bluuebunny/crossref_metadata_2025_split huggingface dataset:
Predicting resprouting ofPlatanus×hispanicafollowing branch pruning by means of machine learning
Authors: Shu Qiguan , Yazdi Hadi , Rötzer Thomas , Ludwig Ferdinand 
Published: August 2023

SummaryResprouting is a crucial survival strategy following the loss of branches, being it by natural events or artificially by pruning. The prediction of resprouting patterns on a physiological basis is a highly complex approach. However, trained gardeners try to predict a tree’s resprouting after pruning purely based on their empirical knowledge and a visual check of the tree’s geometry. In this study, we explore in how far such predictions can also be made by algorithms, especially using machine learning.Table-topped annually prunedPlatanus×hispanicatrees at a nursery were documented with terrestrial LiDAR scanners in two consecutive years. Topological structures for these trees were abstracted from point clouds by cylinder fitting. Then, new shoots and trimmed branches were labelled on corresponding cylinders. Binary and multiclass classification models were tested for predicting the location and number of new sprouts.The accuracy for predicting whether having or not new shoots on each cylinder reaches 90.8% with the LGBMClassifier, the balanced accuracy is 80.3%. The accuracy for predicting the exact numbers of new shoots with GaussianNB model is 82.1% but its balanced accuracy is reduced to 42.9%.The results were validated with a separate evaluation dataset. It proves a feasibility in predicting resprouting patterns after pruning using this approach. Different tree species, tree forms, and other variables should be addressed in further research.
Glutamate helps unmask the differences in driving forces for phase separation versus clustering of FET family proteins in sub-saturated solutions
Authors: Kar Mrityunjoy , Vogel Laura T. , Chauhan Gaurav , Ausserwöger Hannes , Welsh Timothy J. , Kamath Anjana R. , Knowles Tuomas P. J. , Hyman Anthony A. , Seidel Claus A. M. , Pappu Rohit V. 
Published: August 2023

AbstractMultivalent proteins undergo coupled segregative and associative phase transitions. Phase separation, a segregative transition, is driven by macromolecular solubility, and this leads to coexisting phases above system-specific saturation concentrations. Percolation is a continuous transition that is driven by multivalent associations among cohesive motifs. Contributions from percolation are highlighted by the formation of heterogeneous distributions of clusters in sub-saturated solutions, as was recently reported for Fused in sarcoma (FUS) and FET family proteins. Here, we show that clustering and phase separation are defined by a separation of length- and energy-scales. This is unmasked when glutamate is the primary solution anion. Glutamate is preferentially excluded from protein sites, and this enhances molecular associations. Differences between glutamate and chloride are manifest at ultra-low protein concentrations. These differences are amplified as concentrations increase, and they saturate as the micron-scale is approached. Therefore, condensate formation in supersaturated solutions and clustering in sub-saturated are governed by distinct energy and length scales. Glutamate, unlike chloride, is the dominant intracellular anion, and the separation of scales, which is masked in chloride, is unmasked in glutamate. Our work highlights how components of cellular milieus and sequence-encoded interactions contribute to amplifying distinct contributions from associative versus segregative phase transitions.
Interkingdom interactions shape the fungal microbiome of mosquitoes
Authors: Hegde Shivanand , Khanipov Kamil , Hornett Emily A , Nilyanimit Pornjarim , Pimenova Maria , Saldaña Miguel A. , de Becker Charissa , Golovko Georgiy , Hughes Grant L. 
Published: August 2023

