bluuebunny/crossref_metadata_2025_split huggingface dataset:
Investigation of adenosine A1 receptor mediated β-arrestin 2 recruitment using a split-luciferase assay
Authors: Saecker Luisa , Häberlein Hanns , Franken Sebastian 
Published: February 2023

AbstractBackgroundAdenosine A1 receptor (A1AR) plays a prominent role in neurological and cardiac diseases as well as during inflammatory processes. Its endogenous ligand adenosine is known to be one of the key players in the sleep-wake cycle. Like other G protein-coupled receptors, stimulation of A1AR leads to the recruitment of arrestins in addition to the activation of G proteins. So far, little is known about the role of these proteins for signal transduction and regulation of A1AR compared to the activation of G proteins. In this work, we characterized a live cell assay for the A1AR mediated β-arrestin 2 recruitment. We have applied this assay to a set of different compounds that interact with this receptor.MethodsBased on NanoBit®technology, a protein complementation assay was developed in which the A1AR is coupled to the large part of the nanoluciferase (LgBiT) whereas its small part (SmBiT) is fused to the N-terminus of β-arrestin 2. Stimulation of A1AR results in the recruitment of β-arrestin 2 and subsequent complementation of a functional nanoluciferase. For comparison, corresponding data on the effect of receptor stimulation on intracellular cAMP levels were collected for some data sets using the GloSensorTMassay.ResultsThe assay gives highly reproducible results with a very good signal to noise ratio. Capadenoson, in contrast to adenosine, CPA or NECA, shows only partial agonism in this assay with respect to the recruitment of β-arrestin 2, whereas it is a full agonist in the case of the inhibitory effect of A1AR on cAMP production. By using a GRK2 inhibitor, it becomes clear that the recruitment is at least partially dependent on the phosphorylation of the receptor by this kinase. Interestingly, this was also the first time that we were able to demonstrate the A1AR mediated recruitment of β-arrestin 2 by stimulation with a valerian extract.ConclusionThe presented assay is a useful tool for the quantitative study of the A1AR mediated β-arrestin 2 recruitment. It allows data collection for stimulatory, inhibitory as well as modulatory substances and is also suitable for more complex substance mixtures such as valerian extract.
Nutritional stress-induced regulation of microtubule organization and mRNP transport by HDAC1 controlled α-tubulin acetylation
Authors: Wippich Frank , Vaishali  , Hennrich Marco L. , Ephrussi Anne 
Published: February 2023

AbstractIn response to nutritional stress, microtubules in cells of theDrosophilafemale germline are depleted from the cytoplasm and accumulate cortically. This triggers aggregation of mRNPs into large reticulated sponge bodies and oogenesis arrest. Here, we show that hyperacetylation of α-tubulin at lysine 40 (K40) alters microtubule dynamics and sponge body formation. We found that depletion of histone deacetylase 1 (HDAC1) by RNAi phenocopies the nutritional stress response, causing hyperacetylation of α-tubulin, as well as accumulation of maternally deposited mRNPs in reticulated sponge bodies. Usingin vitroandin vivostudies, we identify HDAC1 as a direct regulator of α-tubulin K40 acetylation status. In well-fed flies, HDAC1 maintains low levels of α-tubulin acetylation, enabling the microtubule dynamics required for mRNP transport. Using quantitative phosphoproteomics we identify nutritional stress-induced changes in protein phosphorylation that act upstream of α-tubulin acetylation, including phosphorylation of HDAC1 at S391, which reduces its ability to deacetylate α-tubulin. These results reveal thatDrosophilaHDAC1 senses and relays the nutritional status, which regulates germline development through modulation of cytoskeleton dynamics.
Structural insights into agonist-binding and activation of the human complement C3a receptor
Authors: Yadav Manish K. , Yadav Ravi , Sarma Parishmita , Maharana Jagannath , Soni Chahat , Saha Sayantan , Singh Vinay , Ganguly Manisankar , Saha Shirsha , Khant Htet A. , Banerjee Ramanuj , Shukla Arun K. , Gati Cornelius 
Published: February 2023

