bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Arabidopsis SAC9 Enzyme is enriched in a cortical population of early endosomes and restricts PI(4,5)P2 at the Plasma Membrane
Authors: Lebecq Alexis , Doumane Mehdi , Fangain Aurélie , Bayle Vincent , Leong Jia Xuan , Rozier Frédérique , Marquès-Bueno Maria del Mar , Armengot Laia , Boisseau Romain P. , Simon Mathilde Laetitia Audrey , Franz-Wachtel Mirita , Macek Boris , Üstün Suayb , Jaillais Yvon , Caillaud Marie-Cécile 
Published: September 2021

ABSTRACTMembrane lipids, and especially phosphoinositides, are differentially enriched within the eukaryotic endomembrane system. This generates a landmark code by modulating the properties of each membrane. Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] specifically accumulates at the plasma membrane in yeast, animal and plant cells, where it regulates a wide range of cellular processes including endocytic trafficking. However, the functional consequences of mispatterning PI(4,5)P2 in plants are unknown. Here, we functionally characterized the putative phosphoinositide phosphatase SUPPRESSOR OF ACTIN9 (SAC9) in Arabidopsis thaliana (Arabidopsis). We found that SAC9 depletion led to the ectopic localization of PI(4,5)P2 on cortical intracellular compartments, which depends on PI4P and PI(4,5)P2 production at the plasma membrane. SAC9 localizes to a subpopulation of trans-Golgi Network/early endosomes that are enriched in a region close to the cell cortex and that are coated with clathrin. Furthermore, it interacts and colocalizes with Src Homology 3 Domain Protein 2 (SH3P2), a protein involved in endocytic trafficking. In the absence of SAC9, SH3P2 localization is altered and the clathrin-mediated endocytosis rate is reduced. Together, our results highlight the importance of restricting PI(4,5)P2 at the plasma membrane and illustrate that one of the consequence of PI(4,5)P2 misspatterning in plants is to impact the endocytic trafficking.One-sentence summarySAC9 prevents the accumulation of PI(4,5)P2 in intracellular compartments, thereby contributing to clathrin-mediated endocytosis at the plasma membrane and regulating the cortical localization of its protein partner SH3P2.
A Drosophila Circuit for Habituation Override
Authors: Trisal Swati , Aranha Marcia , Chodankar Ankita , VijayRaghavan K. , Ramaswami Mani 
Published: September 2021

ABSTRACTHabituated animals retain a latent capacity for robust engagement with familiar stimuli. In most instances, the ability to override habituation is best explained by postulating that habituation arises from the potentiation of inhibitory inputs onto stimulus-encoding assemblies and that habituation override occurs through disinhibition. Previous work has shown that inhibitory plasticity contributes to specific forms of olfactory and gustatory habituation in Drosophila. Here we analyze how exposure to a novel stimulus causes override of gustatory (proboscis-extension reflex or “PER”) habituation. While brief sucrose contact with tarsal hairs causes naïve Drosophila to extend their proboscis, persistent exposure reduces PER to subsequent sucrose stimuli. We show that in so habituated animals, either brief exposure of the proboscis to yeast or direct thermogenetic activation of sensory neurons restores PER response to tarsal sucrose stimulation. Similar override of PER habituation can also be induced by brief thermogenetic activation of a population of TH (Tyrosine-Hydroxylase) positive neurons, a subset of which send projections to the subesophagial zone (SEZ). Significantly, sensory-neuron induced habituation override requires transmitter release from these TH-positive cells. Treatments that cause override specifically influence the habituated state, with no effect on the naïve sucrose response across a range of concentrations. Taken together with other findings, these observations in female flies are consistent with a model in which novel taste stimuli trigger activity in dopaminergic neurons which, directly or indirectly, inhibit GABAergic cells that drive PER habituation. The implications of these findings for general mechanisms of attentional and sensory override of habituation are discussed.
mTOR activation induces endolysosomal remodeling and nonclassical secretion of IL-32 via exosomes in inflammatory reactive astrocytes
Authors: Leng Kun , Rooney Brendan , McCarthy Frank , Xia Wenlong , Rose Indigo V.L. , Bax Sophie , Chin Marcus , Fathi Saeed , Herrington Kari A. , Leonetti Manuel , Kao Aimee , Fancy Stephen P.J. , Elias Joshua E. , Kampmann Martin 
Published: September 2021