AbstractThe mosquito microbiome is an important modulator of vector competence and vectoral capacity. Unlike the extensively studied bacterial microbiome, fungal communities in the mosquito microbiome (mycobiome) remain largely unexplored. To work towards getting an improved understanding of the fungi associated with mosquitoes, we sequenced the mycobiome of three field-collected and laboratory-reared mosquito species (Aedes albopictus, Aedes aegypti,andCulex quinquefasciatus). Our analysis showed both environment and host species were contributing to the diversity of the fungal microbiome of mosquitoes. When comparing species,Ae. albopictuspossessed a higher number of diverse fungal taxa thanCx. quinquefasciatus,while strikingly less than 1% of reads fromAe. aegyptisamples were fungal. Fungal reads fromAe. aegyptiwere <1% even after inhibiting host amplification using a PNA blocker, indicating that this species lacked a significant fungal microbiome that was amplified using this sequencing approach. Using a mono-association mosquito infection model, we confirmed that mosquito-derived fungal isolates colonize and forAedesmosquitoes, support growth and development at comparable rates to their bacterial counterparts. Strikingly, native bacterial taxa isolated from mosquitoes impeded the colonization of symbiotic fungi inAe. aegyptisuggesting interkingdom interactions shape fungal microbiome communities. Collectively, this study adds to our understanding of the fungal microbiome of different mosquito species, that these fungal microbes support growth and development, and highlights that microbial interactions underpin fungal colonization of these medically relevent species.
Synergistic control of chloroplast biogenesis byMYB-relatedandGolden2-liketranscription factors
Authors: Frangedakis Eftychios , Yelina Nataliya E. , Billakurthi Kumari , Schreier Tina , Dickinson Patrick J. , Tomaselli Marta , Haseloff Jim , Hibberd Julian M. 
Published: August 2023

AbstractChloroplast biogenesis is dependent on master regulators from the GOLDEN2-LIKE (GLK) family of transcription factors, butglkmutants contain residual chlorophyll and therefore other proteins must also be involved. Here we identify MYB-related transcription factors as regulators of chloroplast biogenesis in the liverwortMarchantia polymorphaand angiospermArabidopsis thaliana. In both species, double mutant alleles in MYB-Related genes show very limited chloroplast development, and photosynthesis gene expression is perturbed to a greater extent than in mutants of GLK. InM. polymorphaMYB-related genes act upstream of GLK, while inA. thalianathis relationship has been rewired. In both species, genes encoding enzymes of chlorophyll biosynthesis are controlled by MYB-related and GLK proteins whilst those allowing CO2fixation, photorespiration and photosystem assembly and repair require the MYB-related proteins. Thus,MYB-relatedand GLK genes have overlapping as well as distinct targets. We conclude that together MYB-related and GLK transcription factors orchestrate chloroplast development in land plants.
Ton Motor Conformational Switch and Peptidoglycan Role in Bacterial Nutrient Uptake
Authors: Zinke Maximilian , Lejeune Maylis , Mechaly Ariel , Bardiaux Benjamin , Boneca Ivo Gomperts , Delepelaire Philippe , Izadi-Pruneyre Nadia 
Published: August 2023

ABSTRACTActive nutrient uptake is fundamental for survival and pathogenicity of Gram-negative bacteria, which operate a multi-protein Ton system to transport essential nutrients like metals and vitamins. This system harnesses the proton motive force at the inner membrane to energize the import through the outer membrane, but the mechanism of energy transfer remains enigmatic. Here, we study the periplasmic domain of ExbD, a crucial component of the proton channel of the Ton system. We show that this domain is a dynamic dimer switching between two conformations representing the proton channel’s open and closed states. Byin vivophenotypic assays we demonstrate that this conformational switch is essential for the nutrient uptake by bacteria. The open state of ExbD triggers a disorder to order transition of TonB, enabling TonB to supply energy to the nutrient transporter. We also reveal the anchoring role of the peptidoglycan layer in this mechanism. Herein, we propose a mechanistic model for the Ton system, emphasizing ExbD duality and the pivotal catalytic role of peptidoglycan. Sequence analysis suggests that this mechanism is conserved in other systems energizing gliding motility and membrane integrity. Our study fills important gaps in understanding bacterial motor mechanism and proposes novel antibacterial strategies.
Synergistic effects of inhibitors targeting PI3K and Aurora Kinase A in preclinical inflammatory breast cancer models
Authors: Al Ali Nadia , Kment Jacob , Young Stephanie , Craig Andrew W.B. 
Published: August 2023

