bluuebunny/crossref_metadata_2025 huggingface dataset:
Precision fMRI reveals that the language-selective network supports both phrase-structure building and lexical access during language production
Authors: Hu Jennifer , Small Hannah , Kean Hope , Takahashi Atsushi , Zekelman Leo , Kleinman Daniel , Ryan Elizabeth , Nieto-Castañón Alfonso , Ferreira Victor , Fedorenko Evelina
Published: September 2021

A fronto-temporal brain network has long been implicated in language comprehension. However, this network’s role in language production remains debated. In particular, it remains unclear whether all or only some language regions contribute to production, and which aspects of production these regions support. Across three fMRI experiments that rely on robust individual-subject analyses, we characterize the language network’s response to high-level production demands. We report three novel results. First, sentence production, spoken or typed, elicits a strong response throughout the language network. Second, the language network responds to both phrase-structure building and lexical access demands, although the response to phrase-structure building is stronger and more spatially extensive, present in every language region. Finally, contra some proposals, we find no evidence of brain regions—within or outside the language network—that selectively support phrase-structure building in production relative to comprehension. Instead, all language regions respond more strongly during production than comprehension, suggesting that production incurs a greater cost for the language network. Together, these results align with the idea that language comprehension and production draw on the same knowledge representations, which are stored in a distributed manner within the language-selective network and are used to both interpret and generate linguistic utterances.
Genetic variations within human gained enhancer elements affect human brain sulcal morphology
Authors: Lemaitre Herve , Le Guen Yann , Tilot Amanda K. , Stein Jason L. , Philippe Cathy , Mangin Jean-François , Fisher Simon E. , Frouin Vincent
Published: September 2021

The expansion of the cerebral cortex is one of the most distinctive changes in the evolution of the human brain. Cortical expansion and related increases in cortical folding may have contributed to emergence of our capacities for high-order cognitive abilities. Molecular analysis of humans, archaic hominins, and non-human primates has allowed identification of chromosomal regions showing evolutionary changes at different points of our phylogenetic history. In this study, we assessed the contributions of genomic annotations spanning 30 million years to human sulcal morphology measured via MRI in more than 18,000 participants from the UK Biobank. We found that variation within brain-expressed human gained enhancers, regulatory genetic elements that emerged since our last common ancestor with Old World monkeys, explained more trait heritability than expected for the left and right calloso-marginal posterior fissures and the right central sulcus. Intriguingly, these are sulci that have been previously linked to the evolution of locomotion in primates and later on bipedalism in our hominin ancestors.
Mechanical Causation of Biological Structure: Productive Pulls Produce Persistent Filaments in a Human Fibroblast Model of Matrix Development
Authors: Silverman Alexandra A. , Olszewski Jason D. , Siadat Seyed Mohammad , Ruberti Jeffrey W.
Published: September 2021

The principal mechanisms driving the synthesis and organization of durable animal structure have been the subject of intense investigation for decades. Here, we present evidence that mechanical strains can direct the formation of extracellular matrix (ECM) filaments via a mechanochemical cascade. This process is driven by cooperative cell contractions (“pulls”) that organize and precipitate structure via strain-induced polymer assembly. In anin vitromodel of ECM synthesis, we use high-resolution optical microscopy to observe the kinematics of cell motion during their growth to confluency and identified cell-to-cell pulls that result in the production of persistent ECM filaments. Using live-cell confocal imaging, we confirmed that these pulls can directly cause the formation of fibronectin filaments that then bind collagen, producing persistent structures aligned with the direction of the pull. The finding suggests a new model for initial durable structure formation in animals based on local cell contraction, extensional strain and polymer mechanochemistry. The results have important implications for ECM development, growth and life-threatening pathologies of the ECM such as fibrosis.
N-myc Downstream Regulated Gene 1 (NDRG1) functions as a molecular switch for cellular adaptation to hypoxia
Authors: Park Jong S. , Gabel Austin M. , Kassir Polina , Kang Lois , Chowdhary Prableen K. , Osei-Ntansah Afia , Tran Neil D. , Viswanathan Soujanya , Canales Bryanna , Ding Pengfei , Lee Young-Sam , Brewster Rachel
Published: September 2021

Lack of oxygen (hypoxia and anoxia) is detrimental to cell function and survival and underlies many disease conditions. Hence, metazoans have evolved mechanisms to adapt to low oxygen. One such mechanism, metabolic suppression, decreases the cellular demand for oxygen by downregulating ATP-demanding processes. However, the molecular mechanisms underlying this adaptation are poorly understood. Here, we report on the role of ndrg1a in hypoxia adaptation of the anoxia-tolerant zebrafish embryo. ndrg1a is expressed in the kidney and ionocytes, cell types that use large amounts of ATP to maintain ion homeostasis. ndrg1a mutants are viable and develop normally when raised under normal oxygen. However, their survival and kidney function is reduced relative to WT embryos following exposure to prolonged anoxia. We further demonstrate that Ndrg1a binds to the energy-demanding sodium-potassium ATPase (NKA) pump under anoxia and is required for its degradation, which may preserve ATP in the kidney and ionocytes and contribute to energy homeostasis. Lastly, we show that sodium azide treatment, which increased lactate levels, was sufficient to trigger NKA degradation in an Ndrg1a-dependent manner. These findings support a model whereby Ndrg1a is essential for hypoxia adaptation and functions downstream of lactate signaling to induce NKA degradation, a process known to conserve cellular energy.
Cytosolic aspartate aminotransferase moonlights as a ribosome binding modulator of Gcn2 activity during oxidative stress
Authors: Crawford Robert A. , Ashe Mark P. , Hubbard Simon J. , Pavitt Graham D.
Published: September 2021

