved with passive exposure to two F0-distinguished test stimuli presented in an oddball ratio to elicit a cortical mismatch negativity (MMN) response. F0 robustly influenced speech categorization when short-term regularities aligned with English but F0 exerted no influence in the context of the accent. The short-term regularities modulated event-related potentials evoked by F0-distinguished test stimuli across both N1 and P3 temporal windows and, for P3 amplitude, there was a strong correlation with perceptual down-weighting of F0. The influence of the short-term regularities persisted to impact MMN in interleaved passive listening blocks when regularities mirrored English but were absent when regularities conveyed the accent. Thus, cortical response is modulated as a function of statistical regularities of the listening context, likely reflecting both early dimension encoding and later categorization.

Significance Statement

Speech perception is a quintessential example of how neurobiological systems flexibly adjust when input departs from the norm. Perception is well-tuned to native-language speech patterns. Yet it adjusts when speech diverges from expectations, as with a foreign accent. We observe that the effectiveness of specific cues in speech, like the pitch of a voice, in signaling phonemes like /b/ versus /p/ is dynamically re-weighted when speech violates native-language expectations. We find that this re-weighting is mirrored in cortical responses that reflect both early acoustic dimension encoding and also in later responses linked to phoneme categorization. The results implicate dynamic adjustments in the mapping of speech to cortical representations, as modulated by statistical regularities experienced across local speech input.
Decoding Neural Patterns for the Processing of Fearful Faces under Different Visual Awareness Conditions: A Multivariate Pattern Analysis
Authors: Qiu Zeguo , Li Xuqian , Pegna Alan J.
Published: December 2022

Mixed findings have been reported for the nonconscious processing of fearful faces. Here, we used multivariate pattern analysis on electroencephalography data from three backward masking experiments to decode the conscious and nonconscious processing of fearful faces. Three groups of participants were shown pairs of faces that were presented either subliminally (16 ms) or supraliminally (266 ms) and were required to complete tasks where the face stimuli were either task-relevant (Experiment 1) or task-irrelevant (Experiments 2 and 3). We decoded the neural activity to examine the temporal dynamics of visual awareness, and to investigate whether the presence and location of a fearful face were processed when levels of awareness varied. The results reveal that the spatial location of fearful faces can be decoded from neural patterns only when they are consciously seen and relevant to participants’ task. Nevertheless, the processing of the mere presence of fearful faces can occur in the absence of visual awareness, and the related neural patterns can be generalised to the conscious, non-spatial processing of fearful faces. Additionally, the flexibility of spatial attention seems to modulate the processing of fearful faces.
Capturing the biology of disease severity in an organoid model of LIS1-lissencephaly
Authors: Zillich Lea , Gasparotto Matteo , Rossetti Andrea Carlo , Fechtner Olivia , Maillard Camille , Hoffrichter Anne , Zillich Eric , Jabali Ammar , Marsoner Fabio , Artioli Annasara , Wilkens Ruven , Schroeter Christina B. , Hentschel Andreas , Witt Stephanie H. , Melzer Nico , Meuth Sven G. , Ruck Tobias , Koch Philipp , Roos Andreas , Bahi-Buisson Nadia , Francis Fiona , Ladewig Julia
Published: December 2022

Lissencephaly is a malformation of cortical development (MCD) characterized by reduced to absent gyri and a disorganized cortex, leading to severe neurological consequences in affected individuals, including epilepsy, intellectual disability, and reduced life expectancy. Treatments are purely symptomatic, and patients often remain refractory to them. Heterozygous mutations in theLIS1gene, encoding a regulator of the microtubule motor dynein, cause LIS1-lissencephaly. For unknown reasons, LIS1-lissencephaly patients show marked differences in disease severity despite each carrying a heterozygous LIS1 mutation. We leveraged forebrain-type organoids from patients diagnosed with mild, moderate, or severe LIS1-lissencephaly to investigate, in a cytoarchitecture and multi-omics approach, disease and severity grade associated phenotypes, mechanisms, and rescue approaches. We identified alterations of the cytoarchitecture, progenitor cell homeostasis, and neurogenesis often with a severity-dependent gradient. Identified disease-linked molecular mechanisms were microtubule destabilization, WNT-signaling, protein metabolism, and perturbed cadherin- and unfolded protein-binding. Some mechanisms exhibited a severity-dependent gradient or were specific to a severe grade. We present strategies to reverse phenotypic changes in LIS1-patient organoids and identify mTOR pathway inhibitors inin silicodrug repurposing analysis as potential novel therapeutic strategy. By probing the top hit drug, the mTOR inhibitor everolimus, we could indeed rescue severity-dependent phenotypic changes in LIS1-patient organoids. This study demonstrates that organoid-based modeling is sensitive in recapitulating disease severity, which presents an important step in patient stratification, and allows the development of novel personalized rescue strategies with therapeutic potential.
FAP-retargeted Ad5 enablesin vivogene delivery to stromal cells in the tumor microenvironment
Authors: Hartmann K. Patricia , van Gogh Merel , Freitag Patrick C. , Kast Florian , Nagy-Davidescu Gabriela , Borsig Lubor , Plückthun Andreas
Published: December 2022

