bluuebunny/crossref_metadata_2025 huggingface dataset:
DNA-encoded immunoassay in picoliter drops: a minimal cell-free approach
Authors: Jacková Barbara , Mottet Guillaume , Rudiuk Sergii , Morel Mathieu , Baigl Damien
Published: September 2022

Based on the remarkably specific antibody-antigen interaction, immunoassays have emerged as indispensable bioanalytical tools for both fundamental research and biomedical applications but necessitate long preliminary steps for the selection, production and purification of the antibody(ies) to be used. Here, we adopt a paradigm shift exploring the concept of creating a rapid and purification-free assay where the antibody is replaced by its coding DNA as a starting material, while exploiting a drop microfluidic format to dramatically decrease sample volume and accelerate throughput and sorting capability. The methodology consists in the co-encapsulation of a DNA coding for the variable domain of the heavy chain of heavy-chain only antibodies (VHH), a reconstituted cell-free expression medium, the target antigen and a capture scaffold where VHH:antigen accumulate to create a detectable signal, inside picoliter drop compartments. We first demonstrate successful synthesis of a functional hemagglutinin (HA)-tagged anti-GFP VHH, referred to as NanoGFP, at a high yield (15.3 ± 2.0 µg·mL-1) in bulk and in less than 3 h using PURExpress cell-free expression medium. We then use a microfluidic device to generate stable water-in-oil drops (30 pL) encapsulating NanoGFP-coding DNA, PURExpress medium, EGFP antigen and HA tag-specific magnetic nanoparticles prior to incubating at 37 °C the resulting emulsion under a magnetic field, inducing bothin situsynthesis of NanoGFP and accumulation of NanoGFP:EGFP complexes on magnetically assembled particles. This allows us to assess, for the first time and in less than 3 hours, the binding of an antigen to a cell-free synthesized antibody, in a large number of picoliter drops down to a DNA concentration as low as 12 plasmids per drop. We also show that the drops of this immunoassay can be further sequentially analyzed at high throughput (500 Hz), thus offering capability for library screening, sorting and/or rare event detection. We finally demonstrate the versatility of this method by using DNA coding for different VHH (e.g., anti-mCherry protein), by characterizing VHH specificity in the presence of antigen mixtures, and by showing that antigens can be either inherently fluorescent or not. We thus anticipate that the ultraminiaturized format (pL), rapidity (3 h), programmability (DNA-encoded approach) and versatility of this novel immunoassay concept will constitute valuable assets for faster discovery, better understanding and/or expanded applications of antibodies.
Femora from an exceptionally large population of coeval ornithomimosaurs yield evidence of sexual dimorphism in extinct theropod dinosaurs
Authors: Pintore R. , Cornette R. , Houssaye A. , Allain R.
Published: September 2022

Sexual dimorphism is challenging to detect among fossils, due to a lack of statistical representativeness. The Angeac-CharenteLagerstätte(France) represents a remarkable “snapshot” from a Berriasian (Early Cretaceous) ecosystem and offers a unique opportunity to study intraspecific variation among a herd of at least 61 coeval ornithomimosaurs. Herein, we investigated the hindlimb variation across the best-preserved specimens from the herd through 3D Geometric Morphometrics and Gaussian Mixture Modelling. Our results based on complete and fragmented femora evidenced a dimorphism characterized by variations in the shaft curvature and the distal epiphysis width. Since the same features vary between sexes among modern avian dinosaurs, crocodilians, and more distant amniotes, we attributed this bimodal variation to sexual dimorphism based on the extant phylogenetic bracketing approach. Documenting sexual dimorphism in fossil dinosaurs allows a better characterization and accounting of intraspecific variations, which is particularly relevant to address ongoing taxonomical and ecological questions relative to dinosaur evolution.
Bacteria evolve macroscopic multicellularity via the canalization of phenotypically plastic cell clustering
Authors: Chavhan Yashraj , Dey Sutirth , Lind Peter A.
Published: September 2022

The evolutionary transition from unicellular to multicellular life was a key innovation in the history of life. Given scarce fossil evidence, experimental evolution has been an important tool to study the likely first step of this transition, namely the formation of undifferentiated cellular clusters. Although multicellularity first evolved in bacteria, the extant experimental evolution literature on this subject has primarily used eukaryotes. Moreover, it focuses on mutationally driven (and not environmentally induced) phenotypes. Here we show that both Gram-negative and Gram-positive bacteria exhibit phenotypically plastic (i.e., environmentally induced) cell clustering. Under high salinity, they grow as elongated ~ 2 cm long clusters (not as individual planktonic cells). However, under habitual salinity, the clusters disintegrate and grow planktonically. We used experimental evolution withEscherichia colito show that such clustering can be canalized successfully: the evolved bacteria inherently grow as macroscopic multicellular clusters, even without environmental induction. Highly parallel mutations in genes linked to cell wall assembly formed the genomic basis of canalized multicellularity. While the wildtype also showed cell shape plasticity across high versus low salinity, it was either canalized or reversed after evolution. Interestingly, a single mutation could canalize multicellularity by modulating plasticity at multiple levels of organization. Taken together, we show that phenotypic plasticity can prime bacteria for evolving undifferentiated macroscopic multicellularity.
The DNA-relaxation-dependent Off-to-On biasing of the type 1 fimbrial genetic switch requires the Fis nucleoid-associated protein
Authors: Conway Colin , Beckett Michael C , Dorman Charles J
Published: September 2022

