bluuebunny/crossref_metadata_2025 huggingface dataset:
A candidate gene cluster for the bioactive natural product gyrophoric acid in lichen-forming fungi
Authors: Singh Garima , Calchera Anjuli , Merges Dominik , Otte Jürgen , Schmitt Imke , Grande Francesco Dal
Published: January 2022

Natural products of lichen-forming fungi are structurally diverse and have a variety of medicinal properties. Despite this, they a have limited implementation in industry, because the corresponding genes remain unknown for most of the natural products. Here we implement a long-read sequencing and bioinformatic approach to identify the biosynthetic gene cluster of the bioactive natural product gyrophoric acid (GA). Using 15 high-quality genomes representing nine GA-producing species of the lichen-forming fungal genusUmbilicaria, we identify the most likely GA cluster and investigate cluster gene organization and composition across the nine species. Our results show that GA clusters are promiscuous withinUmbilicaria, with only three genes that are conserved across species, including the PKS gene. In addition, our results suggest that the same cluster codes for different but structurally similar NPs, i.e., GA, umbilicaric acid and hiascic acid, bringing new evidence that lichen metabolite diversity is also generated through regulatory mechanisms at the molecular level. Ours is the first study to identify the most likely GA cluster, and thus provides essential information to open new avenues for biotechnological approaches to producing and modifying GA and similar lichen-derived compounds. We show that bioinformatics approaches are useful in linking genes and potentially associated natural products. Genome analyses help unlocking the pharmaceutical potential of organisms such as lichens, which are biosynthetically diverse but slow growing, and difficult to cultivate due to their symbiotic nature.

Importance

The implementation of natural products in the pharmaceutical industry relies on the possibility of modifying the natural product (NP) pathway to optimize yields and pharmacological effects. Characterization of genes and pathways underlying natural product biosynthesis is a major bottleneck for the use of natural products in the pharmaceutical industry. Genome mining is a promising and relatively cost- and time-effective approach to exploit unexplored NP resources for drug discovery. In this study, we identify the most likely gene cluster for the lichen-forming fungal depside gyrophoric acid in nineUmbilicariaspecies. This compound shows cytotoxic and antiproliferative properties against several cancer cell lines, and is also a broad-spectrum antimicrobial agent. We identify the putative GA cluster from nineUmbilicariaspecies. This information paves the way for generating GA analogs with modified properties by selective activation/deactivation of genes.
Integrative in situ mapping of single-cell transcriptional states and tissue histopathology in an Alzheimer’s disease model
Authors: Zeng Hu , Huang Jiahao , Zhou Haowen , Meilandt William J. , Dejanovic Borislav , Zhou Yiming , Bohlen Christopher J. , Lee Seung-Hye , Ren Jingyi , Liu Albert , Sheng Hao , Liu Jia , Sheng Morgan , Wang Xiao
Published: January 2022

Amyloid-β plaques and neurofibrillary tau tangles are the neuropathologic hallmarks of Alzheimer’s disease (AD), but the spatiotemporal cellular responses and molecular mechanisms underlying AD pathophysiology remain poorly understood. Here we introduce STARmap PLUS to simultaneously map single-cell transcriptional states and disease marker proteins in brain tissues of AD mouse models at subcellular resolution (200 nm). This high-resolution spatial transcriptomics map revealed a core-shell structure where disease-associated microglia (DAM) closely contact amyloid-β plaques, whereas disease-associated astrocytes (DAA) and oligodendrocyte precursor cells (OPC) are enriched in the outer shells surrounding the plaque- DAM complex. Hyperphosphorylated tau emerged mainly in excitatory neurons in the CA1 region accompanied by the infiltration of oligodendrocyte subtypes into the axon bundles of hippocampal alveus. The integrative STARmap PLUS method bridges single-cell gene expression profiles with tissue histopathology at subcellular resolution, providing an unprecedented roadmap to pinpoint the molecular and cellular mechanisms of AD pathology and neurodegeneration.
NF-κB memory coordinates transcriptional responses to dynamic inflammatory stimuli
Authors: Wang Andrew G. , Son Minjun , Thom Nicholas , Tay Savaş
Published: January 2022

Many scenarios in cellular communication requires cells to interpret multiple dynamic signals. It is unclear how exposure to immune stimuli alters transcriptional responses to subsequent stimulus under inflammatory conditions. Using high-throughput microfluidic live cell analysis, we systematically profiled the NF-κB response to different signal sequences in single cells. We found that NF-κB dynamics stores the history of signals received by cells: depending on the dose and type of prior pathogenic and cytokine signal, the NF-κB response to subsequent stimuli varied widely, from no response to full activation. Using information theory, we revealed that these stimulus-dependent changes in the NF-κB response encode and reflect information about the identity and dose of the prior stimulus. Small-molecule inhibition, computational modeling, and gene expression profiling show that this encoding is driven by stimulus-dependent engagement of negative feedback modules. These results provide a model for how signal transduction networks process sequences of inflammatory stimuli to coordinate cellular responses in complex dynamic environments.
B cells discriminate HIV-1 Envelope protein affinities by sensing antigen binding association rates
Authors: Hossain Md. Alamgir , Anasti Kara , Watts Brian , Cronin Kenneth , Kane Advaiti Pai , Edwards R.J. , Easterhoff David , Zhang Jinsong , Rountree Wes , Ortiz Yaneth , Verkoczy Laurent , Reth Michael , Alam S. Munir
Published: January 2022