Fibroblast activation protein (FAP) is a cell surface serine protease that is highly expressed on reactive stromal fibroblasts, such as cancer-associated fibroblasts (CAFs), and generally absent in healthy adult tissues. FAP expression in the tumor stroma has been detected in more than 90% of all carcinomas, rendering CAFs excellent target cells for a tumor site-specific adenoviral delivery of cancer therapeutics. Here, we present a tropism-modified human adenovirus 5 (Ad5) vector that targets FAP through trivalent, designed ankyrin repeat protein (DARPin)-based retargeting adapters. We describe the development and validation of these adapters via cell-based screening assays and demonstrate adapter-mediated Ad5 retargeting to FAP+fibroblastsin vitroandin vivo. We further show efficientin vivodelivery andin-situproduction of a therapeutic payload by CAFs in the tumor microenvironment (TME), resulting in attenuated tumor growth. We thus propose using our FAP-Ad5 vector to convert CAFs into a ‘biofactory’, secreting encoded cancer therapeutics into the TME to enable a safe and effective cancer treatment.
A duration-dependent interaction between high-intensity light and optical refocus in the drive for myopia control
Authors: Biswas Sayantan , Muralidharan Arumugam R. , Betzler Bjorn Kaijun , Fianza Busoy Joanna Marie , Barathi Veluchamy A. , Tan Royston K. Y. , Shermaine Low Wan Yu , Milea Dan , Kathrani Biten K. , Brennan Noel A. , Najjar Raymond P.
Published: December 2022

PURPOSE

To evaluate the duration-dependent and synergetic impact of high-intensity light (HL) and optical refocus (RF) on lens-induced myopia (LIM) development in chickens.

METHODS

Myopia was induced in one eye in chicks (10 groups, n=126) from day 1 post- hatching (D1) until D8 using -10D lenses. Fellow eyes remained uncovered as controls. Nine groups were exposed daily to continuous 2 hours (h), 4h, or 6h of either HL (15,000 lux); RF (removal of -10D lens); or both (HL+RF). One group served as the LIM group without any interventions. Ocular axial length (AL), refractive error, and choroidal thickness were measured on D1, D4, and D8. Outcome measures are expressed as inter-ocular difference (IOD= experimental - control eye) ±SEM.

RESULTS

By D8, LIM increased AL (0.36±0.04 mm), myopic refraction (-9.02±0.37D), and choroidal thinning (-90.27±16.44 µm) in the LIM group (all, P<0.001). Compared to the LIM group, exposure to 2h, 4h, or 6h of HL, RF, or HL+RF reduced myopic refraction in a duration-dependent manner, with RF being more effective than HL (P<0.05). Only 6h of HL+RF (not 2h or 4h) prevented LIM and was more effective than RF (P=0.004) or HL (P<0.001) in reducing myopic refraction, and more effective than HL (P<0.001) in reducing axial elongation.

CONCLUSION

Daily exposure to 2h, 4h, or 6h of HL, RF, or HL+RF reduced lens-induced myopic refraction in a duration-dependent manner in chickens. Only 6h of HL+RF completely stopped LIM development. The synergetic effect of HL and RF is dependent on the duration of the interventions.
Sensitivity of human sweet taste receptor subunits T1R2 and T1R3 to activation by glucose enantiomers
Authors: Dubovski Nitzan , Galezcki Yaron Ben-Shoshan , Malach Einav , Niv Masha Y.
Published: December 2022

We have previously shown that L-glucose, the non-caloric enantiomer of D-glucose, activates the human sweet taste receptor T1R2/T1R3 transiently expressed in HEK293T cells. Here we show that D- and L-glucose can also activate T1R2 and T1R3 expressed without the counterpart monomer. Serine mutation to alanine in residue 147 in the binding site of T1R3 VFT domain, completely abolishes T1R3S147A activation by either L or D-glucose, while T1R2/T1R3S147A responds in the same way as T1R2 expressed without its counterpart. We further show that the original T1R2 reference sequence (NM_152232.1) is less sensitive by almost an order of magnitude than reference sequence at the time this study was performed (NM_152232.4). We find that out of the four differing positions, it is the R317G in the VFT domain of T1R2, that is responsible for this effect in-vitro. It is important both for practical assay sensitivity, and because glycine is found in this position in ∼20% of the world population. While the effects of the mutations and of the partial transfections were similar for D and L enantiomers, their dose response curves remained distinct, with L-glucose reaching an early plateau.
Multi-modal and multi-model interrogation of large-scale functional brain networks
Authors: Castaldo Francesca , Páscoa dos Santos Francisco , Timms Ryan C , Cabral Joana , Vohryzek Jakub , Deco Gustavo , Woolrich Mark , Friston Karl , Verschure Paul , Litvak Vladimir
Published: December 2022

Current whole-brain models are generally tailored to the modelling of a particular modality of data (e.g., fMRI or MEG/EEG). Although different imaging modalities reflect different aspects of neural activity, we hypothesise that this activity arises from common network dynamics. Building on the universal principles of self-organising delay-coupled nonlinear systems, we aim to link distinct electromagnetic and metabolic features of brain activity to the dynamics on the brain’s macroscopic structural connectome.

To jointly predict dynamical and functional connectivity features of distinct signal modalities, we consider two large-scale models generating local short-lived 40 Hz oscillations with various degrees of realism - namely Stuart Landau (SL) and Wilson and Cowan (WC) models. To this end, we measure features of functional connectivity and metastable oscillatory modes (MOMs) in fMRI and MEG signals - and compare them against simulated data.