Regulation of translation is a fundamental facet of the cellular response to rapidly changing external conditions. Specific RNA-binding proteins (RBPs) co-ordinate the translational regulation of distinct mRNA cohorts during stress. To identify RBPs with previously under-appreciated roles in translational control, we used polysome profiling and mass spectrometry to identify and quantify proteins associated with translating ribosomes in unstressed yeast cells and during oxidative stress and amino acid starvation, which both induce the integrated stress response (ISR). Over 800 proteins were identified across polysome gradient fractions, including ribosomal proteins, translation factors and many others without previously described translation-related roles, including numerous metabolic enzymes. We identified variations in patterns of polysome enrichment in both unstressed and stressed cells and identified proteins enriched in heavy polysomes during stress. Genetic screening of polysome-enriched RBPs identified the cytosolic aspartate aminotransferase, Aat2, as a ribosome-associated protein whose deletion conferred growth sensitivity to oxidative stress. Loss of Aat2 caused aberrantly high activation of the ISR via enhanced eIF2α phosphorylation andGCN4activation. Importantly, non-catalyticAAT2mutants retained polysome association and did not show heightened stress sensitivity. Aat2 therefore has a separate ribosome-associated translational regulatory or ‘moonlighting’ function that modulates the ISR independent of its aspartate aminotransferase activity.
A transcriptome atlas of the mouse iris at single cell resolution defines cell types and the genomic response to pupil dilation
Authors: Wang Jie , Rattner Amir , Nathans Jeremy
Published: September 2021

The iris controls the level of retinal illumination by controlling pupil diameter. It is a site of diverse ophthalmological diseases and it is a potential source of cells for ocular auto-transplantation. The present study provides foundational data on the mouse iris based on single nucleus RNA sequencing. More specifically, this work has (1) defined all of the major cell types in the mouse iris and ciliary body, (2) led to the discovery of two types of iris stromal cells and two types of iris sphincter cells, (3) revealed the differences in cell type-specific transcriptomes in the resting, constricted, and dilated states, and (4) identified and validated antibody and in situ hybridization probes that can be used to visualize the major iris cell types. By immunostaining for specific iris cell-types, we have observed and quantified distortions in nuclear morphology associated with iris dilation and clarified the neural crest contribution to the iris by showing thatWnt1-Cre-expressing progenitors contribute to nearly all iris cell types, whereasSox10-Cre-expressing progenitors contribute only to stromal cells. This work should be useful as a point of reference for investigations of iris development, disease, and pharmacology, for the isolation and propagation of defined iris cell types, and for iris cell engineering and transplantation.
Whole-genome sequencing analysis of wild-caught house mice Mus musculus from Madagascar
Authors: Fujiwara Kazumichi , Ranorosoa Marie C , Ohdachi Satoshi D , Arai Satoru , Sakuma Yuki , Suzuki Hitoshi , Osada Naoki
Published: September 2021

In Madagascar, the house mouse (Mus musculus) is thought to have colonized with human activities and is now one of the most abundant rodents on the island. In this study, we determined the whole-genome sequences of five Madagascar house mice captured from the wild. We examined the evolutionary history of the population by analyzing the mitochondrial and autosomal genomes. We confirmed that the mitochondrial genome lineages of Madagascar house mice formed a monophyletic clade placed at one of the most basal positions in the species. An analysis of autosomal genomic sequences indicates that the Madagascar house mice are genetically members of M. m. castaneus (CAS), but also contain genetic elements of M. m. domesticus (DOM) resulting from hybridization between subspecies. The signature of a strong population bottleneck 1000-3000 years ago was observed in both mitochondrial and autosomal genomic data. All samples showed strong genetic affinity to many CAS samples across a wide range of Indian Ocean coastal and island regions, with divergence time estimated around 4000 years ago. These findings support that the Madagascar house mice started to colonize the island with human agricultural activity, and experienced complex history for the establishment.
Multi-omic Characterization of Pancreatic Cancer-Associated Macrophage Polarization Reveals Deregulated Metabolic Programs Driven by the GMCSF-PI3K Pathway
Authors: Boyer Seth M. , Lee Ho-Joon , Steele Nina G. , Zhang Li , Sajjakulnukit Peter , Andren Anthony , Ward Matthew H. , Basrur Venkatesha , Zhang Yaqing , Nesvizhskii Alexey I. , di Magliano Marina Pasca , Halbrook Christopher J. , Lyssiotis Costas A.
Published: September 2021