HIV-1 Envelope (Env) proteins designed to induce neutralizing antibody responses allow study of the role of affinities (equilibrium dissociation constant, KD) and kinetic rates (association/dissociation rates) on B cell antigen recognition. It is unclear whether affinity discrimination during B cell activation is based solely on Env protein binding KD, and whether B cells discriminate between proteins of similar affinities but that bind with different kinetic rates. Here we used a panel of Env proteins and Ramos B cell lines expressing IgM BCRs with specificity for CD4 binding-site broadly neutralizing (bnAb) or a precursor antibody to study the role of antigen binding kinetic rates on both early (proximal/distal signaling) and late events (BCR/antigen internalization) in B cell activation. Our results support a kinetic model for B cell activation in which Env protein affinity discrimination is based not on overall KD, but on sensing of association rate and a threshold antigen-BCR half-life.
Endothelial transmigration hotspots limit vascular leakage through heterogeneous expression of ICAM1
Authors: Grönloh Max L.B. , Arts Janine J.G. , Martínez Sebastián Palacios , van der Veen Amerens A. , Kempers Lanette , van Steen Abraham C.I. , Roelofs Joris J.T.H. , Nolte Martijn A. , Goedhart Joachim , van Buul Jaap D.
Published: January 2022

Upon inflammation, leukocytes leave the circulation by crossing the endothelial monolayer at specific transmigration ‘hotspot’ regions. Although these regions support leukocyte transmigration, their functionality is not clear. We found that endothelial hotspots function to limit vascular leakage during transmigration events. Using the photo-convertible probe mEos4b, we traced back and identified original endothelial transmigration hotspots. Using this method, we show that the heterogeneous distribution of ICAM-1 determines the location of the transmigration hotspot. Interestingly, loss of ICAM-1 heterogeneity either by CRISPR/Cas9-induced knockout of ICAM-1 or equalizing the distribution of ICAM-1 in all endothelial cells results in loss of TEM hotspots but not necessarily in reduced TEM events. Functionally, loss of endothelial hotspots results in increased vascular leakage during TEM. Mechanistically, we demonstrate that the 3 extracellular Ig-like domains of ICAM-1 are crucial for hotspot recognition. However, the intracellular tail of ICAM-1 and the 4thIg-like dimerization domain are not involved, indicating that intracellular signalling or ICAM-1 dimerization is not required for hotspot recognition. Together, we discovered that hotspots function to limit vascular leakage during inflammation-induced extravasation.
Chemical defense acquired via pharmacophagy can lead to herd protection in a sawfly
Authors: Singh Pragya , Grone Neil , Tewes Lisa Johanna , Müller Caroline
Published: January 2022

Predation is an important selection pressure acting on organisms, with organisms evolving diverse anti-predator strategies to combat it. One such widespread strategy is chemical defense in which organisms either synthesize or extrinsically acquire defensive chemicals. Little is known about the intraspecific transfer of such chemicals and if such chemicals acquired from conspecifics can also serve as defense against predation. Here, we used adults of the turnip sawfly, Athalia rosae, which can acquire neo-clerodane diterpenoids (‘clerodanoids’) via pharmacophagy after exposure to the plant, Ajuga reptans. We show that clerodanoid access mediates protection against predation by mantids for the sawflies, both in a no-choice feeding assay and a microcosm setup. Moreover, even indirect access to clerodanoids, via nibbling on conspecifics that had access to the plant, resulted in protection against predation albeit to a much lower degree than direct access. Furthermore, sawflies that had no direct access to clerodanoids were less consumed by mantids when they were grouped with conspecifics that had direct access. Most, but not all, of such initially undefended sawflies could acquire clerodanoids from conspecifics that had direct access to the plant, although in low quantities. Together our results demonstrate that clerodanoids serve as chemical defense that can be intraspecifically transferred. Moreover, the presence of chemically defended individuals in a group can confer protection onto conspecifics that had no direct access to clerodanoids, suggesting a ‘herd-protection’ effect.
Road encroachment mediates species occupancy, trait filtering and dissimilarity of songbird communities
Authors: Ascensão Fernando , D’Amico Marcello , Revilla Eloy , Pereira Henrique M.
Published: January 2022