We show that both models can represent MEG functional connectivity (FC) and functional connectivity dynamics (FCD) to a comparable degree, by varying global coupling and mean conduction time delay. For both models, the omission of delays dramatically decreased the performance. For fMRI, the SL model performed worse for FCD, highlighting the importance of balanced dynamics for the emergence of spatiotemporal patterns of ultra-slow dynamics. Notably, optimal working points varied across modalities and no model was able to achieve a correlation with empirical FC higher than 0.45 across modalities for the same set of parameters. Nonetheless, both displayed the emergence of FC patterns beyond the anatomical framework. Finally, we show that both models can generate MOMs with empirical-like properties.

Our results demonstrate the emergence of static and dynamic properties of neural activity at different timescales from networks of delay-coupled oscillators at 40 Hz. Given the higher dependence of simulated FC on the underlying structural connectivity, we suggest that mesoscale heterogeneities in neural circuitry may be critical for the emergence of parallel cross-modal functional networks and should be accounted for in future modelling endeavours.
Differential occupancy and regulatory interactions of KDM6A in bladder cell lines
Authors: Özden-Yılmaz Gülden , Savaş Büşra , Bursalı Ahmet , Eray Aleyna , Arıbaş Alirıza , Şentürk Şerif , Karaca Ezgi , Karakülah Gökhan , Erkek-Ozhan Serap
Published: December 2022

Epigenetic deregulation is a critical theme which needs further investigation for bladder cancer research. One of the highly mutated genes in bladder cancer isKDM6A, functioning as a H3K27 demethylase and is part of the MLL3/4 complexes. To decipher the role of KDM6A in normal versus tumor setting, we identified the genomic landscape of KDM6A in normal, immortalized and cancer bladder cells. Our results showed differential KDM6A occupancy at the genes involved in cell differentiation, chromatin organization and Notch signaling depending on the cell type and the mutation status ofKDM6A. Transcription factor motif analysis revealed HES1 to be enriched at KDM6A peaks identified for T24 bladder cancer cell line, which has a truncating mutation in KDM6A, lacking demethylase domain. Our co-immunoprecipitation experiments reveal TLE co-repressors and HES1 as potential truncated and wild type KDM6A interactors. With the aid of structural modeling, we explored how the truncated KDM6A could interact with TLE, HES1, as well RUNX, HHEX transcription factors. These structures provide a solid mean to study the functions of KDM6A independent of its demethylase activity. Collectively, our work provides important contributions to the understanding of KDM6A malfunction in bladder cancer.
Deletion of the transcriptional regulator TFAP4 accelerates c-MYC-driven lymphomagenesis
Authors: Potts Margaret A , Mizutani Shinsuke , Garnham Alexandra L , Li Wai Suen Connie S N , Kueh Andrew J , Tai Lin , Pal Martin , Strasser Andreas , Herold Marco J
Published: December 2022

Many lymphoid malignancies arise from deregulated c-MYC expression in cooperation with additional genetic lesions. While many of these cooperative genetic lesions have been discovered, DNA sequence data suggest that many more do exist. However, their contributions to c-MYC driven lymphomagenesis have not yet been investigated. We identified TFAP4 as a potent suppressor of c-MYC driven lymphoma development in a previous genome-wide CRISPR knockout screen in primary cellsin vivo(Mizutani et al, 2022). CRISPR deletion of TFAP4 inEμ-MYCtransgenic hematopoietic stem and progenitor cells (HSPCs) significantly accelerated c-MYC-driven lymphoma development in mice. TFAP4 deficientEμ-MYClymphomas all arose at the pre-B cell stage. Characterization of the transcriptional profile of pre-leukemic pre-B cells inEμ-MYC/Cas9/sgTFAP4transplanted mice, revealed that TFAP4 deletion reduced expression of several master regulators of B cell differentiation, such asSpi1, SpiBandPax5, which all have been shown to be bound by TFAP4. We therefore conclude that loss of TFAP4 leads to a block in differentiation during early B cell development, causing accelerated c-MYC-driven lymphoma development.
Differential response of patient-derived primary glioblastoma cells to metabolic and adhesion inhibitors
Authors: Rezk Rasha , Basar Fikret , Mediavillo John , Donaldson Rebecca , Watts Colin , Franze Kristian , Kabla Alexandre J
Published: December 2022

Purpose

This study aims to investigate Glioblastoma (GBM) cellular response to adhesion and metabolic inhibitors in the context of cells’ migration and cell-matrix adhesion properties. GBM is the most common incurable brain tumour. Decades of work into GBM chemical and molecular classification have identified mechanisms of drug resistance. Inhibitors targeting cancer cell migration and proliferation rarely take into consideration the heterogeneous migration property amongst cells, which may impact patients’ response to treatment.

Methods

Tissue samples were obtained from spatially distinct locations with different 5-aminolevulinic acid (5-ALA) fluorescent intensities, strong strongly fluorescent tumour cores, a weak fluorescent tumour rim, and nonfluorescent tumour margins. Samples were previously shown to be associated with different motility and adhesion properties. We tested tumour cells’ response to adhesion and metabolic inhibitors using metabolic assays. Cell survival was also monitored using time-lapse microscopy, while cultured on low-modulus polydimethylsiloxane representative of the stiffness of brain tissue.

