bluuebunny/crossref_metadata_2025 huggingface dataset:
Pulsatile electrical stimulation creates predictable, correctable disruptions in neural firing
Authors: Steinhardt Cynthia R. , Mitchell Diana E. , Cullen Kathleen E. , Fridman Gene Y.
Published: August 2021

Electrical stimulation is a key tool in neuroscience, both in brain mapping studies and in many therapeutic applications such as cochlear, vestibular, and retinal neural implants. Due to safety considerations, stimulation is restricted to short biphasic pulses. Despite decades of research and development, neural implants lead to varying restoration of function in patients. In this study, we use computational modeling to provide an explanation for how pulsatile stimulation affects axonal channels and therefore leads to variability in restoration of neural responses. The phenomenological explanation is transformed into equations that predict induced firing rate as a function of pulse rate, pulse amplitude, and spontaneous firing rate. We show that these equations predict simulated responses to pulsatile stimulation with a variety of parameters as well as several features of experimentally recorded vestibular afferent responses to pulsatile stimulation. We then discuss the implications of these effects for improving clinical stimulation paradigms and electrical stimulation-based experiments.
Before trilobite legs:Pygmaclypeatus daziensisreconsidered and the ancestral appendicular organization of Cambrian artiopods
Authors: Schmidt Michel , Hou Xianguang , Zhai Dayou , Mai Huijuan , Belojević Jelena , Chen Xiaohan , Melzer Roland R. , Ortega-Hernández Javier , Liu Yu
Published: August 2021

The Cambrian Stage 3 Chengjiang biota in South China is one of the most influential Konservat-Lagerstätten worldwide thanks to the fossilization of diverse non-biomineralizing organisms through pyritization. Despite their contributions to understanding the evolution of early animals, several Chengjiang species remain poorly known due to their scarcity and/or incomplete preservation. Here, we use micro-computed tomography to reveal in detail the ventral appendage organization of the enigmatic non-trilobite artiopodPygmaclypeatus daziensis– one of the rarest euarthropods in Chengjiang – and explore its functional ecology and broader evolutionary significance.Pygmaclypeatus daziensispossesses a set of uniramous antennae and 14 pairs of post-antennal biramous appendages, the latter of which show an unexpectedly high degree of heteronomy based on the localized differentiation of the protopodite, endopodite and exopodite along the antero-posterior body axis. The small body size (less than 2 cm), presence of delicate spinose endites, and well-developed exopodites with multiple paddle-shaped lamellae on the appendages ofP. daziensisindicate a nekto-benthic mode of life, and a scavenging/detritus feeding strategy.Pygmaclypeatus daziensisshows that appendage heteronomy is phylogenetically widespread within Artiopoda – the megadiverse clade that includes trilobites and their relatives with non-biomineralizing exoskeletons – and suggests that a single exopodite lobe with paddle-like lamellae is ancestral for this clade.

Abstract Figure

Cover image:Morphological reconstruction of the non-trilobite artiopodPygmaclypeatus daziensisfrom the early Cambrian (Stage 3) Chengjiang biota in south China. Artwork by Holly Sullivan (https://www.sulscientific.com/).
Kinetochore-mediated microtubule assembly and Augmin-dependent amplification drive k-fiber maturation in mammals
Authors: Almeida Ana C. , Oliveira Joana , Drpic Danica , Cheeseman Liam P. , Damas Joana , Lewin Harris A. , Larkin Denis M. , Aguiar Paulo , Pereira António J. , Maiato Helder
Published: August 2021

Chromosome segregation in mammals relies on the maturation of a thick bundle of kinetochore-attached microtubules known as k-fibers. How k-fibers mature from initial kinetochore-microtubule attachments remains a fundamental question. By combining functional perturbations in Indian muntjac, a placental mammal with the lowest known chromosome number (n=3) and distinctively large kinetochores, with fixed- and subsecond live-cell super-resolution CH-STED nanoscopy, we uncovered the mechanism by which Augmin mediates k-fiber maturation. Augmin promoted kinetochore microtubule turnover by sustaining microtubule formation from kinetochores and poleward flux, regardless of pioneer centrosomal microtubules. Tracking of microtubule growth events within individual k-fibers revealed a wide angular dispersion, consistent with Augmin-mediated branched microtubule nucleation. Augmin depletion reduced the frequency of microtubule growth events within k-fibers and hampered normal repair after acute k-fiber injury by laser microsurgery. Our work underscores the contribution of microtubule formation from kinetochores and Augmin-mediated microtubule amplification for k-fiber maturation and spindle assembly in mammals.
Comparative secretomics identifies conserved WAxR motif-containing effectors in rust fungi that suppress cell death in plants
Authors: Jaswal Rajdeep , Dubey Himanshu , Kiran Kanti , Rawal Hukam , Rajarammohan Sivasubramanian , Prasad Pramod , Bhardwaj Subhash C , Sonah Humira , Deshmukh Rupesh , Gupta Naveen , Sharma Tilak Raj
Published: August 2021