Assessing the road effects on biodiversity is challenging because impacts may depend on both wildlife responses to roads and on the spatial arrangement of roads. We questioned whether an increase in road encroachment i.e., the advancement of roads into non-urban areas, leads to significant changes (positive and negative) in species occurrence, and if so, whether those changes are linked to specific traits related to perturbation sensitiveness, therefore acting as filters that increase the community compositional dissimilarity. Using a large citizen-science dataset of point-counts performed throughout Iberian Peninsula (4,459 unique survey sites), we modelled the effect of road density on the occurrence of common songbirds (n=58), while accounting for potential confounding effects of environment and survey effort. We then tested if species’ occurrence patterns would be linked to specific traits related to the ability to cope with human presence. Finally, we assessed how road density affect the community compositional dissimilarity. We estimated 24 (41%) and 12 (21%) species to be negatively and positively affected by roads, respectively. Increased road encroachment was positively related with a higher prevalence of urban dwellers and negatively related with the occurrence of species nesting and foraging on the ground. Furthermore, increasing road density translated into an increasing community compositional dissimilarity, mostly due to species turnover. Our study support previous research showing that roads have different effects on the occurrence of different species, but we revealed that at least three species' traits have a clear relation with such road responses. Such trait filtering is probably causing a high species turnover between songbird communities occurring in roaded and nearby roadless areas. Overall, we found that different species-specific responses to roads translates into changes at the community level. Landscape and road-network management should be conceived acknowledging that roads are contributing to biodiversity changes. As so, building upon the concepts of Land Sharing / Land Sparing, conservation actions should be tailored according to the different species responses e.g., road verge management targeting species having a positive relation with road density; and compensation actions targeting species showing a negative response toward roads.
Automatic grading of cervical biopsies by combining full and self-supervision
Authors: Lubrano Mélanie , Lazard Tristan , Balezo Guillaume , Bellahsen-Harrar Yaëlle , Badoual Cécile , Berlemont Sylvain , Walter Thomas
Published: January 2022

In computational pathology, predictive models from Whole Slide Images (WSI) mostly rely on Multiple Instance Learning (MIL), where the WSI are represented as a bag of tiles, each of which is encoded by a Neural Network (NN). Slide-level predictions are then achieved by building models on the agglomeration of these tile encodings. The tile encoding strategy thus plays a key role for such models. Current approaches include the use of encodings trained on unrelated data sources, full supervision or self-supervision. While self-supervised learning (SSL) exploits unlabeled data, it often requires large computational resources to train. On the other end of the spectrum, fully-supervised methods make use of valuable prior knowledge about the data but involve a costly amount of expert time. This paper proposes a framework to reconcile SSL and full supervision, showing that a combination of both provides efficient encodings, both in terms of performance and in terms of biological interpretability. On a recently organized challenge on grading Cervical Biopsies, we show that our mixed supervision scheme reaches high performance (weighted accuracy (WA): 0.945), outperforming both SSL (WA: 0.927) and transfer learning from ImageNet (WA: 0.877). We further shed light upon the internal representations that trigger classification results, providing a method to reveal relevant phenotypic patterns for grading cervical biopsies. We expect that the combination of full and self-supervision is an interesting strategy for many tasks in computational pathology and will be widely adopted by the field.
Evolution of assortative mating following selective introgression of pigmentation genes between two Drosophila species
Authors: David Jean R. , Ferreira Erina A. , Jabaud Laure , Ogereau David , Bastide Héloïse , Yassin Amir
Published: January 2022

Adaptive introgression is ubiquitous in animals but experimental support for its role in driving speciation remains scarce. In the absence of conscious selection, admixed laboratory strains of Drosophila asymmetrically and progressively lose alleles from one parental species and reproductive isolation against the predominant parent ceases after 10 generations. Here, we selectively introgressed during one year light pigmentation genes of D. santomea into the genome of its dark sibling D. yakuba, and vice versa. We found that the pace of phenotypic change differed between the species and the sexes, and identified through genome sequencing common as well as distinct introgressed loci in each species. Mating assays showed that assortative mating between introgressed flies and both parental species persisted even after four years (∼ 60 generations) from the end of the selection. Those results indicate that selective introgression of as low as 0.5% of the genome can beget morphologically-distinct and reproductively-isolated strains, two prerequisites for the delimitation of new species. Our findings hence represent a significant step towards understanding the genome-wide dynamics of speciation-through-introgression.
Partial sex linkage and linkage disequilibrium on the guppy sex chromosome
Authors: Qiu Suo , Yong Lengxob , Wilson Alastair , Croft Darren P. , Graham Chay , Charlesworth Deborah
Published: January 2022