ABSTRACTAstrocytes respond and contribute to neuroinflammation by adopting inflammatory reactive states. Although recent efforts have characterized the gene expression signatures associated with these reactive states, the cell biology underlying inflammatory reactive astrocyte phenotypes remains under-explored. Here, we used CRISPR-based screening in human iPSC-derived astrocytes to identify mTOR activation a driver of cytokine-induced endolysosomal system remodeling, manifesting as alkalinization of endolysosomal compartments, decreased autophagic flux, and increased exocytosis of certain endolysosomal cargos. Through endolysosomal proteomics, we identified and focused on one such cargo – IL-32, a disease-associated pro-inflammatory cytokine not present in rodents, whose secretion mechanism is not well understood. We found that IL-32 was partially secreted in extracellular vesicles likely to be exosomes. Furthermore, we found that IL-32 was involved in the polarization of inflammatory reactive astrocyte states, was upregulated in astrocytes in multiple sclerosis lesions, and preferentially co-localized with astrocytes in hypoxic-ischemic encephalopathy. We believe that our results advance our understanding of cell biological pathways underlying inflammatory reactive astrocyte phenotypes and identify potential therapeutic targets.
Hedgehog signaling can enhance glycolytic ATP production in the Drosophila wing disc
Authors: Nellas Ioannis , Iyer K. Venkatesan , Iglesias-Artola Juan M. , Nadler André , Dye Natalie A. , Eaton Suzanne 
Published: September 2021

ABSTRACTEnergy production and utilization is critically important for animal development and growth. How it is regulated in space and time during tissue growth remains largely unclear. Toward this end, we used a FRET-based adenosine triphosphate (ATP) sensor to dynamically monitor ATP levels across a growing tissue, using the Drosophila wing disc. We discovered that steady-state levels of ATP are spatially uniform across the wing pouch. Pharmacologically inhibiting oxidative phosphorylation, however, reveals spatial heterogeneities in metabolic behavior, whereby signaling centers at compartment boundaries produce more ATP from glycolysis than the rest of the tissue. Genetic perturbations indicate that the conserved Hedgehog (Hh) signaling pathway can enhance ATP production by glycolysis. Collectively, our work reveals a positive feedback loop between Hh signaling and energy metabolism, advancing our understanding of the connection between conserved developmental patterning genes and energy production during animal tissue development.
Predicting long pendant edges in model phylogenies, with applications to biodiversity and tree inference
Authors: Bocharov Sergey , Harris Simon , Kominek Emma , Mooers Arne Ø. , Steel Mike 
Published: September 2021

AbstractIn the simplest phylogenetic diversification model (the pure-birth Yule process), lineages split independently at a constant rate λ for timet. The length of a randomly chosen edge (either interior or pendant) in the resulting tree has an expected value that rapidly converges toastgrows, and thus is essentially independent oft. However, the behaviour of the lengthLof thelongestpendant edge reveals remarkably different behaviour:Lconverges tot/2 as the expected number of leaves grows. Extending this model to allow an extinction rateμ(whereμ< λ), we also establish a similar result for birth-death trees, except thatt/2 is replaced byt/2 · (1 –μ/λ). This ‘complete’ tree may contain subtrees that have died out before timet; for the ‘reduced tree’ that just involves the leaves present at timetand their direct ancestors, the longest pendant edge lengthLagain converges tot/2. Thus, there is likely to be at least one extant species whose associated pendant branch attaches to the tree approximately half-way back in time to the origin of the entire clade. We also briefly consider the length of the shortest edges. Our results are relevant to phylogenetic diversity indices in biodiversity conservation, and to quantifying the length of aligned sequences required to correctly infer a tree. We compare our theoretical results with simulations, and with the branch lengths from a recent phylogenetic tree of all mammals.
The Drosophila TMEM184B ortholog Tmep ensures proper locomotion by restraining ectopic firing at the neuromuscular junction
Authors: Cho Tiffany S. , Beigaitė Eglė , Klein Nathaniel E. , Sweeney Sean T. , Bhattacharya Martha R.C. 
Published: September 2021