The structural genes expressing type 1 fimbriae in Escherichia coli alternate between expressed (phase ON) and non-expressed (phase OFF) states due to inversion of the 314-bp fimS genetic switch. The FimB tyrosine integrase inverts fimS by site-specific recombination, alternately connecting and disconnecting the fim operon, encoding the fimbrial subunit protein and its associated secretion and adhesin factors, to and from its transcriptional promoter within fimS. Site-specific recombination by the FimB recombinase becomes biased towards phase ON as DNA supercoiling is relaxed, a condition that occurs when bacteria approach the stationary phase of the growth cycle. This effect can be mimicked in exponential phase cultures by inhibiting the negative DNA supercoiling activity of DNA gyrase. We report that this bias towards phase ON depends on the presence of the Fis nucleoid-associated protein. We mapped the Fis binding to a site within the invertible fimS switch by DNase I footprinting. Disruption of this binding site by base substitution mutagenesis abolishes both Fis binding and the ability of the mutated switch to sustain its phase ON bias when DNA is relaxed, even in bacteria that produce the Fis protein. In addition, the Fis binding site overlaps one of the sites used by the Lrp protein, a known directionality determinant of fimS inversion that also contributes to phase ON bias. The Fis-Lrp relationship at fimS is reminiscent of that between Fis and Xis when promoting DNA-relaxation-dependent excision of bacteriophage λ from the E. coli chromosome. However, unlike the co-binding mechanism used by Fis and Xis at λ attR, the Fis-Lrp relationship at fimS involves competitive binding. We discuss these findings in the context of the link between fimS inversion biasing and the physiological state of the bacterium.
N6-Adenosine methylation regulates the translation of insulin mRNA
Authors: Wilinski Daniel , Dus Monica
Published: September 2022

Relatively little is known about the first step of insulin synthesis: the translation of the mRNA. Here we show that the translation of D. melanogaster insulin 2 mRNA (dilp2) is controlled by methylation of N6-adenosine (m6A) in the 3’ UTR. Mutations in the m6A writer Mettl3 and methylated-residues in the dilp2 3’UTR decreased the levels of dilp2 mRNA associated with the polysomes, and the total amount of dilp2 protein produced. This resulted in aberrant energy homeostasis and diabetic-like phenotypes, consistent with the specific function of dilp2 in adult metabolism. Conserved m6A signatures were also identified in the 3’ UTRs of vertebrate insulin mRNAs. These data identify m6A as a key regulator of insulin protein synthesis and energy homeostasis in metazoans and demonstrate an essential role for m6A in translation, with important implications for diabetes and metabolic disease.

One-Sentence Summary

The most abundant modification in eukaryotic mRNAs controls the synthesis of insulin protein in D. melanogaster.
Peroxisome biogenesis initiated by protein phase separation
Authors: Ravindran Rini , Bacellar Isabel O. L. , Castellanos-Girouard Xavier , Zhang Zhenghao , Kisley Lydia , Michnick Stephen W.
Published: September 2022

Peroxisomes are organelles that perform beta-oxidation of fatty acids and amino acids. Both rare and prevalent diseases are caused by their disfunction1. Among disease-causing mutant genes are those required for protein transport into the peroxisome. The peroxisomal protein import machinery, also shared with chloroplasts, is unique in transporting folded and large, up to 10 nm in diameter, protein complexes into peroxisomes2 and current models postulate a large pore formed by transmembrane proteins3. To date, however, no pore structure has been observed. In the budding yeast Saccharomyces cerevisiae, the minimum transport machinery includes membrane proteins Pex13 and Pex14 and cargo protein-binding transport receptor, Pex5. Here we show that Pex13 undergoes liquid-liquid phase separation (LLPS) with Pex5-cargo. Intrinsically disordered regions (IDR) in Pex13 and Pex5 resemble those found in nuclear pore complex (NPC) proteins. Cargo transport into peroxisomes depends on the number but not patterns of aromatic residues in these IDRs, consistent with their roles as ‘stickers’ in associative polymer models of LLPS4,5. Finally, imaging Fluorescence Cross-Correlation Spectroscopy (iFCCS) shows that the transport of cargo correlates with transient focusing of GFP-Pex13/14 on the peroxisome membrane. Pex13 and Pex14 form foci in distinct time-frames, suggesting that they may form channels at different saturating concentrations of Pex5-cargo. Our results suggest a model in which LLPS of Pex5-cargo with Pex13/14 results in transient protein transport channels.
Pseudomonas aeruginosa FpvB is a high-affinity transporter for xenosiderophores ferrichrome and ferrioxamine B
Authors: Chan Derek C. K. , Burrows Lori L.
Published: September 2022