Results

Metabolic viability assays, MTT and Cell Titer, showed substantial heterogeneity in drug potency. Highly fluorescent tumour core cells were significantly more resistant to an F0F1 ATP synthase inhibitor (Gboxin), and a FAK inhibitor (GSK2256098), and cell proliferation ceased post-treatment in vitro. Cells derived from non-fluorescent tumour margins exhibited higher potency for the ATP synthase inhibitor (Gboxin). However, cell proliferation persisted post-treatment.

Conclusion

Our study suggests that the adhesive and migration properties of cells account for the sensitivity to therapeutics in different regions of the tumour in individual patients and between patients with GBM.
How Myopia and Glaucoma Influence the Biomechanical Susceptibility of the Optic Nerve Head
Authors: Chuangsuwanich Thanadet , Tun Tin A. , Braeu Fabian A. , Yeoh Clarice H.Y. , Chong Rachel S. , Wang Xiaofei , Aung Tin , Hoang Quan V. , Girard Michaël J.A.
Published: December 2022

Purpose

We aimed to assess optic nerve head (ONH) deformations following acute intraocular pressure (IOP) elevations and horizontal eye movements (adduction and abduction) in control eyes, highly myopic (HM) eyes, HM eyes with glaucoma (HMG), and eyes with pathologic myopia alone (PM) or PM with staphyloma (PM+S).

Methods

We studied 282 eyes, comprising of 99 controls, 51 HM, 35 HMG, 21 PM and 75 PM+S eyes. For each eye, we imaged the ONH using spectral-domain optical coherence tomography (OCT) under the following conditions:(1)primary gaze,(2)20° adduction,(3)20° abduction and(4)primary gaze with acute IOP elevation (to ~35 mmHg) achieved through ophthalmodynamometry. For each OCT volume, we automatically segmented the ONH tissues using deep learning. We performed digital volume correlation (DVC) analysis to compute IOP- and gaze-induced ONH displacements and effective strains (i.e. local deformations). All biomechanical quantities were compared across groups.

Results

Under IOP elevation, we found that HM eyes exhibited significantly lower strains (3.9 ± 2.4 %) than PM eyes (6.9 ± 5.0%, p < 0.001), HMG eyes (4.7 ± 1.8%, p = 0.04) and PM+S eyes (7.0 ± 5.2%, p < 0.001). Under adduction, we found that HM eyes exhibited significantly lower strains (4.8% ± 2.7%) than PM+S eyes (6.0 ± 3.1%, p = 0.02). We also found significant associations between axial length (or refractive error) and strains - eyes with higher axial length and greater myopia were associated with higher strains. IOP-induced strains were also positively correlated with adduction-induced strains.

Conclusion

We found that HMG eyes experienced significantly higher strains under IOP elevations as compared to HM eyes. Additionally, PM+S eyes experienced highest ONH strains as compared to other groups under all biomechanical loads. Our preliminary findings suggest the possibility of using a simple biomechanical test to tease out the susceptibility of HM eyes to further develop glaucoma and/or staphyloma.
Ghost admixture in eastern gorillas
Authors: Pawar Harvinder , Rymbekova Aigerim , Cuadros Sebastian , Huang Xin , de Manuel Marc , van der Valk Tom , Lobon Irene , Alvarez-Estape Marina , Haber Marc , Dolgova Olga , Han Sojung , Esteller-Cucala Paula , Juan David , Ayub Qasim , Bautista Ruben , Kelley Joanna L. , Cornejo Omar E. , Lao Oscar , Andrés Aida M. , Guschanski Katerina , Ssebide Benard , Cranfield Mike , Tyler-Smith Chris , Xue Yali , Prado-Martinez Javier , Marques-Bonet Tomas , Kuhlwilm Martin
Published: December 2022

Archaic admixture has had a significant impact on human evolution with multiple events across different clades, including from extinct hominins such as Neanderthals and Denisovans into modern humans. Within the great apes archaic admixture has been identified in chimpanzees and bonobos, but the possibility of such events has not been explored in other species. Here, we address this question using high-coverage whole genome sequences from all four extant gorilla subspecies, including six newly sequenced eastern gorillas from previously unsampled geographic regions. Using Approximate Bayesian Computation (ABC) with neural networks to model the demographic history of gorillas, we find a signature of admixture from an archaic ‘ghost’ lineage into the common ancestor of eastern gorillas, but not western gorillas. We infer that up to 3% of the genome of these individuals is introgressed from an archaic lineage that diverged more than 3 million years ago from the common ancestor of all extant gorillas. This introgression event took place before the split of mountain and eastern lowland gorillas, likely more than 40 thousand years ago, and may have influenced perception of bitter taste in eastern gorillas. When comparing the introgression landscapes of gorillas, humans and bonobos, we find a consistent depletion of introgressed fragments on the X chromosome across these species. However, depletion in protein-coding content is not detectable in eastern gorillas, possibly as a consequence of stronger genetic drift in this species.
The K2: Open-source simultaneous triple-color TIRF microscope for live-cell and single-molecule imaging
Authors: Niederauer Christian , Seynen Marco , Zomerdijk Jan , Kamp Marko , Ganzinger Kristina A.
Published: December 2022