The pancreatic ductal adenocarcinoma (PDA) microenvironment is composed of a variety of cell types and marked by extensive fibrosis and inflammation. Tumor-associated macrophages (TAM) are abundant, and they are important mediators of disease progression and invasion. TAMs are polarized in situ to a tumor promoting and immunosuppressive phenotype via cytokine signaling and metabolic crosstalk from malignant epithelial cells and other components of the tumor microenvironment (TME). However, the specific distinguishing features and functions of TAMs remain poorly defined. Here, we generated tumor-educated macrophages (TEM)in vitroand performed detailed, multi-omic characterization (i.e. transcriptomics, proteomics, metabolomics). Our results reveal unique genetic and metabolic signatures of TEMs, the veracity of which were queried against our in-house single cell RNA sequencing (scRNA-seq) dataset of human pancreatic tumors. This analysis identified expression of novel, metabolic TEM markers in human pancreatic TAMs, including ARG1, ACLY, and TXNIP. We then utilized our TEM model system to study the role of mutant Kras signaling in cancer cells on TEM polarization. This revealed an important role for GM-CSF and lactate on TEM polarization, molecules released from cancer cells in a mutant Kras-dependent manner. Lastly, we demonstrate that GM-CSF dysregulates TEM gene expression and metabolism through PI3K-AKT pathway signaling. Collectively, our results define new markers and programs to classify pancreatic TAMs, how these are engaged by cancer cells, and the precise signaling pathways mediating polarization.
AgRP neurons coordinate the mitigation of activity-based anorexia
Authors: Sutton Ames K. , Duane Sean C. , Shamma Ahmed M. , Skillings Anna , Krashes Michael J.
Published: September 2021

Anorexia nervosa (AN) is a debilitating and deadly disease characterized by low body mass index due to diminished food intake, and oftentimes concurrent hyperactivity. A high percentage of AN behavioral and metabolic phenotypes can be replicated in rodents given access to a voluntary running wheel and subject to food restriction, termed activity-based anorexia (ABA). Despite the well-documented body weight loss observed in AN human patients and ABA rodents, much less is understood regarding the neurobiological underpinnings of these maladaptive behaviors. Moreover, while exercise has been shown to diminish the activity of hunger-promoting hypothalamic agouti-related peptide (AgRP) neurons, much less is known regarding their activity and function in the mediation of food intake during ABA. Here, feeding microstructure analysis revealed ABA mice decreased food intake due to increased interpellet interval retrieval and diminished meal number. Longitudinal activity recordings of AgRP neurons in ABA animals revealed a maladaptive inhibitory response to food. We then demonstrated that ABA development or progression can be mitigated by chemogenetic AgRP activation through the reprioritization of food intake (increased meal number) over hyperactivity. These results elucidate a potential neural target for the amelioration of behavioral maladaptations present in AN patients.
Combining CRISPR/Cas9 and brain imaging: from genes to molecules to networks
Authors: Marciano Sabina , Ionescu Tudor Mihai , Saw Ran Sing , Cheong Rachel Y. , Kirik Deniz , Maurer Andreas , Pichler Bernd , Herfert Kristina
Published: September 2021

Receptors, transporters and ion channels are important targets for therapy development in neurological diseases including Alzheimeŕs disease, Parkinsońs disease, epilepsy, schizophrenia and major depression. Several receptors and ion channels identified by next generation sequencing may be involved in disease initiation and progression but their mechanistic role in pathogenesis is often poorly understood. Gene editing andin vivoimaging approaches will help to identify the molecular and functional role of these targets and the consequence of their regional dysfunction on whole brain level. Here, we combine CRISPR/Cas9 gene-editing within vivopositron emission tomography (PET) and functional magnetic resonance imaging (fMRI) to investigate the direct link between genes, molecules, and the brain connectome. The extensive knowledge of theSlc18a2gene encoding the vesicular monoamine transporter (VMAT2), involved in the storage and release of dopamine, makes it an excellent target for studying the gene networks relationships while structurally preserving neuronal integrity and function. We edited theSlc18a2in the substantia nigra pars compacta of adult rats and usedin vivomolecular imaging besides behavioral, histological, and biochemical assessments to characterize the CRISPR/Cas9-mediated VMAT2 knockdown. Simultaneous PET/fMRI was performed to investigate molecular and functional brain alterations. We found that stage-specific adaptations of brain functional connectivity follow the selective impairment of presynaptic dopamine storage and release. Our study reveals that recruiting different brain networks is an early response to the dopaminergic dysfunction preceding neuronal cell loss. Our combinatorial approach is a novel tool to investigate the impact of specific genes on brain molecular and functional dynamics which will help to develop tailored therapies for normalizing brain function. The method can easily be transferred to higher-order species allowing for a direct comparison of the molecular imaging findings.
Bacterial supergroup specific “Cost” of Wolbachia infections in Nasonia vitripennis
Authors: Tiwary Alok , Babu Rahul , Sen Ruchira , Raychoudhury Rhitoban
Published: September 2021

The maternally-inherited endosymbiont, Wolbachia, is known to alter the reproductive biology of its arthropod hosts for its benefit and can induce both positive and negative fitness effects in many hosts. Here we describe the effects of the maintenance of two distinct Wolbachia infections, one each from supergroups A and B, on the parasitoid host Nasonia vitripennis. We compare the effect of Wolbachia infections on various traits between the uninfected, single A infected, single B infected, and the double infected strains with their cured versions. Contrary to the previous reports, our results suggest that there is a significant cost associated with the maintenance of Wolbachia infections where traits like family size, fecundity, longevity, and rates of male copulation are compromised in Wolbachia infected strains. The double infected and supergroup B infection strains show higher Wolbachia titer than supergroup A. The double infected Wolbachia strain has the most detrimental impact on the host as compared to single infections. Moreover, there is a supergroup-specific negative impact on these wasps as the supergroup B infections elicit the most pronounced negative effects. These findings raise important questions on the mechanism of survival and maintenance of these reproductive parasites in arthropod hosts.
Mechanistic insight into light-dependent recognition of Timeless by Drosophila cryptochrome
Authors: Lin Changfan , Schneps Connor M. , Chandrasekaran Siddarth , Ganguly Abir , Crane Brian R.
Published: September 2021