Iron is essential for many biological functions in bacteria but its poor solubility is a limiting factor for growth. Bacteria produce siderophores, soluble natural products that bind iron with high affinity, to overcome this challenge. Siderophore-iron complexes return to the cell through specific outer-membrane transporters. The opportunistic pathogen Pseudomonas aeruginosa makes multiple transporters that recognize its own siderophores, pyoverdine and pyochelin, and xenosiderophores produced by other bacteria, which gives it a competitive advantage. Some antibiotics exploit these transporters to bypass the membrane to reach their intercellular targets – including the thiopeptide antibiotic, thiostrepton (TS), which uses the pyoverdine transporters FpvA and FpvB to cross the outer membrane. Here, we assessed TS susceptibility in the presence of various siderophores and discovered that ferrichrome and ferrioxamine B antagonized TS uptake via FpvB. Unexpectedly, we found that FpvB transports ferrichrome and ferrioxamine B with higher affinity than pyoverdine. Site-directed mutagenesis of FpvB coupled with competitive growth inhibition and affinity label quenching studies suggested that the siderophores and antibiotic share a common binding site in an aromatic pocket formed by the plug and barrel domains, but have differences in their binding mechanism and molecular determinants for uptake. This work describes an alternative uptake pathway for ferrichrome and ferrioxamine B in P. aeruginosa and emphasizes the promiscuity of siderophore transporters, with implications for Gram-negative antibiotic development via the Trojan horse approach.
Loss of the first β-strand of human prion protein generates an aggregation-competent partially “open” form
Authors: Hosszu Laszlo L. P. , Sangar Daljit , Batchelor Mark , Risse Emmanuel , Hounslow Andrea M. , Waltho Jonathan P. , Collinge John , Bieschke Jan
Published: September 2022

Prion diseases, a group of incurable, lethal neurodegenerative disorders of mammals including humans, are caused by prions, assemblies of misfolded host prion protein (PrP). The pathway of PrP misfolding is still unclear, though previous data indicate the presence of a structural core in cellular PrP (PrPC), whose cooperative unfolding presents a substantial energy barrier on the path to prion formation. PrP is a GPI-anchored membrane protein, and a number of studies suggest that membrane interactions play an important role in the conversion of PrPC to its disease-associated form, including a transmembrane form of PrP in which a highly conserved region (residues 110 - 136) spans the ER membrane. Insertion of this region results in the detachment of the PrPC first β-strand from the structural core. The effect of this removal on the structure, stability and self-association of the folded domain of PrPC is determined here through a biophysical characterisation of a truncated form of PrPC lacking this region. Whilst markedly destabilised, NMR chemical shifts show that the truncated protein exhibits tertiary structure characteristic of a fully folded protein and retains its native secondary structure elements, including the second strand of the PrP β-sheet, but with altered conformational flexibility in the β2-α2 loop and first α-helix. The latter is destabilised relative to the other helical regions of the protein, with markedly increased solvent exposure. This truncated form of PrP fibrilises more readily than the native form of the protein. These data suggest a stepwise mechanism, in which a destabilised “open” form of PrPC may be a key intermediate in the refolding to the fibrillar, pathogenic form of the protein.
Conserved signaling pathways antagonize and synergize with co-opteddoublesexto control development of novel mimetic butterfly wing patterns
Authors: VanKuren Nicholas W. , Doellman Meredith M. , Sheikh Sofia I. , Palmer Droguett Daniela H. , Massardo Darli , Kronforst Marcus R.
Published: September 2022

Novel phenotypes are increasingly recognized to have evolved by co-option of conserved genes into new developmental contexts, yet the impact of co-option on existing developmental programs remains obscure. Here we provide insight into this process by characterizing the consequences ofdoublesexco-option on wing color pattern development inPapilioswallowtail butterflies. doublesexis the master regulator of insect sex differentiation but has been co-opted to control the switch between discrete mimetic and non-mimetic, male-like color patterns inPapilio polytesand its close relatives. Here we show that development of the mimetic color pattern inP. polytesis caused by a pulse ofdsxexpression early in female wing development that results in a corresponding pulse of differential expression that both alters color pattern development and quickly becomes decoupled fromdsxexpression itself. Differentially expressed genes were enriched in canonical Wnt and Hedgehog signaling pathway genes, but case studies of key genes using RNAi and antibody stains suggested opposing, novel roles for the two pathways in mimetic color pattern development. The pulse of Dsx expression caused Engrailed, the key transcription factor effector of Hh signaling, to gain anterior expression in early pupal wing development. However, Dsx and En became decoupled by mid-pupal development when En pre-figured melanic and red patterns and Dsx pre-figured white patterns. In contrast, Wnt signaling antagonizes Dsx in restricted regions of the wing to refine the mimetic color pattern. Our results therefore provide strong experimental evidence thatdsxco-option significantly altered spatiotemporal activities of conserved wing patterning pathways to promote and refine the development of a novel adaptive color pattern. Altogether, our findings provide strong evidence for how co-opted genes can both cause and elicit changes to established gene regulatory networks during the evolution and development of novel phenotypes.
Critical role of growth medium for detecting drug interactions in Gram-negative bacteria that model in vivo responses
Authors: Davis Kathleen P. , Morales Yoelkys , McCabe Anne L. , Mecsas Joan , Aldridge Bree B.
Published: September 2022