AbstractBackgroundInflammatory breast cancer (IBC) is an aggressive clinical subtype of breast cancer often diagnosed in young women. Lymph node and distant metastases are frequently detected at diagnosis of IBC, and improvements in systemic therapies are needed. For IBC that lack hormone or HER2 expression, no targeted therapies are available. Since the phosphatidyl inositol 3’ kinase (PI3K) pathway is frequently deregulated in IBC, some studies have tested the pan PI3K inhibitor Buparlisib (BKM120). Although the SUM149 IBC cell line was resistant to Buparlisib, a functional genomic screen showed that silencing of Aurora kinase A (AURKA) sensitized cells to killing by Buparlisib. In this study, we tested whether combination treatments of PI3K and AURKA inhibitors act synergistically to kill IBC cells and tumors.MethodsSUM149 cells were treated with increasing doses of PI3K inhibitor Buparlisib (BKM120) and AURKA inhibitor Alisertib as monotherapies or combination therapies. Effects on target pathways, cytotoxicity, cell cycle, soft agar colony growth and cell migration were analyzed. The individual and combined treatments were also tested in a mammary orthotopic SUM149 tumor xenograft model to measure effects on tumor growth and metastasisResultsThe SUM149 IBC cell line treated with Buparlisib showed reduced PI3K/AKT activation but no significant skewing of cell cycle progression. Parallel studies of Alisertib treatment showed that AURKA inhibition led to a significant block in G2/M transition in SUM149 cells. In cytotoxicity assays, Buparlisib and Alisertib combination treatments were highly synergistic compared to monotherapy controls. Evidence of synergy between Buparlisib and Alisertib also extended to soft agar colony growth and wound healing motility in SUM149 cells. The combination of Buparlisib and Alisertib also reduced IBC tumor growth in mammary orthotopic xenograft assays and reduced spontaneous metastases development in lung tissue.ConclusionsAlthough SUM149 IBC cells were relatively resistant to killing by the PI3K inhibitor Buparlisib, our study showed that co-targeting the mitotic kinase AURKA with Alisertib synergized to limit IBC cell growth and motility, as well as IBC tumor growth and metastasis.
Ligand-DisplayingE. coliCells and Minicells for Programmable Delivery of Toxic Payloads via Type IV Secretion Systems
Authors: Li Yang Grace , Kishida Kouhei , Ogawa-Kishida Natsumi , Christie Peter J. 
Published: August 2023

ABSTRACTBacterial type IV secretion systems (T4SSs) are highly versatile macromolecular translocators and offer great potential for deployment as delivery systems for therapeutic intervention. One major T4SS subfamily, the conjugation machines, are well-adapted for delivery of DNA cargoes of interest to other bacteria or eukaryotic cells, but generally exhibit modest transfer frequencies and lack specificity for target cells. Here, we tested the efficacy of a surface-displayed nanobody/antigen (Nb/Ag) pairing system to enhance the conjugative transfer of IncN (pKM101), IncF (F/pOX38), or IncP (RP4) plasmids, or of mobilizable plasmids including those encoding CRISPR/Cas9 systems (pCrispr), to targeted recipient cells.Escherichia colidonors displaying Nb’s transferred plasmids toE. coliandPseudomonas aeruginosarecipients displaying the cognate Ag’s at significantly higher frequencies than to recipients lacking Ag’s. Nb/Ag pairing functionally substituted for the surface adhesin activities of F-encoded TraN and pKM101-encoded Pep, although not conjugative pili or VirB5-like adhesins. Nb/Ag pairing further elevated the killing effects accompanying delivery of pCrispr plasmids toE. coliandP. aeruginosatransconjugants bearing CRISPR/Cas9 target sequences. Finally, we determined that anucleateE. coliminicells, which are clinically safer delivery vectors than intact cells, transferred self-transmissible and mobilizable plasmids toE. coliandP. aeruginosacells. Minicell-mediated mobilization of pCrispr plasmids toE. colirecipients elicited significant killing of transconjugants, although Nb/Ag pairing did not enhance conjugation frequencies or killing. Together, our findings establish the potential for deployment of bacteria or minicells as Programmed Delivery Systems (PDSs) for suppression of targeted bacterial species in infection settings.IMPORTANCEThe rapid emergence of drug-resistant bacteria and current low rate of antibiotic discovery emphasize an urgent need for alternative antibacterial strategies. We engineeredEscherichia colito conjugatively transfer plasmids to specificE. coliandPseudomonas aeruginosarecipient cells through surface display of cognate nanobody/antigen (Nb/Ag) pairs. We further engineered mobilizable plasmids to carry CRISPR/Cas9 systems (pCrispr) for selective killing of recipient cells harboring CRISPR/Cas9 target sequences. In the assembled Programmed Delivery System (PDS), Nb-displayingE. colidonors with different conjugation systems and mobilizable pCrispr plasmids suppressed growth of Ag-displaying recipient cells to significantly greater extents than unpaired recipients. We also showed that anucleate minicells armed with conjugation machines and pCrispr plasmids were highly effective in killing ofE. colirecipients. Together, our findings suggest that bacteria or minicells armed with PDSs may prove highly effective as an adjunct or alternative to antibiotics for antimicrobial intervention.
Structural basis of closed groove scrambling by a TMEM16 protein
Authors: Feng Zhang , Alvarenga Omar E. , Accardi Alessio 
Published: August 2023