AbstractTMEM184B is a putative seven-pass membrane protein that promotes axon degeneration after injury. TMEM184B mutation causes aberrant neuromuscular architecture and sensory and motor behavioral defects in mice. The mechanism through which TMEM184B causes neuromuscular defects is unknown. We employed Drosophila melanogaster to investigate the function of the TMEM184B ortholog, Tmep (CG12004) at the neuromuscular junction. We show that Tmep is required for full adult viability and efficient larval locomotion. Tmep mutant larvae have a reduced body contraction rate compared to controls, with stronger deficits in females. Surviving adult Tmep mutant females show “bang sensitivity,” a phenotype associated with epileptic seizures. In recordings from body wall muscles, Tmep mutants show substantial hyperexcitability, with many post-synaptic potentials fired in response to a single stimulation, consistent with a role for Tmep in restraining synaptic excitability. Neuromuscular junctions in Tmep mutants show modest structural defects and satellite boutons, which could also contribute to poor locomotor performance. Tmep is expressed in endosomes and synaptic vesicles within motor neurons, suggesting a possible role in synaptic membrane trafficking. Using RNAi knockdown, we show that Tmep is required in motor neurons for proper larval locomotion and excitability. Locomotor defects can be rescued by presynaptic knock-down of endoplasmic reticulum calcium channels or by reducing evoked release probability, suggesting that excess synaptic activity drives behavioral deficiencies. Our work establishes a critical function for the TMEM184B ortholog Tmep in the regulation of synaptic transmission and locomotor behavior.
Post-encoding modulation of spatial memory by a GABAA-agonist
Authors: Iggena Deetje , Maier Patrizia M. , Häußler Sophia M. , Menk Mario , Olze Heidi , Larkum Matthew E. , Finke Carsten , Ploner Christoph J. 
Published: September 2021

AbstractWe investigated the role of the post-encoding period for consolidation of self-centered (egocentric) and world-centered (allocentric) spatial memory in neurologically normal human subjects. We used the GABAA-ergic anesthetic propofol to transiently modulate neural activity during the early stage of spatial memory consolidation. A total of 52 patients undergoing minor surgery learned to navigate to a target in a five-armed maze derived from animal experiments immediately prior to injection of propofol (early group) or more than 60 minutes before injection (late group). Two hundred and forty minutes after anesthesia, subjects were tested for memory-guided navigation. Our results show a selective impairment of memory-guided navigation in the early group and near-normal performance in the late group. Both egocentric and allocentric navigation were affected, albeit with distinct error patterns. In the egocentric condition, early group patients navigated significantly more often to a wrong alley of the maze but showed normal navigation times, thus suggesting a deficit mainly for memory of sequences of path segments. By contrast, in the allocentric condition, early group patients mostly navigated to the correct alley of the maze but showed a significant increase in detours and prolonged navigation times, thus suggesting a weakened representation of the relationship between landmarks. We conclude that presumably hippocampus-dependent networks contribute to early consolidation of representations underlying both egocentric and allocentric memory-guided navigation. Distinct aspects of these representations are susceptible to GABAA-ergic modulation within a post-encoding time-window of less than 60 minutes, indicating a redistribution and reconfiguration of spatial memory networks early during consolidation.Significance statementPropofol modulates consolidation of spatial representations underlying human spatial navigation.Following administration of propofol, memory-guided navigation using self-centered (egocentric) and world-centered (allocentric) spatial information is impaired.Error patterns after administration of propofol suggest modulation of a post-encoding integration process relevant for ego- and allocentric memory representations.The transient susceptibility of this process to GABAA-ergic modulation is consistent with rapid reconfiguration of networks for spatial memory shortly after learning.Propofol provides a pharmacological tool to investigate spatial memory consolidation in humans.
SAMD1 Distribution Patterns in Mouse Atherosclerosis Models Suggest Roles in LDL Retention, Antigen Presentation, and Cell Phenotype Modulation
Authors: Campbell Bruce , Bourassa Patricia , Aiello Robert 
Published: September 2021