The rise in infections caused by multidrug-resistant (MDR) bacteria has necessitated a variety of clinical approaches, including the use of antibiotic combinations. Antibiotic susceptibility is affected in part by the growth state of bacteria within various tissues. Here we tested the hypothesis that drug-drug interactions vary in different media, and hence, using a medium that reflects tissue environments will better predict in vivo outcomes. We systematically studied pair-wise antibiotic interactions in three different media (CAMHB, a urine mimetic, and a lung mimetic) using three Gram-negative ESKAPE pathogens, Acinetobacter baumannii (Ab), Klebsiella pneumoniae (Kp), and Pseudomonas aeruginosa (Pa). There were pronounced differences in responses to antibiotic combinations between the three bacterial species grown in the same medium. However, within species, Pa responded to drug combinations similarly when grown in all three different media, whereas Ab responded similarly when grown in CAMHB and a lung mimetic medium. By contrast, drug interactions in Kp were poorly correlated across three different media. To assess whether distinct media were predictive of antibiotic interactions in Kp in the lungs of mice, we developed a treatment strategy and tested three antibiotic combination pairs. Measurements obtained in vitro from lung mimetic medium, but not rich medium, predicted in vivo outcomes. This work demonstrates that antibiotic interactions are highly variable when comparing across three gram-negative pathogens and highlights the importance of growth medium by showing a superior correlation between in vitro interactions in a growth medium that resembles the tissue environment and in vivo outcomes.
Striatal dopamine integrates cost, benefit and motivation
Authors: Eshel Neir , Touponse Gavin C. , Wang Allan R. , Osterman Amber K. , Shank Amei N. , Groome Alexandra M. , Taniguchi Lara , Cardozo Pinto Daniel F. , Tucciarone Jason , Bentzley Brandon S. , Malenka Robert C.
Published: September 2022

Dopamine (DA) release in the ventral and dorsal striatum has been linked to reward processing and motivation, but there are longstanding controversies about whether DA release in these key target structures primarily reflects costs or benefits, and how these signals vary with motivation. Here we apply behavioral economic principles to generate demand curves for rewards while directly measuring DA release in the nucleus accumbens (NAc) and dorsolateral striatum (DLS) via a genetically-encoded sensor. By independently varying costs and benefits, we reveal that DA release in both structures incorporates reward magnitude and sunk cost. Surprisingly, motivation was inversely correlated with reward-evoked DA release; the higher the motivation for rewards the lower the reward-evoked DA release. These relationships between DA release, cost and motivation remained identical when we used optogenetic activation of striatal DA inputs as a reward. Our results reconcile previous disparate findings by demonstrating that during operant tasks, striatal DA release simultaneously encodes cost, benefit and motivation but in distinct manners over different time scales.
MR1 dependent MAIT cell activation is regulated by autophagy associated proteins
Authors: Phalora Prabhjeet , Ussher James , Hester Svenja , Marchi Emanuele , Mak Jeffrey Y. W. , Fairlie David P. , Klenerman Paul
Published: September 2022

The antigen presenting molecule, MR1, presents microbial metabolites to MAIT cells, a population of innate-like, anti-microbial T cells. It also presents an unidentified ligand to MR-1 restricted T cells in the setting of cancer. The cellular co-factors that mediate MR1 antigen presentation have yet to be fully defined. We performed a mass spectrometry-based proteomics screen to identify MR1 interacting proteins and identified the selective autophagy receptor SQSTM1/p62. CRISPR-Cas9-mediated knock out of SQSTM1/p62 increased MAIT cell activation in the presence of E.coli but not the synthetic ligand 5-OP-RU whereas depletion of Atg5 and Atg7, key autophagy proteins, increased MAIT activation irrespective of the ligand used. This regulation appears to occur at an early step in the trafficking pathway. This data implicates distinct roles for autophagy associated proteins in the regulation of MR1 activity and highlights the autophagy pathway as a key regulator of cellular antigen presentation.
Functionally conserved Pkd2, mutated in autosomal dominant polycystic kidney disease, localizes to the endoplasmic reticulum and regulates cytoplasmic calcium homeostasis in fission yeast
Authors: Koyano Takayuki , Kume Kazunori , Onishi Kaori , Matsuyama Makoto , Fukushima Masaki , Toda Takashi
Published: September 2022

Mutations in PKD1 or PKD2 genes lead to autosomal dominant polycystic kidney disease (ADPKD) that is the most frequent family inherited renal disorder. These genes encode polycystin-1/PC-1 and polycycstin-2/PC-2, respectively. Although the genetic basis of ADPKD is well established, the crucial functions of polycystins underlying onset and development of cyst formation remain elusive. Fission yeast Schizosaccharomyces pombe has a single polycystin homolog, Pkd2, which is essential for cell growth. In this study, the truncation analyses of Pkd2 reveal that Pkd2 localizes to not only the plasma membrane but also the endoplasmic reticulum (ER) and regulates cytoplasmic calcium signaling in fission yeast. Internal transmembrane domains within Pkd2 are sufficient for these processes. Surprisingly, more than half of Pkd2 is not required for cell viability. Cytoplasmic calcium levels are mainly regulated through C-terminus of Pkd2. Importantly, human Pkd2 also localizes to the ER and furthermore, fully complements the loss of fission yeast Pkd2. As the functions of polycystin-2 are conserved, fission yeast provides a suitable model to study the mechanism of ADPKD as well as polycystins.
Gut inflammation associated with age and Alzheimer’s disease pathology
Authors: Heston Margo B. , Hanslik Kendra L. , Zarbock Katie R. , Harding Sandra J. , Davenport-Sis Nancy J. , Kerby Robert L. , Chin Nathaniel , Sun Yi , Hoeft Ana , Deming Yuetiva , Vogt Nicholas M. , Betthauser Tobey J. , Johnson Sterling C. , Asthana Sanjay , Kollmorgen Gwendlyn , Suridjan Ivonne , Wild Norbert , Zetterberg Henrik , Blennow Kaj , Rey Federico E. , Bendlin Barbara B. , Ulland Tyler K.
Published: September 2022