The guppy Y chromosome has been considered a model system for the evolution of suppressed recombination between sex chromosomes, and it has been proposed that complete sex-linkage has evolved across about 3 Mb surrounding this fish’s sex-determining locus, followed by recombination suppression across a further 7 Mb of the 23 Mb XY pair, forming younger “evolutionary strata”. Sequences of the guppy genome show that Y is very similar to the X chromosome, making it important to understand which parts of the Y are completely non-recombining, and whether there is indeed a large completely non-recombining region. Here, we describe new evidence that supports a different interpretation of the data that suggested the presence of such a region. We analysed PoolSeq data in samples from multiple natural populations from Trinidad. This yields evidence for linkage disequilibrium (LD) between sequence variants and the sex-determining locus. Downstream populations have higher diversity than upstream ones (which display the expected signs of bottlenecks). The associations we observe conform to predictions for a genome region with infrequent recombination that carries one or more sexually antagonistic polymorphisms. They also suggest the region in which the sex-determining locus must be located. However, no consistently male-specific variants were found, supporting the suggestion that any completely sex-linked region may be very small.
Contrast-enhanced microCT evaluation of degeneration following partial and full width injuries to mouse lumbar intervertebral disc
Authors: Walk Remy E. , Moon Hong Joo , Tang Simon Y. , Gupta Munish C.
Published: January 2022

Study Design

Preclinical animal study

Objective

Evaluation of the degenerative progression resulting from either a partial- or full- width injury to the mouse lumbar intervertebral disc (IVD) using contrast-enhanced micro-computed tomography and histological analyses. We utilized a lateral-retroperitoneal surgical approach to access the lumbar IVD, and the injuries to the IVD were induced by either incising one side of the annulus fibrosus or puncturing both sides of the annulus fibrosus. The full-width injury caused dramatic reduction in nucleus pulposus hydration and significant degeneration. A partial-width injury produces localized deterioration around the annulus fibrosus that resulted in local tissue remodeling without gross degeneration to the IVD.

Methods

Female C57BL/6J mice of 3-4 months age were used in this study. They were divided into three groups to undergo partial-width, full-width, or sham injuries. The L5/L6 and L6/S1 lumbar IVDs were surgically exposed using a lateral-retroperitoneal approach. The L6/S1 IVDs were injured using either a surgical scalpel (partial-width) or a 33G needle (full-width), with the L5/L6 serving as an internal control. These animals were allowed to recover and then sacrificed at 2-, 4-, or 8-weeks post-surgery. The IVDs were assessed for degeneration using contrast-enhanced microCT (CEµCT) and histological analysis.

Results

The high-resolution 3D evaluation of the IVD confirmed that the respective injuries localized within one side of the annulus fibrosus or spanned the full width of the IVD. The full-width injury caused deteriorations in the nucleus pulposus after 2 weeks and progressed to significant degeneration at 8 weeks, while the partial width injury caused localized disruptions that remained limited to the annulus fibrosus.

Conclusion

The use of CEµCT revealed distinct IVD degeneration profiles resulting from partial- and full- width injuries. The partial width injury may serve as an alternative for IVD degeneration resulting from localized annulus fibrosus injuries in humans.
Experimental characterisation of de novo proteins and their unevolved random-sequence counterparts
Authors: Heames Brennen , Buchel Filip , Aubel Margaux , Tretyachenko Vyacheslav , Lange Andreas , Bornberg-Bauer Erich , Hlouchova Klara
Published: January 2022

De novo gene emergence provides a route for new proteins to be formed from previously non-coding DNA. Proteins born in this way are considered random sequences, and typically assumed to lack defined structure. While it remains unclear how likely a de novo protein is to assume a soluble and stable tertiary structure, intersecting evidence from random-sequence and de novo-designed proteins suggests that native-like biophysical properties are abundant in sequence space. Taking putative de novo proteins identified in human and fly, we experimentally characterise a library of these sequences to assess their solubility and structure propensity. We compare this library to a set of synthetic random proteins with no evolutionary history. Bioin-formatic prediction suggests that de novo proteins may have remarkably similar distributions of biophysical properties to unevolved random sequences of a given length and amino acid composition. However, upon expression in vitro, de novo proteins exhibit higher solubility which is further induced by the DnaK chaperone system. We suggest that while synthetic ran-dom sequences are a useful proxy for de novo proteins in terms of structure propensity, de novo proteins may be better integrated in the cellular system given their higher solubility.
Temporal control of late replication and coordination of origin firing by self-stabilizing Rif1-PP1 hubs in Drosophila
Authors: Cho Chun-Yi , Seller Charles A. , O’Farrell Patrick H.
Published: January 2022

In the metazoan S phase, coordinated firing of clusters of origins replicates different parts of the genome in a temporal program. Despite advances, neither the mechanism controlling timing nor that coordinating firing of multiple origins is fully understood. Rif1, an evolutionarily conserved inhibitor of DNA replication, recruits protein phosphatase 1 (PP1) and counteracts firing of origins by S-phase kinases. During the mid-blastula transition (MBT) in Drosophila embryos, Rif1 forms subnuclear hubs at each of the large blocks of satellite sequences and delays their replication. Each Rif1 hub disperses abruptly just prior to the replication of the associated satellite sequences. Here, we show that the level of activity of the S-phase kinase, DDK, accelerated this dispersal program, and that the level of Rif1-recruited PP1 retarded it. Further, Rif1-recruited PP1 supported chromatin association of nearby Rif1. This influence of nearby Rif1 can create a “community effect” counteracting kinase-induced dissociation such that an entire hub of Rif1 undergoes switch-like dispersal at characteristic times that shift in response to the balance of Rif1-PP1 and DDK activities. We propose a model in which the spatiotemporal program of late replication in the MBT embryo is controlled by self-stabilizing Rif1-PP1 hubs, whose abrupt dispersal synchronizes firing of associated late origins.