Cryptochrome (CRY) entrains the fly circadian clock by binding to Timeless (TIM) in light and triggering its degradation. Undocking of a helical C-terminal tail (CTT) in response to photoreduction of the CRY flavin cofactor gates TIM binding. A generally-applicable Select Western-blot-Free Tagged-protein Interaction (SWFTI) assay enables quantification of CRY binding to TIM in dark and light. The assay is utilized to study CRY variants with residue substitutions in the flavin pocket and correlate their TIM affinities with CTT undocking, as measured by pulse-dipolar ESR spectroscopy and evaluated by molecular dynamics simulations. CRY variants with the CTT removed or undocked bind TIM constitutively, whereas those incapable of photoreduction bind TIM weakly. In response to flavin redox state, two conserved histidine residues contribute to a robust on/off switch by mediating CTT interactions with the flavin pocket and TIM. Our approach provides an expeditious means to quantify protein-protein interactions and photoreceptor targeting.
The influence of age and sex on the pre-treatment immune microenvironment of a carcinogen induced murine model of bladder cancer
Authors: Hamade Ali , Li Deyang , Tyryshkin Kathrin , Xu Minqi , Chenard Stephen , Conseil Gwenaelle , Yolmo Priyanka , Siemens D. Robert , Koti Madhuri
Published: September 2021

The incidence of urothelial carcinoma of the bladder is four times higher in males than females; however, females tend to present with a more aggressive disease, a poorer response to immunotherapy and suffer worse clinical outcomes. Recent findings have demonstrated sexual dimorphism in the tumor immune microenvironment of non-muscle invasive bladder cancer and associated clinical outcomes. However, a significant gap in knowledge remains with respect to the current pre-clinical modeling approaches and more precisely recapitulating these differences towards improved therapeutic design. Based on the similarities in mucosal immune physiology between humans and mice, we evaluated the sex and age-related immune alterations in healthy murine bladders. Bulk-RNA sequencing and multiplex immunofluorescence based spatial immune profiling of healthy murine bladders from male and female mice of age groups spanning young to old showed a highly altered immune landscape that exhibited sex and age associated differences, particularly in the context of B cell associated responses. Spatial profiling using markers specific to macrophages, T lymphocytes, B lymphocytes, activated dendritic cells, high endothelial venules, myeloid cells and the PD-L1 immune checkpoint showed sex and age associated differences. Bladders from healthy older female mice showed a higher number of tertiary lymphoid structures (TLSs) compared to both young female and male equivalents. Spatial immune profiling of N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) carcinogen exposed male and female bladders from young and old mice revealed a similar frequency of TLS formation, sex differences in the bladder immune microenvironment and, age differences in latency of tumor induction. These findings support the incorporation of sex and age as factors in pre-clinical modeling of NMIBC and will potentially advance the field of immunotherapeutic drug development to improve clinical outcomes in NMIBC.
HOIL-1-catalysed ubiquitylation of unbranched glucosaccharides and its activation by ubiquitin oligomers
Authors: Kelsall Ian R. , McCrory Elisha H. , Xu Yingqi , Scudamore Cheryl L. , Nanda Sambit K. , Mancebo-Gamella Paula , Wood Nicola T. , Knebel Axel , Matthews Stephen J. , Cohen Philip
Published: September 2021

HOIL-1, a component of the Linear Ubiquitin Assembly Complex (LUBAC), ubiquitylates serine and threonine residues in proteins, forming ester bonds (Kelsall et al, 2019, PNAS 116, 13293-13298). Here we report that mice expressing the E3 ligase-inactive HOIL-1[C458S] mutant accumulate polyglucosan in brain, cardiac muscle and other organs, indicating that HOIL-1’s E3 ligase activity is essential to prevent these toxic polysaccharide deposits from accumulating. We found that HOIL-1 monoubiquitylates glycogen and α1:4-linked maltoheptaose in vitro and identify the C6 hydroxyl moiety of glucose as the site of ester-linked ubiquitylation. The HOIL-1-catalysed monoubiquitylation of maltoheptaose was accelerated >100-fold by Met1-linked or Lys63-linked ubiquitin oligomers, which interact with the catalytic RBR domain of HOIL-1. HOIL-1 also transferred preformed ubiquitin oligomers to maltoheptaose en bloc, producing polyubiquitylated maltoheptaose in one catalytic step. The Sharpin and HOIP components of LUBAC, but not HOIL-1, bound to amylose resin in vitro, suggesting a potential function in targeting HOIL-1 to unbranched glucosaccharides in cells. We suggest that monoubiquitylation of unbranched glucosaccharides may initiate their removal by glycophagy to prevent precipitation as polyglucosan.
Structural and molecular rationale for the diversification of resistance mediated by the Antibiotic_NAT family
Authors: Stogios Peter J. , Bordeleau Emily , Xu Zhiyu , Skarina Tatiana , Evdokimova Elena , Chou Sommer , Diorio-Toth Luke , D’Souza Alaric W. , Patel Sanket , Dantas Gautam , Wright Gerard D. , Savchenko Alexei
Published: September 2021