Age-related disease may be mediated by low levels of chronic inflammation (“inflammaging”). Recent work suggests that gut microbes may contribute to inflammation via degradation of the intestinal barrier. While aging and age-related diseases including Alzheimer’s disease (AD) are linked to altered microbiome composition and higher levels of gut microbial components in systemic circulation, the role of intestinal inflammation and permeabilityper seremains unclear. To test whether greater gut inflammation is associated with older age and AD pathology, we assessed fecal samples from older adults to measure calprotectin, an established marker of intestinal inflammation which is elevated in diseases of gut barrier integrity. Here we found that calprotectin levels are higher with age, and that higher calprotectin was associated with greater amyloid burden among participants with an amyloid-confirmed AD dementia diagnosis. Calprotectin was also associated with cerebrospinal fluid markers of AD pathology and axonal degeneration, as well as with lower verbal memory function among cognitively unimpaired participants. Together, these findings suggest that intestinal inflammation may play a role in pathology development, and that it may exacerbate the progression toward AD.

Intestinal inflammation is correlated with older age, Alzheimer’s disease (AD) dementia, and greater amyloid burden in participants with AD.
Comprehensive analysis of immune responses in CLL patients after heterologous COVID-19 vaccination
Authors: Lee Hye Kyung , Hoechstetter Manuela A. , Buchner Maike , Pham Trang Thu , Huh Jin Won , Müller Katharina , Zange Sabine , Buttlar Heiner von , Girl Philipp , Wölfel Roman , Brandmeier Lisa , Pfeuffer Lisa , Furth Priscilla A. , Wendtner Clemens-Martin , Hennighausen Lothar
Published: September 2022

Patients with chronic lymphocytic leukemia (CLL) treated with B-cell pathway inhibitors and anti-CD20 antibodies exhibit low humoral response rate (RR) following SARS-CoV-2 vaccination. To investigate the relationship between the initial transcriptional response to vaccination with ensuing B and T cell immune responses, we performed a comprehensive immune transcriptome analysis flanked by antibody and T cell assays in peripheral blood prospectively collected from 15 CLL/SLL patients vaccinated with heterologous BNT162b2/ChAdOx1 with follow up at a single institution. The two-dose antibody RR was 40% increasing to 53% after booster. Patients on BTKi, venetoclax ± anti-CD20 antibody within 12 months of vaccination responded less well than those under BTKi alone. The two-dose T cell RR was 80% increasing to 93% after booster. Transcriptome studies revealed that seven patients showed interferon-mediated signaling activation within 2 days and one at 7 days after vaccination. Increasing counts of COVID-19 specific IGHV genes correlated with B-cell reconstitution and improved humoral RR. T cell responses in CLL patients appeared after vaccination regardless of treatment status. A higher humoral RR was associated with BTKi treatment and B-cell reconstitution. Boosting was particularly effective when intrinsic immune status was improved by CLL-treatment.
An expanded genetic toolbox to accelerate the creation ofAcholeplasma laidlawiidriven by synthetic genomes
Authors: Nucifora Daniel P. , Mehta Nidhi D. , Giguere Daniel J. , Karas Bogumil J.
Published: September 2022

Assembling synthetic bacterial genomes in yeast and genome transplantation has enabled an unmatched level of bacterial strain engineering, giving rise to cells with minimal and chemically synthetic genomes. However, this technology is currently limited to members of the Spiroplasma phylogenetic group, mostlyMycoplasmas, within theMollicuteclass. Here, we propose new genetic tools for developing these technologies forAcholeplasma laidlawii, which is phylogenetically distant fromMycoplasmasand, unlike mostMollicutes, uses a standard genetic code. We first investigated a donor-recipient relationship between twoA. laidlawiistrains through whole-genome sequencing. We then created multi-host shuttle plasmids and used them to optimize an electroporation protocol. We also demonstrated the use of evolution to create superior strains for DNA uptake via electroporation. For genome transplantation, we selectedA. laidlawii8195 as the recipient strain and created a PG-8A donor strain by inserting a Tn5 transposon carrying a tetracycline resistance gene. The tools presented here will improveAcholeplasmaresearch and accelerate the effort toward creatingA. laidlawiistrains driven by synthetic genomes.

Graphical Abstract
Metamorphic proteins at the basis of human autophagy initiation and lipid transfer
Authors: Nguyen Anh , Lugarini Francesca , David Céline , Hosnani Pouya , Knotkova Barbora , Patel Anoshi , Parfentev Iwan , Friedrich Annabelle , Urlaub Henning , Meinecke Michael , Stork Björn , Faesen Alex C.
Published: September 2022