AbstractActivation of Ca2+-dependent TMEM16 scramblases induces the externalization of phosphatidylserine, a key molecule in multiple signaling processes. Current models suggest that the TMEM16s scramble lipids by deforming the membrane near a hydrophilic groove, and that Ca2+dependence arises from the different association of lipids with an open or closed groove. However, the molecular rearrangements involved in groove opening and of how lipids reorganize outside the closed groove remain unknown. Using cryogenic electron microscopy, we directly visualize how lipids associate at the closed groove of Ca2+-bound nhTMEM16 in nanodiscs. Functional experiments pinpoint the lipid-protein interaction sites critical for closed groove scrambling. Structural and functional analyses suggest groove opening entails the sequential appearance of two π-helical turns in the groove-lining TM6 helix and identify critical rearrangements. Finally, we show that the choice of scaffold protein and lipids affects the conformations of nhTMEM16 and their distribution, highlighting a key role of these factors in cryoEM structure determination.
Pan-antiviral effects of a PIKfyve inhibitor on respiratory virus infection in human nasal epithelium and mice
Authors: Baker Jonathan , Ombredane Hugo , Daly Leah , Knowles Ian , Rapeport Garth , Ito Kazuhiro 
Published: August 2023

ABSTRACTEndocytosis, or internalization through endosomes is a major cell entry mechanism used by respiratory viruses. Phosphoinositide 5-kinase (PIKfyve) is a critical enzyme for the synthesis of Phosphatidylinositol (3,5)biphosphate (PtdIns(3,5)P2), and has been implicated in virus trafficking via the endocytic pathway. In fact, antiviral effects of PIKfyve inhibitors against SARS-CoV-2 and Ebola have been reported, but there is little evidence regarding other respiratory viruses. In this study we demonstrated the antiviral effects of PIKfyve inhibitors on influenza virus and respiratory syncytial virusin vitroandin vivo. PIKfyve inhibitors, Apilimod mesylate (AM) and YM201636 concentration-dependently inhibited several influenza strains in a MDCK cell-cytopathic assay. AM also reduced the viral load and cytokine release, whilst improving the cell integrity of human nasal air liquid interface cultured epithelium infected with influenza PR8. In PR8-infected mice, AM (2mg/ml), when intranasally treated, exhibited significant reduction of viral load and inflammation and inhibited weight loss caused by influenza infection, with effects being similar to oral oseltamivir (10 mg/kg). In addition, AM demonstrated anti-viral effects in RSV A2 infected human nasal epitheliumin vitroand mousein vivo, with equivalent effect to that of ribavirin. AM also showed anti-viral effects against human rhinovirus and seasonal coronavirusin vitro. Thus, PIKfyve is found to be involved in influenza and RSV infection, and PIKfyve inhibitor is a promising molecule for pan-viral approach against respiratory viruses.
Acute or chronic depletion of macrophages in the dorsal root ganglia induces neuropathic pain after unilateral cervical spinal cord injury
Authors: Chhaya Soha J. , Richards Jonathan Houston , Giddings Grace A. , Detloff Megan Ryan 
Published: August 2023