AbstractThe complement cascade is an integral part of innate immunity, and it plays a crucial role in our body’s innate immune response including combating microbial infections. Activation of the complement cascade results in the generation of multiple peptide fragments, of which complement C3a and C5a are potent anaphylatoxins. The complement C3a binds and activates a G protein-coupled receptor (GPCR) known as C3aR while C5a activates two distinct receptors namely C5aR1 and C5aR2. Our current understanding of complement peptide recognition by their corresponding receptors is limited primarily to biochemical studies, and direct structural visualization of ligand-receptor complexes is still elusive. Here, we present structural snapshots of C3aR in complex with heterotrimeric G-proteins, with the receptor in ligand-free state, activated by full-length complement C3a, and a peptide agonist EP54, derived based on the carboxyl-terminal sequence of C5a. A comprehensive analysis of these structures uncovers the critical residues involved in C3a-C3aR interaction, and also provides molecular insights to rationally design carboxyl-terminal fragments of C3a and C5a to act as potent agonists of the receptor. Surprisingly, a comparison of C3a-C3aR structure with C5a-C5aR1 structure reveals diagonally opposite placement of these two complement peptides on their respective receptors, which helps explain the subtype selectivity of these complement peptides. Finally, taking lead from the structural insights, we also identify EP141, a peptide derived from the carboxyl-terminus of C3a, as a G-protein-biased agonist at C3aR. Taken together, our study illuminates the structural mechanism of complement C3a recognition by C3aR, and it also offers the first structural template for designing novel C3aR ligands with therapeutic potential for inflammatory disorders.
The iron-dopamine D1 coupling modulates neural signatures of working memory across adult lifespan
Authors: Gustavsson Jonatan , Johansson Jarkko , Falahati Farshad , Andersson Micael , Papenberg Goran , Avelar-Pereira Bárbara , Bäckman Lars , Kalpouzos Grégoria , Salami Alireza 
Published: February 2023

AbstractBrain iron overload and decreased integrity of the dopaminergic system have been independently reported as brain substrates of cognitive decline in aging. Dopamine (DA), and iron are co-localized in high concentrations in the striatum and prefrontal cortex (PFC), but follow opposing age-related trajectories across the lifespan. DA contributes to cellular iron homeostasis and the activation of D1-like DA receptors (D1DR) alleviates oxidative stress-induced inflammatory responses, suggesting a mutual interaction between these two fundamental components. Still, a direct in-vivo study testing the iron-D1DR relationship and their interactions on brain function and cognition across the lifespan is rare. Using PET and MRI data from the DyNAMiC study (n=180, age=20-79, %50 female), we showed that elevated iron content was related to lower D1DRs in DLPFC, but not in striatum, suggesting that dopamine-rich regions are less susceptible to elevated iron. Critically, older individuals with elevated iron and lower D1DR exhibited less frontoparietal activations during the most demanding task, which in turn was related to poorer working-memory performance. Together, our findings suggest that the combination of elevated iron load and reduced D1DR contribute to disturbed PFC-related circuits in older age, and thus may be targeted as two modifiable factors for future intervention.HighlightsFirst study demonstrating the association between regional iron and dopamine D1DR in adult humans.The interplay between age-related elevated iron and diminished D1DR explained lower task-related brain activity, which in turn was related to poorer task performance.Our findings iron-DA coupling can help progress the understanding of the mechanisms behind DA-related neurodegeneration.
It takes tau to tango: Investigating the fuzzy interaction between the R2-repeat domain and tubulin C-terminal tails
Authors: Marien Jules , Prévost Chantal , Sacquin-Mora Sophie 
Published: February 2023

AbstractThe microtubule-associated protein (MAP) tau plays a key role in the regulation of microtubule assembly and spatial organisation. Tau hyperphosphorylation affects its binding on the tubulin surface and has been shown to be involved in several pathologies such as Alzheimer disease. As the tau binding site on the microtubule lays close to the disordered and highly flexible tubulin C-terminal tails (CTTs), these are likely to impact the tau-tubulin interaction. Since the disordered tubulin CTTs are missing from the available experimental structures, we used homology modeling to build two complete models of tubulin heterotrimers with different isotypes for the β-tubulin subunit (βI/αI/ βI and βIII/αI/βIII). We then performed long timescale classical Molecular Dynamics simulations for the tau-R2/tubulin assembly (in systems with and without CTTs) and analyzed the resulting trajectories to obtain a detailed view of the protein interface in the complex and the impact of the CTTs on the stability of this assembly. Additional analyses of the CTTs mobility in the presence, or in the absence, of tau also highlight how tau might modulate the CTTs activity as hooks that are involved in the recruitment of several MAPs. In particular, we observe awrappingmechanism, where the β-tubulin CTTs form a loop over tau-R2, thus stabilizing its interaction with the tubulin surface and simultaneously reducing the CTTs availability for interactions with other MAPs.
Zika virus E protein alters blood-brain barrier by modulating brain microvascular endothelial cell and astrocyte functions
Authors: Kaur Guneet , Pant Pallavi , Bhagat Reshma , Seth Pankaj 
Published: February 2023