Autophagy is a conserved intracellular degradation pathway that uses de novo doublemembrane vesicle (autophagosome) formation to target a wide range of cytoplasmic material for lysosomal degradation. In multicellular organisms, autophagy initiation requires the timely assembly of a contact site between the ER and the nascent autophagosome. Here, we report the in vitro reconstitution of a full-length sevensubunit human autophagy initiation supercomplex and found at its core ATG13-101 and transmembrane protein ATG9. Assembly of this core complex requires the rare ability of ATG13 and ATG101 to adopt topologically distinct folds. The slow spontaneous conversion between folds creates a rate-limiting step to regulate selfassembly of the super-complex. The interaction of the core complex with ATG2-WIPI4 enhances tethering of membrane vesicles and accelerates lipid transfer of ATG2 by both ATG9 and ATG13-101. Our work uncovers the molecular basis of the contact site and its assembly mechanisms imposed by the metamorphosis of ATG13-101 to regulate autophagosome biogenesis in space and time.
Sequential Activation and Local Unfolding Control Poly(A)-Binding Protein Condensation
Authors: Chen Ruofan , Kahan Darren , Shangguan Julia , Sachleben Joseph R. , Riback Joshua A. , Drummond D. Allan , Sosnick Tobin R.
Published: September 2022

Eukaryotic cells form biomolecular condensates to sense and adapt to their environment1,2. Poly(A)-binding protein (Pab1), a canonical stress granule marker3,4, condenses upon heat shock or starvation, promoting adaptation5. The molecular basis of condensation has remained elusive due to a dearth of techniques to probe structure directly in condensates. Here we apply hydrogen-deuterium exchange/mass spectrometry (HDX-MS) to investigate the molecular mechanism of Pab1’s condensation. We find that Pab1’s four RNA recognition motifs (RRMs) undergo different levels of partial unfolding upon condensation, and the changes are similar for thermal and pH stresses. Although structural heterogeneity is observed, the ability of MS to describe individual subpopulations allows us to identify which regions become partially unfolded and contribute to the condensate’s interaction network. Our data yield a clear molecular picture of Pab1’s stress-triggered condensation, which we term sequential activation, wherein each RRM becomes activated at a temperature where it partially unfolds and associates with other likewise activated RRMs to form the condensate. This model thus implies that sequential activation is dictated by the underlying free energy surface, an effect we refer to as thermodynamic specificity. Our study represents a methodological advance for elucidating the interactions that drive biomolecular condensation that we anticipate will be widely applicable. Furthermore, our findings demonstrate how condensation can use thermodynamic specificity to perform an acute response to multiple, stresses, a potentially general mechanism for stress-responsive proteins.
LEAP2 is a more conserved ligand than ghrelin for fish GHSRs
Authors: Li Hao-Zheng , Shao Xiao-Xia , Wang Ya-Fen , Liu Ya-Li , Xu Zeng-Guang , Guo Zhan-Yun
Published: September 2022

Recently, liver-expressed antimicrobial peptide 2 (LEAP2) was identified as an endogenous competitive antagonist and an inverse agonist of the ghrelin receptor GHSR. However, its functions in lower vertebrates are not well understood. Our recent study demonstrated that both LEAP2 and ghrelin are functional towards a fish GHSR from Latimeria chalumnae, an extant coelacanth believed to be one of the closest ancestors of tetrapods. However, amino acid sequence alignment identified that the 6.58 position (Ballesteros-Weinstein numbering system) of most fish GHSRs are not occupied by an aromatic Phe residue, which is absolutely conserved in all known GHSRs from amphibians to mammals, and is responsible for human GHSR binding to its agonist, ghrelin. To test whether these unusual fish receptors are functional, we studied the ligand binding properties of three representative fish GHSRs, two from Danio rerio (zebrafish) and one from Larimichthys crocea (large yellow croaker). After overexpression in human embryonic kidney 293T cells, the three fish GHSRs retained normal binding to all tested LEAP2s, except for a second LEAP2 from L. crocea. However, they displayed almost no binding to all chemically synthesized n-octanoylated ghrelins, despite these ghrelins all retaining normal function towards human and coelacanth GHSRs. Thus, it seems that LEAP2 is a more conserved ligand than ghrelin towards fish GHSRs. Our results not only provided new insights into the interaction mechanism of GHSRs with LEAP2s and ghrelins, but also shed new light on the functions of LEAP2 and ghrelin in different fish species.
Online environmentally adaptive trajectory planning for rotorcraft unmanned aerial vehicles
Authors: Tong Chunming , Liu Zhenbao , Dang Qingqing , Wang Jingyan , Cheng Yao
Published: September 2022

Purpose

This paper aims to propose an environmentally adaptive trajectory planning system considering the dynamic characteristics of unmanned aerial vehicles (UAVs) and the distance between obstacles and the UAV. The system generates a smooth and safe flight trajectory online.

Design/methodology/approach

First, the hybrid A* search method considering the dynamic characteristics of the quadrotor is used to find the collision-free initial trajectory. Then, environmentally adaptive velocity cost is designed for environment-adaptive trajectory optimization using environmental gradient data. The proposed method adaptively adjusts the autonomous flight speed of the UAV. Finally, the initial trajectory is applied to the multi-layered optimization framework to make it smooth and dynamically viable.

Findings

The feasibility of the designed system is validated by online flight experiments, which are in unknown, complex situations.

Practical implications

The proposed trajectory planning system is integrated into a vision-based quadrotor platform. It is easily implementable onboard and computationally efficient.