SIGNIFICANCE

Seventy years ago, it was discovered that large domains of the eukaryotic genome replicate at different times. Detailed descriptions left significant questions unresolved. How are the many origins in the large domains coordinated to fire in unison? What distinguishes different domains and gives rise to a temporal program? When Drosophila embryos first establish late replication, an inhibitor of DNA replication, Rif1, forms hubs over domains of late replicating DNA. Rif1 recruits protein phosphatase 1 (PP1), which prevents kinases from dispersing Rif1 hubs or activating associated origins. When kinase activity eventually exceeds a hub-specific threshold, the self-stabilization of Rif1-PP1 breaks down, hubs disperse abruptly, and all associated origins are free to initiate.
Somatodendritic release of cholecystokinin potentiates GABAergic synapses onto ventral tegmental area dopamine cells
Authors: Damonte Valentina Martinez , Pomrenze Matthew B. , Casper Caroline , Wolfden Annie M. , Malenka Robert C. , Kauer Julie A.
Published: January 2022

Neuropeptides are contained in nearly every neuron in the central nervous system and can be released from somatodendritic sites as well as from nerve terminals. Cholecystokinin (CCK), among the most abundant neuropeptides in the brain, is expressed in the majority of midbrain dopamine neurons. Here we report that ventral tegmental area (VTA) dopamine neurons release CCK from somatodendritic regions, where it triggers long-term potentiation of GABAergic synapses. The somatodendritic release occurs with trains of action potentials or prolonged but modest depolarization and is dependent on synaptotagmin 7 and T-type Ca2+ channels. Depolarization-induced LTP is blocked by the CCK2R antagonist, LY225910, and mimicked by exogenously added CCK. To test the behavioral role of CCK, we infused it into the mouse VTA. Ca2+ imaging in vivo demonstrated that infused CCK reduced dopamine cell signals during fasted food consumption. Moreover, local infusion of CCK also inhibited food consumption and decreased distance traveled in an open field test. Together our experiments introduce somatodendritic neuropeptide release as a previously unknown feedback regulator of VTA dopamine cell excitability and dopamine-related behaviors.
conST: an interpretable multi-modal contrastive learning framework for spatial transcriptomics
Authors: Zong Yongshuo , Yu Tingyang , Wang Xuesong , Wang Yixuan , Hu Zhihang , Li Yu
Published: January 2022

Motivation

Spatially resolved transcriptomics (SRT) shows its impressive power in yielding biological insights into neuroscience, disease study, and even plant biology. However, current methods do not sufficiently explore the expressiveness of the multi-modal SRT data, leaving a large room for improvement of performance. Moreover, the current deep learning based methods lack interpretability due to the “black box” nature, impeding its further applications in the areas that require explanation.

Results

We propose conST, a powerful and flexible SRT data analysis framework utilizing contrastive learning techniques. conST can learn low-dimensional embeddings by effectively integrating multi-modal SRT data, i.e. gene expression, spatial information, and morphology (if applicable). The learned embeddings can be then used for various downstream tasks, including clustering, trajectory and pseudotime inference, cell-to-cell interaction, etc. Extensive experiments in various datasets have been conducted to demonstrate the effectiveness and robustness of the proposed conST, achieving up to 10% improvement in clustering ARI in the commonly used benchmark dataset. We also show that the learned embedding can be used in complicated scenarios, such as predicting cancer progression by analyzing the tumour microenvironment and cell-to-cell interaction (CCI) of breast cancer. Our framework is interpretable in that it is able to find the correlated spots that support the clustering, which matches the CCI interaction pairs as well, providing more confidence to clinicians when making clinical decisions.
Structural Insights into Sphingosine-1-phosphate Receptor Activation
Authors: Yu Leiye , He Licong , Gan Bing , Ti Rujuan , Xiao Qingjie , Hu Hongli , Zhu Lizhe , Wang Sheng , Ren Ruobing
Published: January 2022