The environmental microbiome harbors a vast repertoire of antibiotic resistance genes (ARGs) which can serve as evolutionary predecessors for ARGs found in pathogenic bacteria, or can be directly mobilized to pathogens in the presence of selection pressures. Thus, ARGs from benign environmental bacteria are an important resource for understanding clinically relevant resistance. Here, we conduct a comprehensive functional analysis of the Antibiotic_NAT family of aminoglycoside acetyltransferases. We determined a pan-family antibiogram of 21 Antibiotic_NAT enzymes, including 8 derived from clinical isolates and 13 from environmental metagenomic samples. We find that environment-derived representatives confer high-level, broad-spectrum resistance, including against the atypical aminoglycoside apramycin, and that a metagenome-derived gene likely is ancestral to an AAC(3) gene found in clinical isolates. Through crystallographic analysis, we rationalize the molecular basis for diversification of substrate specificity across the family. This work provides critical data on the molecular mechanism underpinning resistance to established and emergent aminoglycoside antibiotics and broadens our understanding of ARGs in the environment.
Elongated sample plot shapes produce higher measurement precision in clustering populations
Authors: McGarvey Richard , Burch Paul , Matthews Janet M.
Published: September 2021

Monitoring the density of natural populations is crucial for ecosystem management decision making and natural resource management. The most widely used method to measure the population density of animal and plant species in natural habitats is to count organisms in sample plots. Yet evaluation of survey performance by different sample plot shapes, e.g. quadrats compared with transects, has been largely neglected since the 1990s and has not been undertaken using simulation. Simulating populations and surveys, we evaluated population density measurement precision for 900 cases, testing 30 sample plot survey designs in each of 30 spatially clustered populations. We varied three design options: elongation of plot shape while keeping sample area constant, systematic or random plot allocation, and sample size. Survey design performance varied markedly: elongating the plot shape always improved survey precision; allocating plots systematically sometimes did. (i) Averaged across all tested populations, elongated (1:100) transect plot shapes were 2-to-3 times more precise than square (10:10) quadrats. (ii) The precision of systematic surveys accelerated with sample plot number, increasing faster than the (known) linear increase under simple random sampling. This non-linear, concave upward, dependence of systematic precision on sample size has not previously been reported. (iii) The most precise design we evaluated used long narrow transects allocated systematically. Averaging among all 30 tested populations, a researcher would need 600 random square (10:10) quadrats to equal the precision achieved by 100 systematic (1:100) transects. Finding this average efficiency difference of 600% for a survey sample size of 100 plots, these simulation results imply that field trips requiring five sampling days using random quadrats could achieve equal precision in one or two days using systematic elongated transects. For all clustered populations we tested, long narrow transects resulted in a more efficient design for sample plot survey.
Population differentiation of polygenic score predictions under stabilizing selection
Authors: Yair Sivan , Coop Graham
Published: September 2021

1

Given the many small-effect loci uncovered by genome-wide association studies (GWAS), polygenic scores have become central to the drive for genomic medicine and have spread into various areas including evolutionary studies of adaptation. While promising, these scores are fraught with issues of portability across populations, due to mis-estimated effect sizes and missing causal loci across populations unrepresented in large-scale GWAS. The poor portability of polygenic scores at first seems at odds with the view that much of common genetic variation is shared among populations. Here we investigate one potential cause of this discrepancy, stabilizing selection on complex traits. Somewhat counter-intuitively, while stabilizing selection to the same optimum phenotype leads to lower phenotypic differentiation among populations, it increases genetic differentiation at GWAS loci because it accelerates the turnover of polymorphisms underlying trait variation within populations. We develop theory to show how stabilizing selection impacts the utility of polygenic scores when applied to unrepresented populations. Specifically, we quantify their reduced prediction accuracy and find they can substantially overstate average genetic differences of phenotypes among populations. Our work emphasizes stabilizing selection to the same optimum as a useful null evolutionary model to draw connections between patterns of allele frequency and polygenic score differentiation.
Transmembrane region dimer structures of Type 1 receptors readily sample alternate configurations: MD simulations using the Martini 3 coarse grained model compared to AlphaFold2 Multimer
Authors: Sahoo Amita R. , Souza Paulo C. T. , Meng Zhiyuan , Buck Matthias
Published: September 2021