Originality/value

A hybrid A* path searching method is proposed to generate feasible motion primitives by dispersing the input space uniformly. The proposed method considers the control input of the UAV and the search time as the heuristic cost. Therefore, the proposed method can provide an initial path with the minimum flying time and energy loss that benefits trajectory optimization. The environmentally adaptive velocity cost is proposed to adaptively adjust the flight speed of the UAV using the distance between obstacles and the UAV. Furthermore, a multi-layered environmentally adaptive trajectory optimization framework is proposed to generate a smooth and safe trajectory.
A strategy of the longitudinal control for the tandem wing configuration design
Authors: Figat Marcin
Published: September 2022

Purpose

This paper presents first sight on the longitudinal control strategy for an aircraft in the tandem wing configuration. It is an aerodynamic strongly coupled configuration that needs a lot of detailed aerodynamic analysis which describes the mutual impact of the main parts of the aircraft. The purpose of this paper is to build the numerical model that allows to make an analysis of necessary flaps (front and rear) deflection and prepare the control strategy for this kind of aircraft.

Design/methodology/approach

Aircrafts’ aerodynamic characteristics were obtained using the MGAERO software which is a commercial computing fluid dynamics tool created by Analytical Methods, Inc. This software uses the Euler flow model. Results from this software were used in the static stability evaluation and trim condition analysis. The trim conditions are the outcome of the optimisation process whose goal was to find the best front and rear flap deflection to achieve the best lift to drag (L/D) ratio.

Findings

The main outcome of this investigation is the proposal of strategy for the front and rear flap deflection which ensured the maximum L/D ratio and satisfied the trim condition. Moreover, the analysis of the mutual impact of the front and rear wings and the analysis of the control surface impact on the aerodynamic characteristic of the aircraft are presented.

Research limitations/implications

In terms of aerodynamic computation, MGAERO software uses an inviscid flow model. However, this research is for the conceptual stage of the design and the MGAERO software grantee satisfied accurate respect to relatively low time of computations.

Practical implications

The ultimate goal is to build an aircraft in a tandem wing configuration and to conduct flying tests or wind tunnel tests. The presented result is one of the milestones to achieve this goal.

Originality/value

The aircraft in the tandem wing configuration is an aerodynamic-coupled configuration that needs detailed analysis to find the mutual interaction between the front and rear wings. Moreover, the mutual impact of the front and rear flaps is necessary too. Obtaining these results allowed this study to build the numerical model of the aircraft in the tandem wing configuration. It allows to find the best strategy of flap deflection, which allows to obtain the maximum L/D ratio and satisfy the trim condition.
Sociocultural factors that affect the identification and development of talent in children and adolescents
Authors: Olszewski-Kubilius Paula , Subotnik Rena F. , Worrell Frank C.
Published: September 2022

Purpose

The purpose of this paper is to inform readers about the nature of talent development prior to post-secondary education; describe the obstacles that individuals face because of poverty, racism or geography; and recommend asset-based approaches that can enable more individuals to be prepared to make significant contributions to society within their domain of talent.

Design/methodology/approach

The methodology used was to review research from the fields of education and psychology about talent in varied domains of sport, academics and the arts, as it relates to key components (domain pathways, opportunities and psychosocial skills) of the talent development megamodel proposed by Subotnik et al. (2011).

Findings

Findings include a delineation of the challenges that many nations face in cultivating talent among its young citizens particularly related to their socioeconomic status, race and ethnicity and geography. Findings include recommendations for new approaches to identification; a substantial increase in school and community-based, domain-specific opportunities; teacher training; and deliberate cultivation of psychosocial skills that can support achievement.

Originality/value

This paper emphasizes the importance of focusing efforts on talent development at earlier stages, which is critical to creating pathways for marginalized youths to maximize their potential and contributions to the workplace.
Explosion behaviors of IPN/air mixture at high temperature and high pressure
Authors: Wan Hang-Wei , Wen Yu-Quan , Zhang Qi
Published: September 2022

Purpose

The reaction dynamics of combustible clouds at high temperatures and pressures are a common form of energy output in aerospace and explosion accidents. The cloud explosion process is often affected by the external initial conditions. This study aims to numerically study the effects of airflow velocity, initial temperature and fuel concentration on the explosion behavior of isopropyl nitrate/air mixture in a semiconstrained combustor.

Design/methodology/approach

The discrete-phase model was adopted to consider the interaction between the gas-phase and droplet particles. A wave model was applied to the droplet breakup. A finite rate/eddy dissipation model was used to simulate the explosion process of the fuel cloud.

Findings

The peak pressure and temperature growth rate both decrease with the increasing initial temperature (1,000–2,200 K) of the combustor at a lower airflow velocity. The peak pressure increases with the increase of airflow velocity (50–100 m/s), whereas the peak temperature is not sensitive to the initial high temperature. The peak pressure of the two-phase explosion decreases with concentration (200–1,500 g/m3), whereas the peak temperature first increases and then decreases as the concentration increases.

Practical implications

Chain explosion reactions often occur under high-temperature, high-pressure and turbulent conditions. This study aims to provide prevention and data support for a gas–liquid two-phase explosion.

Originality/value

Sustained turbulence is realized by continuously injecting air and liquid fuel into a semiconfined high-temperature and high-pressure combustor to obtain the reaction dynamic parameters of a two-phase explosion.
Impact of COVID-19 on convergence in Indian districts
Authors: Chakrabarty Manisha , Mukherjee Subhankar
Published: September 2022

Purpose

The purpose of this paper is to estimate the impact of the COVID-19 pandemic on the patterns of convergence/divergence among the districts in India. Specifically, this paper investigates if the impact is heterogeneous among different cohorts of districts (based on income distribution). The differential impact may lead to heterogeneous long-run growth paths, resulting in unbalanced development across regions within the country. A study of convergence can ascertain the possible trajectory of such development across regions. Investigation of this phenomenon is the primary aim of this study.