Imaging the dynamics and interactions of biomolecules at the single-molecule level in live cells and reconstituted systems has generated unprecedented knowledge about the biomolecular processes underlying many cellular functions. To achieve the speed and sensitivity needed to detect and follow individual molecules, these experiments typically require custom-built microscopes or custom modifications of commercial systems. The costs of such single-molecule microscopes, their technical complexity and the lack of open-source documentation on how to build custom setups therefore limit the accessibility of single-molecule imaging techniques. To advance the adaptation of dynamic single-molecule imaging by a wider community, we present the “K2”: an open-source, simultaneous triple-color total internal reflection fluorescence (TIRF) microscope specifically designed for live-cell and single-molecule imaging. We explain our design considerations and provide step-by-step building instructions, parts list and full CAD models. The flexible design of this TIRF microscope allows users to customize it to their scientific and financial needs, or to re-use parts of our design to improve the capabilities of their existing setups without necessarily having to build a full copy of the K2 microscope.
Non-equilibrium processes prevail in shaping species richness and functional diversity of terrestrial vertebrates in a global hotspot
Authors: Moroti Matheus de Toledo , Skeels Alexander , Da Silva Fernando R. , Provete Diogo B.
Published: December 2022

Aim

The effects of equilibrium and nonequilibrium processes are generally investigated using species richness on a single biological group. However, little is known about how these two classes of processes also affect trait diversity, considering multiple taxa within the same geographical template. Here, we evaluated which variables representing equilibrium (topography, climate, and primary productivity) and nonequilibrium (diversification rate and evolutionary time) processes best explain species richness and trait diversity of four clades of vertebrates within the same global hotspot. We also investigated how trait disparity has accumulated over time and whether there are congruent spatial patterns between groups.

Location

Atlantic Rainforest

Time period

Contemporary.

Major taxa studied

Terrestrial vertebrates.

Methods

We tested whether the spatial pattern of Functional Dispersion (FDis), richness, diversification rate, and evolutionary time of each group are correlated. We used a spatially explicit structural equation model to test how species richness and functional dispersion are influenced by variables representing equilibrium and nonequilibrium mechanisms. Additionally, we explored how trait disparity accumulated over time in the four groups.

Results

We found that non-equilibrium proxies, evolutionary time and diversification rate, played a primary role in driving species richness and trait diversity, with elevation and climate variables having only an indirect effect species and trait diversity via diversification rate and evolutionary time. We found a congruent pattern of species richness among all groups, except among ectotherms. In contrast, the spatial distribution of evolutionary time was distinct for each group.

Main conclusions

Despite nonequilibrium processes were more important for generating large-scale diversity patterns within the same geographical template, the interplay between evolutionary time and dispersal ability have disparately determined the assembly of communities.
HBM-CITEseq: a uniform CITE-seq processing pipeline for the HuBMAP Consortium
Authors: Lu Xinyue , Ruffalo Matthew
Published: December 2022

As part of the HuBMAP Consortium we have been developing methods for uniformly processing and indexing multiple single-cell datasets, which enable efficient integration of data from different platforms. HuBMAP computational pipelines have so far focused on unimodal data types such as single-cell/nucleus RNA sequencing and single nucleus ATAC-seq. Here we present HBM-CITEseq, a processing pipeline for Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) datasets, the first multimodal sequencing processing pipeline for the HuBMAP Consortium, with transcriptomic outputs from HBM-CITEseq matching those of the HuBMAP RNA-seq pipeline.

HBM-CITEseq is a CWL workflow wrapping command-line tools encapsulated in Docker images. It is freely available on GitHub athttps://github.com/hubmapconsortium/citeseq-pipeline.
Different B cell activation patterns in asymptomatic and symptomatic COVID-19 patients
Authors: Pham Nhung , Talih Nuray , Ehrhart Friederike , Evelo Chris T , Kutmon Martina
Published: December 2022

Early and persistent defects in B cell subsets such as memory B cells were shown to be correlated with poor outcomes in COVID-19 patients. This research aimed to develop a molecular pathway model to understand the B cell development in COVID-19. A B cell transcriptomics dataset, obtained from COVID-19 patients, was analyzed on the resulting pathway model to study B cell activation. The pathway showed two distinct gene expression profiles between asymptomatic and symptomatic patients. In asymptomatic patients, there is an increase in transcript levels of antiviral interferon-stimulated genes such as ISG15, IFITM1, and NEAT1 and a driving gene for the extrafollicular pathway CXCR4 indicating a formation of plasmablast. In symptomatic patients, the results suggest an inhibition occurring at the germinal center hinting at a reduction in memory B cell production. Transcripts of driver gene CXCR5 involved in germinal center development is one of the most downregulated genes. This could contribute to the shortage in the formation of memory B cells in COVID-19. Concluding, in SARS-CoV-2 infection, B cells follow different activation routes in asymptomatic and symptomatic patients. In this study, we constructed a pathway that allowed us to analyze and interpret activation patterns of B cells in COVID-19 patients and their link to disease severity. Importantly, the pathway and approach can be reused for further research in COVID-19 or other diseases.
Genomic diversity ofStreptococcus uberisisolated from clinical mastitis of cattle in selected areas of Bangladesh
Authors: Hassan Jayedul , Bag Md. Abdus Sattar , Kabir Ajran , Ali Md. Wohab , Hossain Maqsud , Rahman Md. Tanvir , Islam Md. Shafiqul , Khan Md. Shahidur Rahman
Published: December 2022