As a critical sphingolipid metabolite, sphingosine-1-phosphate (S1P) plays an essential role in immune and vascular systems. There are five S1P receptors, designated as S1PR1-5, encoded in the human genome, and their activities are governed by endogenous S1P, lipid-like S1P mimics, or non-lipid-like therapeutic molecules. Among S1PRs, S1PR1 stands out due to its non-redundant functions, such as the egress of T and B cells from the thymus and secondary lymphoid tissues, making it a potential therapeutic target. However, the structural basis of S1PR1 activation and regulation by various agonists remains unclear. Here we reported four atomic resolution cryo-EM structures of Gi-coupled human S1PR1 complexes: bound to endogenous agonist d18:1 S1P, benchmark lipid-like S1P mimic phosphorylated Fingolimod ((S)-FTY720-P), or non-lipid-like therapeutic molecule CBP-307 in two binding modes. Our results revealed the similarities and differences of activation of S1PR1 through distinct ligands binding to the amphiphilic orthosteric pocket. We also proposed a two-step “shallow to deep” transition process of CBP-307 for S1PR1 activation. Both binding modes of CBP-307 could activate S1PR1, but from shallow to deep transition may trigger the rotation of the N-terminal helix of Gαi and further stabilize the complex by increasing the Gαi interaction with the cell membrane. We combine with extensive biochemical analysis and molecular dynamic simulations to suggest key steps of S1P binding and receptor activation. The above results decipher the common feature of the S1PR1 agonist recognition and activation mechanism and will firmly promote the development of therapeutics targeting S1P receptors.
Sugar shock: Probing Streptococcus pyogenes metabolism through bioluminescence imaging
Authors: Davis Richard W. , Muse Charlotte G. , Eggleston Heather , Hill Micaila , Panizzi Peter
Published: January 2022

Streptococcus pyogenes (S. pyogenes) can thrive in its host during an infection, and, as a result, it must be able to respond to external stimuli and available carbon sources. The preclinical use of engineered pathogens capable of constitutive light production may provide realtime information on microbial-specific metabolic processes. Here we mapped the central metabolism of a luxABCDE-modified S. pyogenes Xen20 (Strep. Xen20) to its de novo synthesis of luciferase substrates as assessed by the rate of light production in response to different environmental triggers. Previous characterization predicted that the lux operon was under the myo-inositol iolE promotor. Here we show that supplementation with myoinositol generated increased Xen20 luminescence. Surprisingly, when supplemented with infection-relevant carbon sources, such as glucose or glycine, light production was diminished. This was presumably due to the scavenging of pyruvate by L-lactate dehydrogenase (LDH). Inhibition of LDH by its inhibitor, oxamate, partially restored luminescent signal in the presence of glucose, presumably by allowing the resulting pyruvate to proceed to acetyl-coenzyme A (CoA). This phenomenon appeared specific to the lactic acid bacterial metabolism as glucose or glycine did not reduce signal in an analogous luxABCDE-modified Gram-positive pathogen, Staph. Xen29. The Strep. Xen20 cells produced light in a concentration-dependent manner, inversely related to the amount of glucose present. Taken together, our measures of microbial response could provide new information regarding the responsiveness of S. pyogenes metabolism to acute changes in its local environments and cellular health.
The Cif proteins from Wolbachia prophage WO modify sperm genome integrity to establish cytoplasmic incompatibility
Authors: Kaur Rupinder , Leigh Brittany A. , Ritchie Isabella T. , Bordenstein Seth R.
Published: January 2022

Inherited microorganisms can selfishly manipulate host reproduction to drive through populations. In Drosophila melanogaster, germline expression of the native Wolbachia prophage WO proteins CifA and CifB cause cytoplasmic incompatibility (CI) in which embryos from infected males and uninfected females suffer catastrophic mitotic defects and lethality; however, in infected females, CifA expression rescues the embryonic lethality and thus imparts a fitness advantage to the maternally-transmitted Wolbachia. Despite widespread relevance to sex determination, evolution, and vector control, the mechanisms underlying when and how CI impairs male reproduction remain unknown and a topic of debate. Here we use cytochemical, microscopic, and transgenic assays in D. melanogaster to demonstrate that CifA and CifB proteins of wMel localize to nuclear DNA throughout the process of spermatogenesis. Cif proteins cause abnormal histone retention in elongating spermatids and protamine deficiency in mature sperms that travel to the female reproductive tract with Cif proteins. Notably, protamine gene knockouts enhance wild type CI. In ovaries, CifA localizes to germ cell nuclei and cytoplasm of early-stage egg chambers, however Cifs are absent in late-stage oocytes and subsequently in fertilized embryos. Finally, CI and rescue are contingent upon a newly annotated CifA bipartite nuclear localization sequence. Together, our results strongly support the Host Modification model of CI in which Cifs initially modify the paternal and maternal gametes to bestow CI-defining embryonic lethality and rescue.
Metal-binding propensity in the mitochondrial dynamin-related protein 1
Authors: Roy Krishnendu , Pucadyil Thomas J.
Published: January 2022