Determination of the structure and dynamics of transmembrane (TM) regions of single-transmembrane receptors is key to understanding their mechanism of signal transduction across the plasma membrane. Although many studies have been performed on isolated soluble extra- and intracellular receptor domains in aqueous solutions, limited knowledge exists on the lipid embedded TM domain. In this study, we examine the assembly of configurations of receptor TM region dimers using the Martini 3 force field for coarse-grain (CG) molecular dynamics simulations. This recently published version of Martini has new bead types and sizes, which allows more accurate predictions of molecular interactions compared to the previous versions. At first glance our results with Martini 3 simulations show only a reasonable agreement withab initiopredictions using PREDDIMER (for TM domains only), AlphaFold2 Multimer and with available NMR derived structures for TM helix dimers. Specifically, 6 of 11 CG TM structures are similar to the NMR structures (within < 3.5 Å mainchain RMSD) compared to 10 of 11 and 9 of 11 using PREDDIMER and Alphafold, respectively (7 structures of the latter are within 1.5 Å) Surprisingly, AlphaFold2 predictions are more comparable with NMR structures when the database of 2001 (mainly composed of soluble proteins) instead of 2020 PDB structures are used. While there are some differences in the conditions used, the CG simulations primarily reveal that alternate configurations of the TM dimers that are sampled, which readily interconvert with a predominant population. The implications of these findings for our understanding of the signalling mechanism of TM receptors are discussed, including opportunities for the development of new pharmaceuticals, some of which are peptide based.
Prediction and validation of the three-dimensional structure of glucokinase-1 from Phytophthora infestans
Authors: Villalobos-Piña Liara , Rojas Ascanio , Acosta Héctor
Published: September 2021

According to its primary structure, the [PITG_06016] gene encodes for one of the 7 glucokinases present in Phytophthora infestans (PiGlcK-1), the causal agent of late blight disease. Currently, there are no structural studies of any of its enzymes, being the determination of the three-dimensional (3D) structure of PiGlcK-1 a necessary contribution in the deduction of its functions, its interaction with ligands, and possible regulatory mechanisms. In this work we present the first structural model obtained by in silico tools for PiGlcK-1. For the prediction of this model, different algorithms were used to find the best annealing, refinement, and qualitative evaluation of them. A structural comparison of the predicted model with other structures of crystallized kinase enzymes allowed us to identify the regions of interaction with their classical substrates (glucose and ATP), as well as to identify the amino acid residues involved in the binding of other substrates such as fructose and ADP. In addition, we propose a possible recognition region of PPi, an activator of kinase activity that includes the GXGE motif, conserved in enzymes of the ribokinase (RK) family, which distinguishes this PiGlcK-1 from a classical glucokinase. Accordingly, these findings suggest PPi-binding motif as potential targets for the development of inhibitors of PiGlcK-1 activity.
AlphaFold Protein Structure Database for Sequence-Independent Molecular Replacement
Authors: Chai Lawrence , Zhu Ping , Chai Jin , Pang Changxu , Andi Babak , McSweeney Sean , Shanklin John , Liu Qun
Published: September 2021

Crystallographic phasing recovers the phase information that is lost during a diffraction experiment. Molecular replacement is a dominant phasing method for the crystal structures in the protein data bank. In one form it uses a protein sequence to search a structure database for finding suitable templates for phasing. However, such sequence information is not always available such as when proteins are crystallized with unknown binding partner proteins or when the crystal is that of a contaminant. The recent development of AlphaFold has resulted in the availability of predicted protein structures for all proteins from twenty species. In this work, we tested whether AlphaFold-predicted E. coli protein structures were accurate enough for sequence-independent phasing of diffraction data from two crystallization contaminants for which we had not identified the protein. Using each of more than 4000 predicted structures as a search model, robust molecular replacement solutions were obtained which allowed the identification and structure determination of both structures, YncE and YadF. Our results advocate a general utility of AlphaFold-predicted structure database with respect to crystallographic phasing.
Engineered bacteria detect tumor DNA
Authors: Cooper Robert M. , Wright Josephine A. , Ng Jia Q. , Goyne Jarrad M. , Suzuki Nobumi , Lee Young K. , Ichinose Mari , Radford Georgette , Ryan Feargal , Kumar Shalni , Thomas Elaine M. , Vrbanac Laura , Knight Rob , Woods Susan L. , Worthley Daniel L. , Hasty. Jeff
Published: September 2021

Advances in bacterial engineering have catalysed the development of living cell diagnostics and therapeutics1–3, including microbes that respond to gut inflammation4, intestinal bleeding5, pathogens6 and hypoxic tumors7. Bacteria can access the entire gastrointestinal tract8 to produce outputs measured in stool4 or urine7. Cellular memory, such as bistable switches4,9,10 or genomic rearrangements11, allows bacteria to store information over time. However, living biosensors have not yet been engineered to detect specific DNA sequences or mutations from outside the cell. Here, we engineer naturally competent Acinetobacter baylyi to detect donor DNA from the genomes of colorectal cancer (CRC) cells, organoids and tumors. We characterize the functionality of the biosensors in vitro with co-culture assays and then validate in vivo with sensor bacteria delivered to mice harboring colorectal tumors. We observe horizontal gene transfer from the tumor to the sensor bacteria in our mouse model of CRC. The sensor bacteria achieved 100% discrimination between mice with and without CRC. This Cellular Assay of Targeted, CRISPR-discriminated Horizontal gene transfer (CATCH), establishes a framework for biosensing of mutations or organisms within environments that are difficult to sample, among many other potential applications. Furthermore, the platform could be readily expanded to include production and delivery of antibiotic or antineoplastic therapeutic payloads at the detection site.
A case for glycerol as an acceptable additive for single particle cryoEM samples
Authors: Basanta Benjamin , Hirschi Marscha M. , Grotjahn Danielle A. , Lander Gabriel C.
Published: September 2021