Identification of novel effectors with conserved features has always remained a challenge in plant-pathogen interaction studies. The introduction of the genomics era in plant-pathogen studies has led to the identification of significant candidate effectors with novel motifs such as RxLR and dEER motifs. However, in the case of fungal pathogens, limited conserved motifs associated with effectors have been discovered yet. In the present study, we have performed comparative secretome analysis for major plant pathogens of diverse nutrition mechanisms with the aim of dissecting the features underlying their corresponding secretome and conserved motifs. We showed that rust fungi possess the lowest Cell wall degrading enzymes (CWDEs) consortium lower than other biotrophic pathogens. We also showed rust fungi possess the highest secretory superoxide dismutase (SOD) than other studied plant pathogens. Further, we prioritized the candidate secretory effectors proteins (CSEPs) of all the studied pathogens by combining various effector mining parameters to highlight the candidates with potential effector features. A novel WAxR motif in conjugation with the Y/F/WxC (FGC) motif was identified in the effectors of variousP. striiformisraces present globally. The WAxR/WAxR like motifs ( WxxR, WAxx, xAxR) containing effectors were also found in the secretome of other rust fungi. Further, the functional validation of two candidate effectors with WAxR motif fromP. striiformis Yr9showed that these effectors localize to the nucleus as well as cytoplasm, and are able to suppress BAX induced cell death inNicotiana benthamiana. The mutation analysis of individual residues of the WAxR motif (W, A, R) however did not affect the cell death suppression nor subcellular localization of these effectors. Overall, the current study reports the presence of novel motifs in large numbers of effectors of rust fungi with cell death suppression features.

Highlights

Secretome analysis of various plant pathogens performed

A prioritization list for candidate effectors designed

A novel WAxR motif was discovered in rust fungal effector proteins

Two WAxR candidate effectors were functionally validated
How synonymous mutations alter enzyme structure and function over long time scales
Authors: Jiang Yang , Neti Syam Sundar , Sitarik Ian , Pradhan Priya , To Philip , Xia Yingzi , Fried Stephen D. , Booker Squire J. , O’Brien Edward P.
Published: August 2021

The specific activity of enzymes can be altered over long time scales in cells by synonymous mutations, which change an mRNA molecule’s sequence but not the encoded protein’s primary structure. How this happens at the molecular level is unknown. Here, we investigate this issue by applying multiscale modeling to threeE. colienzymes - type III chloramphenicol acetyltransferase, D-alanine–D-alanine ligase B, and dihydrofolate reductase. This modeling involves coarse-grained simulations of protein synthesis and post-translational behavior, all-atom simulations as a test of robustness, and QM/MM calculations to characterize enzymatic function. We first demonstrate that our model predicts experimentally measured changes in specific activity due to synonymous mutations. Then, we show that changes in codon translation rates induced by synonymous mutations cause shifts in co-translational and post-translational folding pathways that kinetically partition molecules into subpopulations that very slowly interconvert to the native, functional state. These long-lived states exhibit reduced catalytic activity, as demonstrated by their increased activation energies for the reactions they carry out. Structurally, these states resemble the native state, with localized misfolding near the active sites of the enzymes. The localized misfolding involves noncovalent lasso entanglements - a topology in which the protein backbone forms a loop closed by noncovalent native contacts which is then threaded by another portion of the protein. Such entanglements are often kinetic traps, as they can require a large proportion of the protein to unfold, which is energetically unfavorable, before they can disentangle and attain the native state. The near-native structures of these misfolded states allow them to bypass the proteostasis machinery and remain soluble, as they exhibit similar hydrophobic surface areas as the native state. These entangled structures persist in all-atom simulations as well, indicating that these conclusions are independent of model resolution. Moreover, the structures of long-lived entangled states are supported by agreement with limited-proteolysis mass spectrometry results. Thus, synonymous mutations cause shifts in the co- and post-translational structural ensemble of proteins, whose altered subpopulations lead to long-term changes in the specific activities of some enzymes. The formation of entangled subpopulations is a plausible mechanism through which changes in translation elongation rate alter ensemble-averaged specific activities, which can ultimately affect the efficiency of biochemical pathways and phenotypic traits.
Improved radiation expression profiling in blood by sequential application of sensitive and specific gene signatures
Authors: Mucaki Eliseos J , Shirley Ben C , Rogan Peter K
Published: August 2021