AbstractNeurotropic viruses can cross the otherwise dynamically regulated blood-brain barrier (BBB) and affect the brain cells. Zika virus (ZIKV) is an enveloped neurotropicFlavivirusknown to cause severe neurological complications, such as encephalitis and foetal microcephaly. In the present study, we used human brain microvascular endothelial cells (hBMECs) and human progenitor derived astrocytes to form a physiologically relevant BBB model. We used this model to investigate the effects of ZIKV envelope (E) protein on properties of cells comprising the BBB. E protein is the principal viral protein involved in interaction with host cell surface receptors, facilitating the viral entry. Our findings show that ZIKV E protein results in activation of both hBMECs and astrocytes. hBMECs showed reduced expression of endothelial junction proteins - ZO-1, Occludin and VE-Cadherin, which are crucial in establishing and maintaining the BBB. As a result, ZIKV E protein triggered alteration in BBB integrity and permeability. We also found upregulation of genes involved in leukocyte recruitment along with increased proinflammatory chemokines and cytokines upon exposure to E protein. Furthermore, E protein resulted in astrogliosis as seen by increased expression of GFAP and Vimentin. Both BBB cell types exhibited inflammatory response following exposure to E protein which may influence viral access into the central nervous system (CNS), resulting in infection of other CNS cells. Overall, our study provides valuable insights into the transient changes that occur at the site of BBB upon ZIKV infection.
Pumping Iron: A Multi-omics Analysis of Two Extremophilic Algae Reveals Iron Economy Management
Authors: Davidi Lital , Gallaher Sean D. , Ben-David Eyal , Purvine Samuel O. , Filmore Thomas L. , Nicora Carrie D. , Craig Rory J. , Schmollinger Stefan , Roje Sanja , Blaby-Haas Crysten E. , Auber Robert P. , Wisecaver Jennifer , Merchant Sabeeha S. 
Published: February 2023

AbstractMarine algae are responsible for half of the world’s primary productivity, but this critical carbon sink is often constrained by insufficient iron. One species of marine algae,Dunaliella tertiolecta, is remarkable for its ability to maintain photosynthesis and thrive in low-iron environments. A related species,Dunaliella salinaBardawil, shares this attribute but is an extremophile found in hyper-saline environments. To elucidate how algae manage their iron requirements, we produced high-quality genome assemblies and transcriptomes for both species to serve as a foundation for a comparative multi-omics analysis. We identified a host of iron-uptake proteins in both species, including a massive expansion of transferrins and a novel family of siderophore-iron uptake proteins. Complementing these multiple iron-uptake routes, ferredoxin functions as a large iron reservoir that can be released by induction of flavodoxin. Proteomic analysis revealed reduced investment in the photosynthetic apparatus coupled with remodeling of antenna proteins by dramatic iron-deficiency induction of TIDI1, an LHCA-related protein found also in other chlorophytes. These combinatorial iron scavenging and sparing strategies makeDunaliellaunique among photosynthetic organisms.Significance StatementDespite their small size, microalgae play a huge role in CO2uptake via photosynthesis, and represent an important target for climate crisis mitigation efforts. Most photosynthesis proteins require iron as a co-factor so that insufficient iron often limits algal CO2sequestration. With this in mind, we examined a genus of microalgae calledDunaliellathat is exceptionally well-adapted to low iron environments. We produced complete genomes, transcriptomes, and proteomes for two species ofDunaliellathat hail from radically different environments: one from coastal ocean waters and the other from salt flats. We identified dozens of genes and multiple, complementary strategies that both species utilize for iron-uptake and management that explainDunaliella’sremarkable ability to thrive on low iron.
DNA modifications impact natural transformation ofAcinetobacter baumannii
Authors: Vesel Nina , Iseli Christian , Guex Nicolas , Lemopoulos Alexandre , Blokesch Melanie 
Published: February 2023