Buffer composition and sample preparation guidelines for cryo-electron microscopy are geared toward maximizing imaging contrast and reducing electron beam-induced motion. These pursuits often involve the minimization or complete removal of additives that are commonly used to facilitate proper protein folding and minimize aggregation. Among these admonished additives is glycerol, a widely used osmolyte that aids protein stability. In this work, we show that inclusion of glycerol does not preclude high-resolution structure determination by cryoEM, as demonstrated by a ∼2.3 Å reconstruction of mouse apoferritin (∼500 kDa) and a ∼3.3 Å reconstruction of rabbit muscle aldolase (∼160 kDa) in presence of 20% v/v glycerol. While we found that generating thin ice that is amenable for high-resolution imaging requires long blot times, the addition of glycerol did not result in increased beam-induced motion nor an inability to pick particles. Overall, our findings indicate glycerol should not be discounted as a cryoEM sample buffer additive, particularly for large, fragile complexes that are prone to disassembly or aggregation upon its removal.
The histone deacetylase activity of HDAC1/2 is required to safeguard zygotic genome activation in mice and cattle
Authors: Dang Yanna , Li Shuang , Zhao Panpan , Xiao Lieying , Wang Lefeng , Shi Yan , Luo Lei , Wang Shaohua , Wang Huanan , Zhang Kun
Published: September 2021

The genome is transcriptionally inert at fertilization and must be activated through a remarkable developmental process called zygotic genome activation (ZGA). The gene expression pattern formed over the course of ZGA is required for establishing totipotency in early embryos and subsequent development. Substantial epigenetic reprogramming contributes significantly to the pronounced change in gene expression during ZGA, however the mechanism has yet to be resolved. Here, we find histone deacetylase 1 and 2 (HDAC1/2) are critical histone modifiers that regulate ZGA through the histone deacetylase activity. Notably, we show that H3K27ac level declines dramatically during ZGA with a dynamic change in its genome-wide distribution. In mouse embryos, ectopic expression of HDAC1/2 dominant negative mutant leads to a failure of H3K27ac removal and a developmental arrest at 2-cell stage. RNA-seq results reveal a remarkable transcriptomic change with 6565 differentially expressed genes identified. Further analysis shows 64% of down-regulated genes are ZGA genes and 49% of up-regulated genes are developmental genes. Low input ChIP-seq analysis exhibits an increase and decrease of H3K27ac enrichment at the promoter region of up- and down-regulated genes, respectively. Moreover, HDAC1 mutants prohibited removal of broad H3K4me3 domain via impeding the expression of Kdm5s during ZGA. Importantly, the developmental block can be greatly overcome through injection of Kdm5b mRNA and expression of the majority of dysregulated genes partially corrected. Similar functional significance of HDAC1/2 in ZGA is conserved in bovine embryos. Together, we propose that HDAC1/2 is indispensable for mouse and bovine ZGA via creating correct transcriptional repressive and active states.
Addressing Enzymatic-Independent Tumor-Promoting Function of NAMPT via PROTAC-Mediated Degradation
Authors: Zhu Xiaotong , Liu Haixia , Chen Li , Cang Yong , Jiang Biao , Yang Xiaobao , Fan Gaofeng
Published: September 2021

The rate-limiting enzyme of salvage pathway for NAD+ synthesis, NAMPT, is aberrantly overexpressed in a variety of tumor cells and is a poor prognosis factor for patient survival. NAMPT plays a major role in tumor cell proliferation, acting concurrently as an NAD+ synthase and unexpectedly, an extracellular ligand for several tumor-promoting signaling pathways. While previous efforts to modulate NAMPT activity were limited to enzymatic inhibitors with low success in clinical studies, protein degradation offers a possibility to simultaneously disrupt NAMPT’s enzyme activity and ligand capabilities. Here, we report the development of two highly selective NAMPT-targeted proteolysis-targeting chimeras (PROTACs), which promoted rapid and potent NAMPT degradation in a cereblon-dependent manner in multiple tumor cell lines. Notably, both PROTAC degraders outperform a clinical candidate, FK866, in killing effect on hematological tumor cells. These results emphasize the importance and feasibility of applying PROTACs as a better strategy for targeting proteins like NAMPT with dual tumor-promoting functions, which are not easily achieved by conventional enzymatic inhibitors.
Planning and optimization of robotic pick-and-place operations in highly constrained industrial environments
Authors: Tipary Bence , Kovács András , Erdős Ferenc Gábor
Published: September 2021

Purpose

The purpose of this paper is to give a comprehensive solution method for the manipulation of parts with complex geometries arriving in bulk into a robotic assembly cell. As bin-picking applications are still not reliable in intricate workcells, first, the problem is transformed to a semi-structured pick-and-place application, then by collecting and organizing the required process planning steps, a methodology is formed to achieve reliable factory applications even in crowded assembly cell environments.

Design/methodology/approach

The process planning steps are separated into offline precomputation and online planning. The offline phase focuses on preparing the operation and reducing the online computational burdens. During the online phase, the parts laying in a semi-structured arrangement are first recognized and localized based on their stable equilibrium using two-dimensional vision. Then, the picking sequence and corresponding collision-free robot trajectories are planned and optimized.