Streptococci are the major etiology in mastitis, a cause of huge economic losses in the dairy industries.Streptococcus(S.)agalactiae, S. dysagalactiaeandS. uberisare mostly encountered in bovine mastitis; however, data on the diversity and characteristics of Streptococcus in clinical mastitis of cattle in Bangladesh is lacking. Thus, the present study was aimed to determine the diversity and antimicrobial resistance pattern ofStreptococcusspp. isolated from clinical mastitis of cattle reared in Bangladesh. A total of 105 milk samples comprising eighty (80) from cattle with clinical mastitis (CCM) and twenty-five (25) from apparently healthy cattle (AHC) in four prominent dairy farms and one dairy community were purposively collected and examined in this study. Milk samples were enriched in Luria Bertani broth (LB) andStreptococcusspp. was isolated on Modified Edwards Medium and identified by 16S rRNA gene sequencing. Among eighty (80) clinical samples, eighteen (18) were positive forStreptococcusspp. while none of the milk from AHC revealed Streptococcus by cultural and molecular examination. Sequencing and phylogenetic analysis identified 55.6%, 33.3%, 5.6% and 5.6% of the Streptococcus isolates asS. uberis, S. agalactiae, S. hyovaginalisandS. urinalis, respectively. Antibiotic sensitivity testing with antimicrobials commonly used to treat clinical mastitis revealed 100%, 100%, and 30% of theS. agalactiae,S. hyovaginalis, andS. uberisas multidrug-resistant, respectively. Molecular characterization through whole genome sequencing of five (5)S. uberisisolates identified at least two novel ST types ofS. uberiscirculating in the study areas with one ST (4/5 isolates) clustered with the isolates from China, India and Thailand, and the other (1/5) with UK, Ireland and Australia. Pan-genome analysis and phylogeny of the core genome sequences also clustered the isolates into two sub-clusters, indicating the presence of at least two different subtypes ofS. uberisin the study area. On virulence profiling, all the isolates of this study were found to harbor at least 35 virulence and putative virulence genes probably associated with intramammary infection (IMI) indicating all theS. uberisisolated in this study as potential pathogen. From the overall findings it was evident that Strepococcus occurring in bovine mastitis are diverse andS. uberisgenome carries an array of putative virulence factors which need to be investigated genotypically and phenotypically to identify a specific trait or determinant governing the virulence and fitness of this bacterium. Moreover, Streptococcus isolated in this study carried multidrug resistance which needs careful consideration during the selection of a treatment regimen for mastitis.
How to tuna fish: constraint, convergence and integration in the neurocranium of pelagiarian fishes
Authors: Knapp Andrew , Rangel-de Lázaro Gizéh , Goswami Anjali , Friedman Matt , Evans Kory M , Giles Sam , Beckett Hermione T , Johanson Zerina
Published: December 2022

Morphological evolution of the vertebrate skull has been explored across a wide range of tetrapod clades, but teleost fishes, accounting for roughly half of all vertebrate species, have largely been overlooked. Here we present the results of a study investigating three-dimensional morphological evolution across 114 species of Pelagiaria, a morphologically diverse clade of open-ocean teleost fishes that includes tuna and mackerel. Despite showing high shape disparity, the majority of taxa are concentrated in fairly restricted regions of morphospace, with taxa from all families falling into three distinct clusters. Phylogenetic signal in shape data is significant but low (Kmult= 0.27,p= 0.001) and a single-rate Ornstein-Uhlenbeck model of evolution is supported, revealing convergence in shape within and between families. Shape is significantly correlated with body elongation (R2= 0.35,p=0.001), but correlation with size, diet, and habitat depth is weak. Integration of the neurocranium is high, supporting the hypothesis that high integration may promote the evolution of more extreme morphologies. Taken together, these results suggest that shape evolution in the pelagiarian neurocranium is constrained by a number of factors, resulting in the repeated evolution of a restricted range of morphologies.
Hippocampal ripples coincide with “up-state” and cortical spindles in Retrosplenial Cortex
Authors: Pedrosa Rafael , Nazari Mojtaba , Kergoat Loig , Bernard Christophe , Mohajerani Majid , Stella Federico , Battaglia Francesco
Published: December 2022

During NREM sleep hippocampal Sharp-wave ripples (SWR) events are thought to stabilize memory traces for long-term storage in downstream neocortical structures. Within the neocortex, Default Mode Network (DMN) areas interact preferentially with the hippocampus purportedly to consolidate those traces. Transient bouts of slow oscillations and sleep spindles in DMN areas are often observed around SWRs, suggesting that these two activities are related and that their interplay possibly contributes to memory consolidation. To investigate how SWRs interact with the DMN and spindles, we combined cortical wide-field voltage imaging, ECoG, and hippocampal LFP recordings in anesthetized and sleeping mice. Here we show that, during SWR, “up-states” and spindles reliably co-occur in a cortical subnetwork centered around the Retrosplenial cortex. Furthermore, Retrosplenial transient activations and spindles predict Slow Gamma oscillations in CA1 during SWRs. Together, our results suggest that Retrosplenial-hippocampal interaction may be a central source of information exchange between cortex and hippocampus.
Calcium flux through ER-TGN contact sites facilitates cargo export
Authors: Ramazanov Bulat R. , Di Martino Rosaria , Kumar Abhishek , Parchure Anup , Kim Yeongho , Griesbeck Oliver , Schwartz Martin A. , Luini Alberto , von Blume Julia
Published: December 2022