ABSTRACTAcinetobacter baumanniiis a dangerous nosocomial pathogen, especially due to its ability to rapidly acquire new genetic traits, including antibiotic resistance genes (ARG). InA. baumannii, natural competence for transformation, one of the primary modes of horizontal gene transfer (HGT), is thought to contribute to ARG acquisition and has therefore been intensively studied. However, knowledge regarding the potential role of epigenetic DNA modification(s) on this process remains lacking. Here, we demonstrate that the methylome pattern of diverseA. baumanniistrains differs substantially and that these epigenetic marks influence the fate of transforming DNA. Specifically, we describe a methylome-dependent phenomenon that impacts intra- and inter-species DNA exchange by the competentA. baumanniistrain A118. We go on to identify and characterize an A118-specific restrictionmodification (RM) system that impairs transformation when the incoming DNA lacks a specific methylation signature. Collectively, our work contributes towards a more holistic understanding of HGT in this organism and may also aid future endeavors towards tackling the spread of novel ARGs. In particular, our results suggest that DNA exchanges between bacteria that share similar epigenomes are favored and could therefore guide future research into identifying the reservoir(s) of dangerous genetic traits for this multi-drug resistant pathogen.
Lung injury induces a polarized immune response by self antigen-specific Foxp3+regulatory T cells
Authors: Shin Daniel S. , Ratnapriya Sneha , Cashin Creel Ng , Kuhn Lucy F. , Rahimi Rod A. , Anthony Robert M. , Moon James J. 
Published: February 2023

SummarySelf antigen-specific T cells are prevalent in the mature adaptive immune system, but are regulated through multiple mechanisms of tolerance. However, inflammatory conditions such as tissue injury may provide these T cells with an opportunity to break tolerance and trigger autoimmunity. To understand how the T cell repertoire responds to the presentation of self antigen under highly stimulatory conditions, we used peptide:MHCII tetramers to track the behavior of endogenous CD4+T cells with specificity to a lung-expressed self antigen in mouse models of immune-mediated lung injury. Acute injury resulted in the exclusive expansion of regulatory T cells (Tregs) that was dependent on self antigen recognition and IL-2. Conversely, conventional T cells of the same self antigen specificity remained unresponsive, even following Treg ablation. Thus, the self antigen-specific T cell repertoire is poised to serve a regulatory function during acute tissue damage to limit further damage and the possibility of autoimmunity.
Synergistic induction of blood-brain barrier properties
Authors: Porkoláb Gergő , Mészáros Mária , Szecskó Anikó , Vigh Judit P. , Walter Fruzsina R. , Figueiredo Ricardo , Kálomista Ildikó , Hoyk Zsófia , Vizsnyiczai Gaszton , Gróf Ilona , Jan Jeng-Shiung , Gosselet Fabien , Pirity Melinda K. , Vastag Monika , Hudson Natalie , Campbell Matthew , Veszelka Szilvia , Deli Mária A. 
Published: February 2023

AbstractBlood-brain barrier (BBB) models derived from human stem cells are powerful tools to improve our understanding of cerebrovascular diseases and to facilitate drug development for the human brain. Yet providing stem cell-derived endothelial cells with the right signaling cues to acquire BBB characteristics while also retaining their vascular identity remains challenging. Here, we show that the simultaneous activation of cyclic AMP and Wnt/β-catenin signaling, and inhibition of the TGF-β pathway in endothelial cells robustly induce BBB propertiesin vitro. To target this novel interaction, we present a small molecule cocktail named cARLA, which synergistically enhances barrier tightness in a range of BBB models across species. Mechanistically, we reveal that the three pathways converge on Wnt/β-catenin signaling to mediate the effect of cARLAviathe tight junction protein claudin-5. We demonstrate that cARLA shifts the gene expressional profile of human stem cell-derived endothelial cells towards thein vivobrain endothelial signature, with a higher glycocalyx density and efflux pump activity, lower rates of endocytosis and a characteristic endothelial response to proinflammatory cytokines. Finally, we illustrate how cARLA can improve the predictive value of human BBB models regarding the brain penetration of drugs and targeted nanoparticles. Due to its synergistic effect, high reproducibility and ease of use, cARLA has the potential to advance drug development for the human brain by improving BBB models across laboratories.Significance StatementThe blood-brain barrier (BBB) hinders drug delivery to the brain and is implicated in neurological diseases. To better understand these processes in humans, there is a need for culture models that mimic the complexity of the BBB. However, state-of-the-art human BBB models either suffer from a non-physiological, mixed epithelial-endothelial identity or have weak barrier tightness, which greatly limits their usability. We identified a molecule combination that synergistically enhances barrier tightness in severalin vitromodels and induces complex BBB properties in human stem cell-derived endothelial cells by targeting a novel link between three signaling pathways. The molecule combination has the potential to improve BBB culture models across laboratories to advance both basic research and drug development for the human brain.
Repetitive execution of a reach-and-lift task is associated with inter-trial changes in movement-related EEG features and their predictive power
Authors: Paek Andrew , Prashad Shikha 
Published: February 2023