Purpose. Combinations of expressed genes can discriminate radiation-exposed from normal control blood samples by machine learning based signatures (with 8 to 20% misclassification rates). These signatures can quantify therapeutically-relevant as well as accidental radiation exposures. The prodromal symptoms of Acute Radiation Syndrome (ARS) overlap those present in Influenza and Dengue Fever infections. Surprisingly, these human radiation signatures misclassified gene expression profiles of virally infected samples as false positive exposures. The present study investigates these and other confounders, and then mitigates their impact on signature accuracy. Methods. This study investigated recall by previous and novel radiation signatures independently derived from multiple Gene Expression Omnibus datasets on common and rare non-malignant blood disorders and blood-borne infections (thromboembolism, S. aureus bacteremia, malaria, sickle cell disease, polycythemia vera, and aplastic anemia). Normalized expression levels of signature genes are used as input to machine learning-based classifiers to predict radiation exposure in other hematological conditions. Results. Except for aplastic anemia, these blood-borne disorders modify the normal baseline expression values of genes present in radiation signatures, leading to false-positive misclassification of radiation exposures in 8 to 54% of individuals. Shared changes, predominantly in DNA damage response and apoptosis-related gene transcripts in radiation and confounding hematological conditions, compromise the utility of these signatures for radiation assessment. These confounding conditions (sickle cell disease, thromboembolism, S. aureus bacteremia, malaria) induce neutrophil extracellular traps, initiated by chromatin decondensation, DNA damage response and fragmentation followed by programmed cell death. Riboviral infections (for example, Influenza or Dengue fever) have been proposed to bind and deplete host RNA binding proteins, inducing R-loops in chromatin. R-loops that collide with incoming replication forks can result in incompletely repaired DNA damage, inducing apoptosis and releasing mature virus. To mitigate the effects of confounders, we evaluated predicted radiation-positive samples with novel gene expression signatures derived from radiation-responsive transcripts encoding secreted blood plasma proteins whose expression levels are unperturbed by these conditions. Conclusions. This approach identifies and eliminates misclassified samples with underlying hematological or infectious conditions, leaving only samples with true radiation exposures. Diagnostic accuracy is significantly improved by selecting genes that maximize both sensitivity and specificity in the appropriate tissue using combinations of the best signatures for each of these classes of signatures.
Utilizing Digital Pathology to Quantify Stromal Caveolin-1 Expression in Malignant and Benign Ovarian Tumors: Associations with Clinicopathological Parameters and Clinical Outcomes
Authors: Saeed-Vafa Daryoush , Marchion Douglas C. , McCarthy Susan M , Hakam Ardeshir , Lopez Alexis , Wenham Robert M. , Apte Sachin M. , Chen Dung-Tsa , Magliocco Anthony M. , Lancaster Jonathan M. , Reid Brett M. , Permuth Jennifer B.
Published: August 2021

Loss of stromal caveolin-1 (Cav-1) is a biomarker of a cancer-associated fibroblast (CAF) phenotype and is related to progression, metastasis, and poor outcomes in several cancers. The objective of this study was to evaluate the clinical significance of Cav-1 expression in invasive epithelial ovarian cancer (OvCa). Epithelial and stromal Cav-1 expression were quantified in serous OvCa and benign ovarian tissue in two, independent cohorts – one quantified expression using immunohistochemistry (IHC) and the other using multiplex immunofluorescence (IF) with digital image analysis designed to target CAF-specific expression. Cav-1 expression was significantly downregulated in OvCa stroma compared to non-neoplastic stroma using both the IHC (p=0.002) and IF (p=1.8×10-13) assays. OvCa stroma showed Cav-1 downregulation compared to tumor epithelium with IHC (p=1.2×10-24). Conversely, Cav-1 expression was higher in OvCa stroma compared to tumor epithelium with IF (p=0.002). There was moderate correlation between IHC and IF methods for stromal Cav-1 expression (r2= 0.69, p=0.006) whereas there was no correlation for epithelial expression (r2=0.006, p=0.98). Irrespective of the staining assay, neither response to therapy or overall survival correlated with the expression level of Cav-1 in the stroma or tumor epithelium. Our findings demonstrate a loss of stromal Cav-1 expression in ovarian serous carcinomas. Studies are needed to replicate these findings and explore therapeutic implications, particularly for immunotherapy response.
The Mechanosensitive Ion Channel MSL10 Modulates Susceptibility toPseudomonas syringaeinArabidopsis thaliana
Authors: Basu Debarati , Codjoe Jennette M. , Veley Kira M. , Haswell Elizabeth S.
Published: August 2021