Dynamin-related protein1 (Drp1) functions to divide mitochondria and peroxisomes by binding specific adaptor proteins and lipids, both of which are integral to the limiting organellar membrane. In efforts to understand how such multivalent interactions regulate Drp1 functions,in vitroreconstitution schemes rely on recruiting soluble portions of the adaptors appended with genetically encoded polyhistidine tags onto membranes containing Ni2+-bound chelator lipids. These strategies are facile and circumvent the challenge in working with membrane proteins but assume that binding is specific to proteins carrying the polyhistidine tag. Here, we find using chelator lipids and chelator beads that both native and recombinant Drp1 directly bind Ni2+ions. Metal-binding therefore represents a potential strategy to deplete or purify Drp1 from native tissue lysates. Importantly, high concentrations of the metal in solution inhibit GTP hydrolysis and renders Drp1 inactive in membrane fission. Together, our results emphasize a metal-binding propensity, which could significantly impact Drp1 functions.
Cryo-EM reconstruction of the human 40S ribosomal subunit at 2.15 Å resolution
Authors: Pellegrino Simone , Dent Kyle C. , Spikes Tobias , Warren Alan J.
Published: January 2022

The chemical modification of ribosomal RNA and proteins is critical for ribosome assembly, for protein synthesis and may drive ribosome specialization in development and disease. However, the inability to accurately visualize these modifications has limited mechanistic understanding of the role of these modifications in ribosome function. Here we report the 2.15 Å resolution cryo-EM reconstruction of the human 40S ribosomal subunit. We directly visualize post-transcriptional modifications within the 18S rRNA and post-translational modifications at the N-termini of two ribosomal proteins. Additionally, we interpret the solvation shells in the core regions of the 40S ribosomal subunit and reveal how potassium and magnesium ions establish both universally conserved and eukaryote-specific coordination to promote the stabilization and folding of key ribosomal elements. This work provides unprecedented structural details for the human 40S ribosomal subunit that will serve as an important reference for unraveling the functional role of ribosomal RNA modifications.
Outer membrane protein A (OmpA) deficient Salmonella Typhimurium displays enhanced susceptibility towards β-lactam antibiotics: third-generation cephalosporins (ceftazidime) and carbapenems (meropenem)
Authors: Chowdhury Atish Roy , Mukherjee Debapriya , Singh Ashish Kumar , Chakravortty Dipshikha
Published: January 2022

The invasive non-typhoidal serovar of Salmonella enterica, namely Salmonella Typhimurium ST313, causes bloodstream infection in sub-Saharan Africa. Like other bacterial pathogens, the development of antimicrobial resistance is a severe problem in curing non-typhoidal Salmonella infection. In this work, we have investigated the role of four prominent outer membrane porins of S. Typhimurium, namely OmpA, OmpC, OmpD, and OmpF, in resistance against broad-spectrum β-lactam antibiotics-ceftazidime and meropenem. We found that deleting OmpA from Salmonella makes the bacteria susceptible to β-lactam drugs. The MIC for both the antibiotics reduced significantly for STM ΔompA compared to the wild-type and the ompA complemented strains. Despite the presence of antibiotics, the uninterrupted growth of STM ΔompC, ΔompD, and ΔompF endorsed the dispensability of these three porins in antimicrobial resistance. The β-lactam antibiotics caused massive depolarization in the outer membrane of the bacteria in the absence of OmpA. We have proved that none of the extracellular loops but the complete structure of perfectly folded OmpA is required by the bacteria for developing antimicrobial resistance. Our data revealed that STM ΔompA consumed more antibiotics than the wild-type and the complemented strain, resulting in severe damage of the bacterial outer membrane and subsequent killing of the pathogen by antibiotic-mediated oxidative stress. Upon deleting ompA, the steady decrease in the relative proportion of antibiotic-resistant persisters and the clearance of the STM ΔompA from the liver and spleen of C57BL/6 mice upon treatment with ceftazidime proved the role of OmpA in rendering protection against β-lactam antibiotics.
EFCAB4B (CRACR2A) genetic variants associated with COVID-19 fatality
Authors: Wang Dapeng , Wiktor Sabina D , Cheng Chew W , Simmons Katie J , Money Ashley , Pedicini Lucia , Carlton Asya , Breeze Alexander L , McKeown Lynn
Published: January 2022

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in more than 235 million cases worldwide and 4.8 million deaths (October 2021). Severe COVID-19 is characterised in part by vascular thrombosis and a cytokine storm due to increased plasma concentrations of factors secreted from endothelial and T-cells. Here, using patient data recorded in the UK Biobank, we demonstrate the importance of variations in Rab46 (CRACR2A) with clinical outcomes. Using logistic regression analysis, we determined that three single nucleotide polymorphisms (SNPs) in the gene EFCAB4B cause missense mutations in Rab46, which are associated with COVID-19 fatality independently of risk factors. All three SNPs cause changes in amino acid residues that are highly conserved across species, indicating their importance in protein structure and function. Two SNPs, rs17836273 (A98T) and rs36030417 (H212Q), cause amino acid substitutions in important functional domains: the EF-hand and coiled-coil domain respectively. By using molecular modelling, we suggest that the substitution of threonine at position 98 causes structural changes in the EF-hand calcium binding domain. Since Rab46 is a Rab GTPase that regulates both endothelial cell secretion and T-cell signalling, these missense variations may play a role in the molecular mechanisms underlying the thrombotic and inflammatory characteristics observed in patients with severe COVID-19 outcomes.
The language of profit warnings: a case of denial, defiance, desperation and defeat
Authors: Edgar Victoria C. , Brennan Niamh M. , Power Sean Bradley
Published: January 2022

Purpose

Taking a communication perspective, the paper explores management's rhetoric in profit warnings, whose sole purpose is to disclose unexpected bad news.