AbstractThe theory that lesions formed by retention of circulating LDL can then progress to complicated atherosclerotic lesions has been a subject of debate, as has the mechanism of retention. In earlier work, we identified SAMD1, a protein expressed by intimal smooth muscle cells in human lesions that appears to irreversibly bind apoB-Lps in extracellular matrix near the lumen. We hypothesized this binding could contribute to the formation of lesions in mice, and that inhibiting binding could reduce lesion growth. In mouse models of atherosclerosis, we found that SAMD1 binds LDL; that SAMD1/apoB complex is ingested by intimal cells; and that recognizable epitopes of the SAMD1/apoB complex survive some degree of catabolism in foam cell. These data appear to support the SAMD1/LDL retention hypothesis of lesion growth. Despite apparently irreversible binding of human LDL to full-length human SAMD1, efficient anti-SAMD1-antibody inhibitors were created. In vivo lesion targeting of inhibitors was demonstrated by MRI, ultrasound, and ex vivo microscopy. However, only non-statistically significant reductions in spontaneous lesion size in apoE-/- mice were seen after 12 weeks of treatment with PEG-fab inhibitors of SAMD1/LDL binding. In contrast, these inhibitors substantially reduced LDL retention in carotid injury lesions in apoE-/- and LDLR-/- mice 7 days after injury. The most obvious difference between injury lesions and early spontaneous lesions is the presence of numerous smooth muscle cells and associated extracellular matrix in the injury lesions. Thus, SAMD1 may be involved in retention of apoB-Lps in mouse lesions, but not until smooth muscle cells have entered the intima. In addition, SAMD1 is seen throughout arteries in changing patterns that suggest broader and more complicated roles in atherosclerosis.
Computation of the mitochondrial age distribution along the axon length
Authors: Kuznetsov Ivan A. , Kuznetsov Andrey V. 
Published: September 2021

AbstractWe describe a compartmental model of mitochondrial transport in axons, which we apply to compute mitochondrial age at different distances from the soma. The model predicts that at the tip of an axon that has a length of 1 cm, the average mitochondrial age is approximately 22 hours. The mitochondria are youngest closest to the soma and their age scales approximately linearly with distance from the soma. To the best of the authors’ knowledge, this is the first attempt to predict the spatial distribution of mitochondrial age within an axon. A sensitivity study of the mean age of mitochondria to various model parameters is also presented.
Full-length NLRP3 forms oligomeric cages to mediate NLRP3 sensing and activation
Authors: Andreeva Liudmila , David Liron , Rawson Shaun , Shen Chen , Pasricha Teerithveen , Pelegrin Pablo , Wu Hao 
Published: September 2021

AbstractThe nucleotide-binding domain and leucine-rich-repeat (LRR) containing protein 3 with a pyrin domain (NLRP3) is emerging to be a critical intracellular inflammasome sensor of membrane integrity and a highly important clinical target against chronic inflammation. Here we report that the endogenous, stimulus-responsive form of full-length NLRP3 is a 12-16 mer double ring cage held together by LRR-LRR interactions with the pyrin domains shielded within the assembly to avoid premature activation. Surprisingly, this NLRP3 form is predominantly membrane localized, which is consistent with previously noted localization of NLRP3 at various membrane organelles. Structure-guided mutagenesis reveals that trans-Golgi network dispersion into vesicles, an early event observed for all NLRP3 activating stimuli, requires the double ring cages of NLRP3. Double ring-defective NLRP3 mutants further abolish inflammasome punctum formation, caspase-1 processing and cell death. Thus, unlike other inflammasome sensors that are monomeric when inactive, our data uncover a unique NLRP3 oligomer on membrane that is poised to sense diverse signals to induce inflammasome activation.
Therapeutic efficacy of an oral nucleoside analog of remdesivir against SARS-CoV-2 pathogenesis in mice
Authors: Schäfer Alexandra , Martinez David R. , Won John J. , Moreira Fernando R. , Brown Ariane J. , Gully Kendra L. , Kalla Rao , Chun Kwon , Du Pont Venice , Babusis Darius , Tang Jennifer , Murakami Eisuke , Subramanian Raju , Barrett Kimberly T , Bleier Blake J. , Bannister Roy , Feng Joy Y. , Bilello John P. , Cihlar Tomas , Mackman Richard L. , Montgomery Stephanie A. , Baric Ralph S. , Sheahan Timothy P. 
Published: September 2021