Ca2+influx into the trans-Golgi Network (TGN) promotes secretory cargo sorting by the Ca2+-ATPase SPCA1 and the luminal Ca2+binding protein Cab45. Cab45 oligomerizes upon a local Ca2+influx, and Cab45 oligomers sequester and separate soluble secretory cargo from the bulk flow of proteins in the TGN. However, how this Ca2+flux into the lumen of the TGN is achieved remains elusive, as the cytosol has a very low steady-state Ca2+concentration. The TGN forms membrane contact sites (MCS) with the Endoplasmic Reticulum (ER), whereby the close apposition of the two organelles allows the protein-mediated exchange of molecular species such as lipids. Here we show that the TGN export of Cab45 clients requires the integrity of ER-TGN MCS and IP3R-dependent Ca2+fluxes in the MCS, suggesting Ca2+transfer between these organelles. Using an MCS-targeted Ca2+FRET sensor module, we measure the Ca2+flow in these sites in real-time. These data show for the first time that ER-TGN MCS facilitates Ca2+transfer required for SPCA1-dependent cargo sorting and export from the TGN, thus solving a fundamental question in cell biology.

The current study demonstrates that the trafficking of COMP and LyzC relies on Ca2+flux between the endoplasmic reticulum (ER) and trans-Golgi Network (TGN). This process requires the activity of IP3 receptors, present in ER membranes, and depends on the integrity of the membrane contact site between these two organelles.
Cas phosphorylation regulates focal adhesion assembly
Authors: Kumar Saurav , Stainer Amanda , Dubrulle Julien , Simpkins Christopher , Cooper Jonathan A
Published: December 2022

Integrin-mediated cell attachment rapidly induces tyrosine kinase signaling. Despite years of research, the role of this signaling in integrin activation and focal adhesion assembly is unclear. We provide evidence that the Src-family kinase (SFK) substrate Cas (Crk-associated substrate, p130Cas, BCAR1) is phosphorylated, and associated with its effectors, Crk/CrkL, in clusters that are precursors of focal adhesions. The initial phospho-Cas clusters contain integrin β1 in its inactive, bent closed, conformation. Later, phospho-Cas and total Cas levels decrease as integrin β1 is activated and core focal adhesion proteins including vinculin, talin, kindlin and paxillin are recruited. Cas is required for cell spreading and focal adhesion assembly in epithelial and fibroblast cells on collagen and fibronectin. Cas cluster formation requires Cas, Crk/CrkL, SFKs and Rac1 but not vinculin. Rac1 provides positive feedback onto Cas through reactive oxygen, opposed by negative feedback from the ubiquitin proteasome system. The results suggest a two-step model for focal adhesion assembly in which clusters of phospho-Cas, effectors and inactive integrin β1 grow through positive feedback prior to integrin activation and recruitment of core focal adhesion proteins.
Dynamic Interplay of Autophagy and Membrane Repair DuringMycobacterium tuberculosisInfection
Authors: Augenstreich Jacques , Phan Anna T. , Allen Charles N.S. , Poddar Anushka , Srinivasan Lalitha , Briken Volker
Published: December 2022

Autophagy plays a crucial role in the host response to Mycobacterium tuberculosis (Mtb) infection, yet the dynamics and regulation of autophagy induction on mycobacterial phagosomes remain partially understood. In this study, we employed time-lapse confocal microscopy to investigate in real time the recruitment of LC3B (LC3), a key autophagy marker, to Mtb-containing vacuoles (MCVs) at the single cell level with our newly developed workflow for single cell and single MCV tracking and fluorescence quantification. The results reveal that approximately 70% of MCVs exhibited LC3 recruitment but was lost in about 40% of those MCVs. The LC3 recruitment to MCVs displayed a high variability in timing that was independent of the size of the MCV or the bacterial burden. Most notably, the LC3-positive MCVs did not acidify, indicating that LC3 recruitment does not necessarily lead to the formation of mature autophagolysosomes. In addition, interferon-gamma (IFN-γ) pre-treatment did not affect LC3 recruitment frequency or autophagosome maturation, but increased the susceptibility of the macrophage to Mtb-induced cell death. Instead, LC3 recruitment and lysotracker staining were mutually exclusive events alternating on some MCVs multiple times showing a new reversible aspect of this autophagy response. It also suggested a role of autophagy in membrane repair of the MCV. Consistently, LC3 recruitment was strongly associated with galectin-3 and oxysterol-binding protein 1 staining, indicating a correlation with membrane damage and repair mechanisms. However, knockdown of ATG7 did not impact membrane repair, suggesting that autophagy is not directly involved in this process but is coregulated by the membrane damage of MCVs.