AbstractBrain Machine Interfaces (BMIs) can help restore motor function to individuals with paralysis. These systems allow users to control an assistive device through the detection of movement-related brain activity. Such neural signatures are found through machine learning algorithms and training datasets that are generated from participants performing repetitive motor tasks. We anticipate that the movement-related brain waves of interest can attenuate over time due to neural efficiency, where the brain becomes more efficient with practice in a motor task. To explore this hypothesis, we used three open-access EEG datasets where participants performed a simple reach-and-lift task. From each trial, time windows associated with resting and movement periods were segmented. Alpha- and beta-band spectral power was estimated for each epoch, and event-related desynchronization (ERD) was estimated as the suppression in spectral power from rest to movement. These ERDs were compared between early and late trials in the dataset. We also used linear discriminant analysis to assess a machine learning algorithm’s accuracy in classifying whether the time windows belonged to rest or movement based on spectral power. In some cases, the ERDs were significantly different between earlier and later trials, and these differences led to changes in predicting the presence of movement from these ERDs. These results call for a reevaluation of BMI performance in datasets with numerous trials and an exploration of strategies that can compensate for longitudinal changes in movement-related brain activity used for BMIs.
Frataxin-deficient human brain microvascular endothelial cells lose polymerized actin and are paracellularly permeable –implications for blood-brain barrier integrity in Friedreich’s Ataxia
Authors: Smith Frances M. , Kosman Daniel J 
Published: February 2023

AbstractBackgroundFriedreich’s Ataxia (FRDA) is the most prevalent inherited ataxia; the disease results from loss of Frataxin, an essential mitochondrial iron trafficking protein. FRDA presents as neurodegeneration of the dorsal root ganglion and cerebellar dentate nuclei, followed by brain iron accumulation in the latter. End stage disease includes cardiac fibrosis that contributes to hypertrophic cardiomyopathy. The microvasculature plays an essential barrier role in both the brain and heart, thus an investigation of this tissue system in FRDA is essential to the delineation of the cellular dysfunction in this genetic disorder. Here, we investigate brain microvascular endothelial cell integrity in FRDA in a model of the blood-brain barrier (BBB).MethodsWe used lentiviral mediated shRNA delivery to generate a novel FRDA model in immortalized human brain microvascular endothelial cells (hBMVEC) that compose the microcapillaries of the BBB. We verified known cellular pathophysiologies of FXN knockdown including increased oxidative stress, loss of energy metabolism, and increased cell size. Furthermore, we investigated cytoskeletal architecture including the abundance and organization of filamentous actin, and barrier physiologyviatransendothelial electrical resistance and fluorescent tracer flux.ResultsshFXN hBMVEC display the known FRDA cell morbidity including increased oxidative stress, decreased energy metabolism, and an increase in cell size. We demonstrate that shFXN hBMVEC have less overall filamentous actin, and that filamentous actin is lost at the cell membrane and cortical actin ring. Consistent with loss of cytoskeletal structure and anchorage, we found decreased barrier strength and increased paracellular tracer flux in the shFXN hBMVEC transwell model.ConclusionWe identified that insufficient FXN levels in the hBMVEC BBB model causes changes in cytoskeletal architecture and increased barrier permeability, cell pathologies that may be related to patient brain iron accumulation, neuroinflammation, neurodegeneration, and stroke. Our findings implicate other barrier cells,e.g.,the cardiac microvasculature, likely contributory also to disease pathology in FRDA.