AbstractThe COVID-19 pandemic remains uncontrolled despite the rapid rollout of safe and effective SARS-CoV-2 vaccines, underscoring the need to develop highly effective antivirals. In the setting of waning immunity from infection and vaccination, breakthrough infections are becoming increasingly common and treatment options remain limited. Additionally, the emergence of SARS-CoV-2 variants of concern with their potential to escape therapeutic monoclonal antibodies emphasizes the need to develop second-generation oral antivirals targeting highly conserved viral proteins that can be rapidly deployed to outpatients. Here, we demonstrate the in vitro antiviral activity and in vivo therapeutic efficacy of GS-621763, an orally bioavailable prodrug of GS-441524, the parental nucleoside of remdesivir, which targets the highly conserved RNA-dependent RNA polymerase. GS-621763 exhibited significant antiviral activity in lung cell lines and two different human primary lung cell culture systems. The dose-proportional pharmacokinetic profile observed after oral administration of GS-621763 translated to dose-dependent antiviral activity in mice infected with SARS-CoV-2. Therapeutic GS-621763 significantly reduced viral load, lung pathology, and improved pulmonary function in COVID-19 mouse model. A direct comparison of GS-621763 with molnupiravir, an oral nucleoside analog antiviral currently in human clinical trial, proved both drugs to be similarly efficacious. These data demonstrate that therapy with oral prodrugs of remdesivir can significantly improve outcomes in SARS-CoV-2 infected mice. Thus, GS-621763 supports the exploration of GS-441524 oral prodrugs for the treatment of COVID-19 in humans.
Paramagnetic spin labeling of a bacterial DnaB helicase for solid-state NMR
Authors: Zehnder Johannes , Cadalbert Riccardo , Yulikov Maxim , Künze Georg , Wiegand Thomas 
Published: September 2021

AbstractLabeling of biomolecules with a paramagnetic probe for nuclear magnetic resonance (NMR) spectroscopy enables determining long-range distance restraints, which are otherwise not accessible by classically used dipolar coupling-based NMR approaches. Distance restraints derived from paramagnetic relaxation enhancements (PREs) can facilitate the structure determination of large proteins and protein complexes. We herein present the site-directed labeling of the large oligomeric bacterial DnaB helicase from Helicobacter pylori with cysteine-reactive maleimide tags carrying either a nitroxide radical or a lanthanide ion. The success of the labeling reaction was followed by quantitative continuous-wave electron paramagnetic resonance (EPR) experiments performed on the nitroxide-labeled protein. PREs were extracted site-specifically from 2D and 3D solid-state NMR spectra. A good agreement with predicted PRE values, derived by computational modeling of nitroxide and Gd3+ tags in the low-resolution DnaB crystal structure, was found. Comparison of experimental PREs and model-predicted spin label-nucleus distances indicated that the size of the “blind sphere” around the paramagnetic center, in which NMR resonances are not detected, is slightly larger for Gd3+ (~14 Å) than for nitroxide (~11 Å) in 13C-detected 2D spectra of DnaB. We also present Gd3+-Gd3+ dipolar electron-electron resonance EPR experiments on DnaB supporting the conclusion that DnaB was present as a hexameric assembly.
Experimental confirmation that an uncommon, yet clinically relevant mutation (G878A) in the rrs gene of Mycobacterium tuberculosis confers resistance to streptomycin
Authors: Domenech Pilar , Mouhoub Esma , Reed Michael B. 
Published: September 2021

ABSTRACTThe effective treatment of patients diagnosed with drug resistant tuberculosis (TB) is highly dependent upon the ability to rapidly and accurately determine the antibiotic resistance/ susceptibility profile of the Mycobacterium tuberculosis isolate(s) involved. Thus, as more and more clinical microbiology laboratories advance towards the routine use of DNA sequence-based diagnostics, it is imperative that their predictive functions extend beyond the well-known resistance-conferring mutations, in order to also encompass as many of the lower-frequency mutations as possible. However, in most cases, the fundamental experimental proof that links these uncommon mutations with phenotypic resistance is still lacking. One such example is the G878A polymorphism within the rrs gene encoding the 16s rRNA. We, and others, have identified this mutation within a small number of drug-resistant M. tuberculosis isolates, although prior to this study a consensus regarding exactly which aminoglycoside antibiotic(s) it conferred resistance toward seems not to have been reached. Here we have employed oligo-mediated recombineering to specifically introduce the G878A polymorphism into the rrs gene of M. bovis BCG - a species very closely related to M. tuberculosis - and demonstrate that it confers low-level resistance to streptomycin alone. In our hands, it does not confer cross-resistance towards amikacin, capreomycin, nor kanamycin. We also demonstrate that the rrsG878A mutation exerts a substantial fitness defect in vitro, that may at least in part explain why clinical M. tuberculosis isolates bearing this mutation appear to be quite rare. Overall, this study provides clarity to the resistance phenotype attributable to the rrsG878A mutation and is relevant to the future implementation of genomics-based diagnostics, as well as the clinical management of patients in situations where this particular polymorphism is encountered.