In summary, our findings provide novel insights into the dynamic and variable nature of LC3 recruitment and autophagy to the MCVs over time during Mtb infection. Our data suggests that there is no major role of autophagy in cell autonomous defense against Mtb nor membrane repair of the MCV in human macrophages. However, the combined dynamics of LC3 recruitment and Lysoview staining emerged as promising markers for future research focused on directly investigating the damage and repair processes of phagosomal membranes.
CMA-ES-Rosetta: Blackbox optimization algorithm traverses rugged peptide docking energy landscapes
Authors: Claussen Erin R. , Renfrew P. Douglas , Müller Christian L. , Drew Kevin
Published: December 2022

Energy minimization is necessary for virtually all modeling and design tasks and involves traversing extremely rugged energy landscapes. Although the gradient descent based minimization routines in Rosetta have fast runtimes, due to these rugged landscapes, minimization often converges into high-energy local minima. Alternative numerical optimization techniques, such as evolution strategies, are more robust to rugged landscapes and have been shown to be highly successful on a diverse set of problems. Here we explore the Covariance Matrix Adaptation Evolution Strategy (CMA-ES), a state-of-the-art derivative-free optimization algorithm, as a complementary approach to the default minimizer in Rosetta. We used a benchmark of 26 peptides, from the FlexPepDock Benchmark, to assess the performance of three algorithms in Rosetta, specifically, CMA-ES, Rosetta’s default minimizer, and a Monte Carlo protocol of small backbone perturbations. We test the algorithms’ performance on their ability to dock an idealized peptide to a series of hotspots residues (i.e. constraints) along a native peptide. Of the three methods, CMA-ES was able to find the lowest energy conformation for 23 out of 26 benchmark peptides. The application of CMA-ES allows for an alternative optimization method for macromolecular modeling problems with rough energy landscapes.
Dynamics of activation in the voltage-sensing domain of Ci-VSP
Authors: Guo Spencer C. , Shen Rong , Roux Benoît , Dinner Aaron R.
Published: December 2022

TheCiona intestinalisvoltage-sensing phosphatase (Ci-VSP) is a membrane protein containing a voltagesensing domain (VSD) that is homologous to VSDs from voltage-gated ion channels responsible for cellular excitability. Two crystal structures of Ci-VSD in putative resting and active conformations suggest a helical-screw voltage sensing mechanism in which the S4 helix translocates and rotates to enable exchange of salt-bridge partners. By combining extensive molecular dynamics simulations with a computational framework based on dynamical operators, we elucidate the microscopic mechanism of the resting-active transition at physiological membrane potential. Sparse regression reveals a small set of coordinates that distinguish intermediates hidden from electrophysiological measurements. The intermediates arise from a noncanonical helical-screw mechanism in which translocation, rotation, and side-chain movement of the S4 helix are only loosely coupled. These results provide new insights into existing experimental and computational findings on voltage sensing and suggest ways of further probing its mechanism.
Transcriptome guided metabolic network analysis reveals rearrangements of carbon flux distribution inNeisseria gonorrhoeaeduring neutrophil co-culture
Authors: Potter Aimee D. , Baiocco Christopher M. , Papin Jason A. , Criss Alison K.
Published: December 2022

The ability of bacterial pathogens to metabolically adapt to the environmental conditions of their hosts is critical to both colonization and invasive disease. Infection withNeisseria gonorrhoeae(the gonococcus, Gc) is characterized by the influx of neutrophils (PMNs), which fail to clear the bacteria and make antimicrobial products that can exacerbate tissue damage. The inability of the human host to clear Gc infection is particularly concerning in light of the emergence of strains that are resistant to all clinically recommended antibiotics. Bacterial metabolism represents a promising target for the development of new therapeutics against Gc. Here, we generated a curated genome-scale metabolic network reconstruction (GENRE) of Gc strain FA1090. This GENRE links genetic information to metabolic phenotypes and predicts Gc biomass synthesis and energy consumption. We validated this model with published data and in new results reported here. Contextualization of this model using the transcriptional profile of Gc exposed to PMNs revealed substantial rearrangements of Gc central metabolism and induction of Gc nutrient acquisition strategies for alternate carbon source use. These features enhanced the growth of Gc in the presence of neutrophils. From these results we conclude that the metabolic interplay between Gc and PMNs helps define infection outcomes. The use of transcriptional profiling and metabolic modeling to reveal new mechanisms by which Gc persists in the presence of PMNs uncovers unique aspects of metabolism in this fastidious bacterium, which could be targeted to block infection and thereby reduce the burden of gonorrhea in the human population.

Importance

The World Health Organization (WHO) designatedNeisseria gonorrhoeae(Gc) as a high priority pathogen for research and development of new antimicrobials. Bacterial metabolism is a promising target for new antimicrobials, as metabolic enzymes are widely conserved among bacterial strains and are critical for nutrient acquisition and survival within the human host. Here we used genome-scale metabolic modeling to characterize the core metabolic pathways of this fastidious bacterium, and to uncover the pathways used by Gc during culture with primary human immune cells. These analyses revealed that Gc relies on different metabolic pathways during co-culture with human neutrophils than in rich media. Conditionally essential genes emerging from these analyses were validated experimentally. These results show that metabolic adaptation in the context of innate immunity is important to Gc pathogenesis. Identifying the metabolic pathways used by Gc during infection can highlight new therapeutic targets for drug-resistant gonorrhea.