Design/methodology/approach

Adopting a close-reading approach to text analysis, the authors analyse three profit warnings of the now-collapsed Carillion, contrasting the rhetoric with contemporaneous investor conference calls to discuss the profit warnings and board minutes recording boardroom discussions of the case company's precarious financial circumstances. The analysis applies an Aristotelian framework, focussing on logos (appealing to logic and reason), ethos (appealing to authority) and pathos (appealing to emotion) to examine how Carillion's board and management used language to persuade shareholders concerning the company's adverse circumstances.

Findings

As non-routine communications, the language in profit warnings displays and mimics characteristics of routine communications by appealing primarily to logos (logic and reason). The rhetorical profiles of investor conference calls and board meeting minutes differ from profit warnings, suggesting a different version of the story behind the scenes. The authors frame the three profit warnings as representing three stages of communication as follows: denial, defiance and desperation and, for our case company, ultimately, culminating in defeat.

Research limitations/implications

The research is limited to the study of profit warnings in one case company.

Originality/value

The paper views profit warnings as a communication artefact and examines the rhetoric in these corporate documents to elucidate their key features. The paper provides novel insights into the role of profit warnings as a corporate communication vehicle/genre delivering bad news.
Designing interpretive communities toward justice: indexicality in classroom discourse
Authors: Storm Scott , Jones Karis , Beck Sarah W.
Published: January 2022

Purpose

This study aims to investigate how, through text-based classroom talk, youth collaboratively draw on and remix discourses and practices from multiple socially indexed traditions.

Design/methodology/approach

Drawing on data from a year-long social design experiment, this study uses qualitative coding and traces discoursal markers of indexicality.

Findings

The youth sustained, remixed and evaluated interpretive communities in their navigation across disciplinary and fandom discourses to construct a hybrid classroom interpretive community.

Originality/value

This research contributes to scholarship that supports using popular texts in classrooms as the focus of a scholarly inquiry by demonstrating how youth in one high school English classroom discursively index interpretive communities aligned with popular fandoms and literary scholarship. This study adds to understandings about the social nature of literary reading, interpretive whole-class text-based talk and literary literacies with multimodal texts in diverse, high school classrooms.
Why are we doing this? Teacher and student perspectives on literary reading
Authors: Hall Allison H. , Goldman Susan R.
Published: January 2022

Purpose

This paper aims to examine the extent to which students’ experiences and perceptions of their literature classroom align with their teacher’s instructional goals for literary inquiry and what teachers can learn from gaining access to students’ perspectives on their classroom experiences.

Design/methodology/approach

Thematic analyses were used to examine the data sources: mid-year and end-of-year interviews with six students, audio recordings of the teacher’s rationale for her instructional designs and a reflective discussion with the teacher upon reading the student interviews three years later.

Findings

Much of what the teacher intended students to get out of her instruction was what they expressed learning and experiencing in the class, yet some understood the purpose of the class to be far from her intentions. All the interviewed students had deeply personal and varied ways of relating what they learned in class to the world and their own lives. The teacher’s reflection on the interviews highlighted the importance of making space for multiple meanings and perspectives on literary works.

Originality/value

This paper speaks to the importance of surfacing students’ individual and varied ways of making sense of literary texts as part of instruction that values students’ thinking as well as the epistemic commitments of literary reading.
Women in hospitality and tourism: a study of the top-down and bottom-up dynamics
Authors: Ali Muhammad , Grabarski Mirit K. , Konrad Alison M.
Published: January 2022

Purpose

This study aims to investigate the impact of women’s representation at one hierarchical level on women’s representation above or below that level. No past research investigated these effects in the hospitality and tourism industries. The mixed results of research in other industries and across industries demand tests of curvilinearity and moderators.

Design/methodology/approach

Using annual equality reports, a panel data set for 2010–2019 was created for the hospitality and tourism industries. The sample of 581 organizations had up to 5,810 observations over the 10 years.

Findings

The analyses show the following effects of women’s representation: an inverted U-shape from management to non-management, a U-shape from non-management to management and a U-shape from management to the executive team, with more pronounced effect in small organizations.

Practical implications

To increase the number of female employees, organizations should invest their resources in hiring and retaining female managers until a gender balance is reached while managing any backlash from men. The results suggest that organizations with more than 40% of women non-management employees and 50% of women managers start `experiencing positive bottom-up dynamics. Thus, efforts need to be made to attract and retain a women’s pipeline at the non-management and management levels.

Originality/value

This study delivers pioneering evidence of the top-down and bottom-up phenomena in hospitality and tourism. It refines evidence of such effects found in past research conducted in other industries and across industries.