Findings

The proposed method was evaluated in a geometrically complex experimental workcell, where it ensured precise, collision-free operation. Moreover, the applied planning processes could significantly reduce the execution time compared to heuristic approaches.

Research limitations/implications

The methodology can be further generalized by considering multiple part types and grasping modes. Additionally, the automation of grasp planning and the enhancement of part localization, sequence planning and path smoothing with more advanced solutions are further research directions.

Originality/value

The paper proposes a novel methodology that combines geometrical computations, image processing and combinatorial optimization, adapted to the requirements of flexible pick-and-place applications. The methodology covers each required planning step to reach reliable and more efficient operation.
Sulfide stress cracking inhibition of a supermartensitic stainless steel by using an imidazol obtained from palm oil
Authors: Gonzalez-Rodriguez Jose-Gonzalo , Carmona Hernandez Andres , Vázquez-Vélez E. , Contreras-Cuevas A. , Uruchurtu Chavarin Jorge
Published: September 2021

Purpose

This paper aims to use an imidazole-based n-ionic Gemini surfactant derived from palm oil to inhibit the sulfide stress corrosion cracking of a supermartensitic stainless steel.

Design/methodology/approach

The slow strain rate testing technique, hydrogen permeation tests and potentiodynamic polarization curves have been used.

Findings

Addition of the inhibitor below the critical micelle concentration (CMC) decreased the corrosion current density (icorr), but not enough to avoid embrittlement due to the entry of hydrogen into the steel. Instead, the addition of the inhibitor close to the CMC decreased the icorr, suppressed the entry of hydrogen and inhibited the sulfide stress cracking of steel. Finally, the addition of inhibitor above the CMC led to a slight increase of icorr and promoted localized corrosion, however, the sulfide stress cracking of steel was inhibited.

Originality/value

A green sulfide stress corrosion cracking inhibitor of a supermartensitic stainless steel has been obtained.
Effect of Cu addition to low-alloy structural steel on the resistance against corrosion by the Pseudomonas aeruginosa biofilm
Authors: Li Zhong , Lei Qing , Huang Luyao , Liu Chao
Published: September 2021

Purpose

Low-alloy structural steels (LASS) face severe microbiologically influenced corrosion (MIC) in their service environments. To mitigate this issue, Cu is often used as an alloying element owing to its intrinsic antimicrobial activity. However, the antibacterial performance and biofilm resistance of Cu-containing LASS (Cu-LASS) are still unclear. This study aims to analyze the effect of Cu addition to 420 MP LASS on its MIC by the Pseudomonas aeruginosa biofilm.

Design/methodology/approach

Scanning electron microscope, confocal laser scanning microscope and X-ray photoelectron spectroscopy were used to analyze the surface morphology and composition of corrosion products. The antibacterial activities of Cu-LASS were analyzed by the spread-plate method. In addition, electrochemical analysis was conducted to characterize the corrosion behavior of the produced alloy.

Findings

Bacterial analysis and morphological observation confirmed a reduced sessile cell count and inactivation of the P. aeruginosa biofilm on the surface of Cu-LASS coupons. Electrochemical measurements showed that Cu-LASS exhibited large polarization and charge-transfer resistances, which indicated excellent MIC resistance. This significantly enhanced resistance to MIC could be explained by the synergistic effect of released Cu2+ from the Cu-LASS surface and immediate contact to Cu-rich phase in the surface and the release of Cu2+ ions from the Cu-LASS surface.

Originality/value

The effect of Cu addition on the MIC resistance and antibacterial performance of LASS is seldom reported. It is necessary to investigate the corrosion resistance of Cu-LASS and clarify its antibacterial mechanism. This paper fulfills this need.
Big Five personality traits and green consumption: bridging the attitude-intention-behavior gap
Authors: Duong Cong Doanh
Published: September 2021

Purpose

This study examines the roles of Big Five personality traits, including conscientiousness, agreeableness, extraversion, neuroticism and openness to experience, in shaping green consumption behavior, as well as bridging the attitude-intention-behavior gap in environmentally friendly consumption and testing the gender differences between these associations.

Design/methodology/approach

A dataset of 611 consumers was collected by means of mall-intercept surveys in major Vietnamese cities. Structural equation modeling (SEM) via AMOS 24.0 was employed to test the proposed conceptual framework and hypotheses, while the PROCESS approach was utilized to estimate mediation standardized regression coefficients.

Findings

The study revealed that in addition to extraversion, other personality traits (agreeableness, conscientiousness, openness to experience and neuroticism) were strongly associated with green consumption. Moreover, attitude towards green products and intention to buy environmentally friendly products were determined to have key roles in explaining consumers' pro-environmental behavior. There was also a notable difference in the impact of personality traits on men's and women's green consumption.

Practical implications

This study provides useful recommendations for administrational practices seeking to understand consumer behavior, build appropriate marketing and communication campaigns and attract customers to buy environmentally friendly products.

Originality/value

This study makes efforts to resolve the attitude-intention-behavior gap, a recurring theme in the green consumption literature, as well as illustrates the significance of Big Five personality traits in explaining attitude, intention and behavior when purchasing green products. This research also demonstrates that Big Five personality traits have significantly different effects on green consumption attitudes and intention to carry out pro-behavioral consumption.