AbstractThe inflammatory response at the spinal cord injury (SCI) epicenter and heightened macrophage presence in the dorsal root ganglia (DRG) has been well characterized after SCI and correlates with neuropathic pain. CCL2, a chemokine that acts as a macrophage chemoattractant and neuromodulator, is implicated in pain development, however, the role of the CCL2-CCR2 axis in the development of pain after SCI has not been explored. Here, we examined the role of CCL2-CCR2 signaling in macrophage recruitment to the DRG as well as the prolonged presence of macrophages in the DRG on the development and persistence of pain after SCI. Adult female Sprague-Dawley rats received a moderate, unilateral C5 contusion. Sandwich ELISA revealed that CCL2 is upregulated in the ipsilesional C7 and C8 DRGs in the first 24 hours post injury (hpi) and returns to naïve levels by 72 hpi. To prevent monocyte-derived macrophage recruitment to the DRG, additional SCI rats received vehicle or INCB3344, a CCR2 antagonist, intravenously at the time of SCI and at 24 and 48 hpi. INCB3344 administration induced transient forepaw allodynia at 7dpi in nearly all rats (88%) compared to only 33% in vehicle controls that resolves partially by 28 dpi, as measured by von Frey and mechanical conflict avoidance paradigms. As expected, qPCR analyses of whole DRG revealed that INCB3344 reduced macrophage markers and inflammatory cytokines in the ipsilesional C7 and C8 DRGs at 7 dpi compared to vehicle treated rats. By 28 dpi, there were no significant differences between INCB3344 or vehicle-treated groups, indicating that SCI-induced macrophage presence in the DRG is delayed by INCB3344 treatment. Moreover, gene expression of markers of macrophage polarity and cytokines suggest a pro-inflammatory environment in the DRG at 28dpi. DRG macrophage ablation via liposomal clodronate at 21dpi did not ameliorate hypersensitive pain behavior, though their ablation did reduce paw withdrawal thresholds in SCI rats that did not previously demonstrate pain behavior. Collectively, these data suggest that driving macrophages to a pro-reparative phenotype may be a viable and effective analgesic strategy that acts by modulating both the immune response and the experience of pain.
Conformational ensemble dependent lipid recognition and segregation by prenylated intrinsically disordered regions in small GTPases
Authors: Araya Mussie K. , Gorfe Alemayehu A. 
Published: August 2023

AbstractWe studied diverse prenylated intrinsically disordered regions (PIDRs) of Ras and Rho family small GTPases using long timescale atomistic molecular dynamics simulations in an asymmetric model membrane of phosphatidylcholine (PC) and phosphatidylserine (PS) lipids. We show that conformational plasticity is a key determinant of lipid sorting by polybasic PIDRs and provide evidence for lipid sorting based on both headgroup and acyl chain structures. We further show that conformational ensemble-based lipid recognition is generalizable to all polybasic PIDRs, and that the sequence outside the polybasic domain (PBD) modulates the conformational plasticity, bilayer adsorption, and interactions of PIDRs with membrane lipids. Specifically, we found that palmitoylation, the ratio of basic to acidic residues, and the hydrophobic content of the sequence outside the PBD significantly impact the diversity of conformational substates and hence the extent of conformation-dependent lipid interactions. We thus propose that the PBD is required but not sufficient for the full realization of lipid sorting by prenylated PBD-containing membrane anchors, and that the membrane anchor is not only responsible for high affinity membrane binding but also directs the protein to the right target membrane where it participates in lipid sorting.
Targeted Protein Degradation through Recruitment of the CUL4A Complex Adaptor Protein DDB1
Authors: Meyers Margot , Cismoski Sabine , Panidapu Anoohya , Chie-Leon Barbara , Nomura Daniel K. 
Published: August 2023

AbstractTargeted protein degradation has arisen as a powerful therapeutic modality for eliminating proteins. Thus far, most heterobifunctional Proteolysis Targeting Chimeras (PROTACs) have utilized recruiters against substrate receptors of Cullin RING E3 ubiquitin ligases, such as cereblon and VHL. However, previous studies have surprisingly uncovered molecular glue degraders that exploit a CUL4A adaptor protein DDB1 to degrade neosubstrate proteins. Here, we sought to investigate whether DDB1 recruiters can be discovered that can be exploited for PROTAC applications. We utilized activity-based protein profiling and cysteine chemoproteomic screening to identify a covalent recruiter that targets C173 on DDB1 and exploited this recruiter to develop PROTACs against BRD4 and androgen receptor (AR). We demonstrated that the BRD4 PROTAC results in selective degradation of the short BRD4 isoform over the long isoform in a proteasome, NEDDylation, and DDB1-dependent manner. We also demonstrated degradation of AR with the AR PROTAC in prostate cancer cells. Our study demonstrated that covalent chemoproteomic approaches can be used to discover recruiters against Cullin RING adapter proteins and that these recruiters can be used for PROTAC applications to degrade neo-substrates.