Design/methodology/approach

This paper uses the panel regression method for estimation. This paper uses high-frequency nighttime light intensity data as a proxy for aggregate output.

Findings

The authors observe a significant reduction in the convergence rate as a result of the pandemic. Across the cluster of districts, the drop in ß-convergence rate, compared to the pre-pandemic period, varied from approximately 33% for the poorer districts to close to zero for the richest group of districts. These findings suggest that the pandemic may lead to a wider disparity among different regions within the country.

Originality/value

This paper contributes to the literature in the following ways. First, to the best of the authors’ knowledge, this is the first paper to investigate the impact of COVID-19 on the convergence rate. A detailed look into the possible disparity in convergence among various regions is critical because a larger drop in convergence, especially among the poorer regions, may call for policy attention to attain long-term equitable development. The authors perform this exercise by dividing the districts into four quantile groups based on the distribution of night-light intensity. Second, while previous studies on convergence using nighttime light data have used a cross-sectional approach, this study is possibly the first attempt to use the panel regression method on this data. The application of this method can be useful in tackling district-level omitted variables bias. Finally, the heterogeneity analysis using different quantiles of the distribution of night-light intensity may help in designing targeted policies to mitigate the disparity across districts due to the shock.
Loneliness in the workplace: feelings and expressivity of hotel employees
Authors: Dedeoğlu Bekir Bora , Çalışkan Caner , Chen Tzu-Ling , Borzyszkowski Jacek , Okumus Fevzi
Published: September 2022

Purpose

This study investigates the relationship between feelings of loneliness in the workplace, life satisfaction, affect, hope and expressivity among hotel employees.

Design/methodology/approach

The research model was tested via structural equation modeling based on the empirical data collected from hotel employees in Antalya, Turkey.

Findings

The research findings suggest that emotional deprivation and social companionship have a significant impact on life satisfaction, that life satisfaction has a significant impact on positive and negative emotions, and that positive and negative emotions have the same impact on pathways and agencies.

Originality/value

The research findings should assist researchers and practitioners to understand the behaviors of hotel employees in continuous interaction and relationship with individuals to motivate them while providing more effective services.
Public policy responses to COVID-19 and the survival of ethnic minority businesses (EMBs): does entrepreneurial orientation (EO) matter?
Authors: Kabir Ibrahim , Abubakar Yazid Abdullahi
Published: September 2022

Purpose

The global economic crisis triggered by the coronavirus disease 2019 (COVID-19) has caused the closure of countless ethnic minority businesses (EMBs) worldwide, partly due to the public policy responses. This paper investigates whether EMBs’ entrepreneurial orientation (EO) mediates the impact of public policy responses to COVID-19 on their survival.

Design/methodology/approach

Utilizing institutional theory, the authors developed a novel conceptual framework that divides policy responses to COVID-19 into aggressive (imposing restrictions on movement, e.g. lockdowns) and less aggressive policy responses (not imposing restrictions on movement, e.g. social distancing). The authors then surveyed intra-regional EMBs, specifically businesses owned by ethnic minorities in the Kano and Katsina provinces of Nigeria, and analysed the data using structural equation modelling and analysis of variance (ANOVA).

Findings

The authors found that intra-regional EMBs in developing countries are very vulnerable to the public policy responses imposed by governments to curb COVID-19. Aggressive policy responses have a more significant negative effect on the survival of intra-regional EMBs than their less aggressive counterparts. Furthermore, the authors found that EO as a crisis response strategy significantly supports intra-regional EMBs in managing their vulnerability to the hostile institutional environment, reduces the adverse effect of public policy responses and stimulates their survival during the COVID-19 pandemic.

Originality/value

This paper contributes to the institutional theory of small and medium-sized enterprises (SMEs)/entrepreneurship and the literature on EMBs by showing the role of EO in mediating the effects of COVID-19 institutional policies on the survival of intra-regional EMBs.
Audit quality and liquidity policy
Authors: Hendijani Zadeh Mohammad
Published: September 2022

Purpose

The purpose of this study is to examine whether audit quality influences auditees' liquidity policy.

Design/methodology/approach

The author uses ordinary least squares (OLS) estimators, and we focus on a panel of US publicly traded companies (36,118 company-year observations) over the period of 2004–2019 to examine the effect of audit quality on auditees' cash reserves.

Findings

The author finds that high quality audits are negatively related to auditees' cash reserves. Additional analyses show that the potential channel by which audit quality influences these reserves is financial constraints (FC). Particularly, his results suggest that an auditee's FC serve as an intermediary in the association between audit quality and auditee's cash reserves. Ultimately, we show that high quality audits raise the market value relevance of an extra dollar in cash reserves.

Originality/value

By linking two distinct research lines of audit quality and corporate cash reserves, this study adds to both lines of literature, as it is a novel one (to the best of the author’s knowledge) to provide evidence about the effect of audit quality on the auditees' liquidity policy (a real economic decision and internal financial policy) that ultimately boosts the auditees' investment efficiency. The author’s findings are consistent with influential monitoring and an insurance-like function of high quality audits in reducing information asymmetry and its consequences. His results also support the argument that auditees' transparency through high quality audits can be a pivotal determinant of their liquidity policy.