Plants sense and respond to molecular signals associated with the presence of pathogens and their virulence factors. Mechanical signals generated during pathogenic invasion may also be important, but their contributions have rarely been studied. Here we investigate the potential role of a mechanosensitive ion channel, MscS-Like (MSL)10, in defense against the bacterial pathogenPseudomonas syringaeinArabidopsis thaliana. We previously showed that overexpression of MSL10-GFP, phospho-mimetic versions of MSL10, and the gain-of-function allelemsl10-3Gall produce dwarfing, spontaneous cell death, and the hyperaccumulation of reactive oxygen species. These phenotypes are shared by many autoimmune mutants and are frequently suppressed by growth at high temperature in those lines. Here, we found that the same was true for all threeMSL10hypermorphs. In addition, we show that the SGT1/RAR1/HSP90 co-chaperone complex was required for dwarfing and ectopic cell death, PAD4 and SID2 were partially required, and the immune regulators EDS1 and NDR1 were dispensable. AllMSL10hypermorphs exhibited reduced susceptibility to infection byP. syringaestrainPtoDC3000,PtoDC3000 expressing the avirulence genesavrRpt2oravrRpm1, but notPto DC3000 hrpL, and showed an accelerated induction ofPR1expression compared to wild-type plants. Nullmsl10-1mutants were delayed inPR1induction and displayed modest susceptibility to infection by COR-deficientPst. Finally, stomatal closure was reduced inmsl10-1loss-of-function mutants in response toPst COR−. These data show that MSL10 modulates pathogen responses and begin to address the possibility that mechanical signals are exploited by the plant for pathogen perception.
Self-organization of human dorsal-ventral forebrain structures by light induced SHH
Authors: De Santis Riccardo , Etoc Fred , Rosado-Olivieri A. Edwin , Brivanlou Ali H.
Published: August 2021

Organizing centers secrete morphogens that specify the emergence of germ layers and the establishment of the body’s axes during embryogenesis. While traditional experimental embryology tools have been instrumental in dissecting the molecular aspects of organizers in model systems, they are impractical in human in-vitro model systems to quantitatively dissect the relationships between signaling and fate along embryonic coordinates. To systematically study human embryonic organizer centers, we devised a collection of optogenetic ePiggyBac vectors to express a photoactivatable Cre-loxP recombinase, that allows the systematic induction of organizer structures by shining blue-light on hESCs. We used a light stimulus to geometrically confine SHH expression in neuralizing hESCs. This led to the self-organization of mediolateral neural patterns from the organizer. scRNA-seq analysis established that these structures represent the dorsal-ventral forebrain, at the end of the first month of development. Here, we show that morphogen light-stimulation is a scalable tool that induces self-organizing centers.
Cap-independent translation and a precisely localized RNA sequence enable SARS-CoV-2 to control host translation and escape anti-viral response
Authors: Slobodin Boris , Sehrawat Urmila , Lev Anastasia , Ogran Ariel , Fraticelli Davide , Hayat Daniel , Zuckerman Binyamin , Ulitsky Igor , Ben-Shmuel Amir , Bar-David Elad , Levy Haim , Dikstein Rivka
Published: August 2021

Translation of SARS-CoV-2-encoded mRNAs by the host ribosomes is essential for its propagation. Following infection, the early expressed viral protein NSP1 binds the ribosome, represseses translation and induces mRNA degradation, while the host elicits anti-viral response. The mechanisms enabling viral mRNAs to escape this multifaceted repression remain obscure. Here we show that expression of NSP1 leads to destabilization of multi-exon cellular mRNAs, while intron-less transcripts, such as viral mRNAs and anti-viral interferon genes, remain relatively stable. We identified a conserved and precisely located cap-proximal RNA element devoid of guanosines that confers resistance to NSP1-meidated translation inhibition. Importantly, the primary sequence rather than the secondary structure is critical for protection. We further show that the genomic 5’UTR of SARS-CoV-2 exhibits an IRES-like activity and promotes expression of NSP1 in an eIF4E-independent and Torin-1 resistant manner. Upon expression, NSP1 enhances cap-independent translation. However, the sub-genomic 5’UTRs are highly sensitive to eIF4E availability, rendering viral propagation partially sensitive to Torin-1. The combined NSP1-mediated degradation of spliced mRNAs and translation inhibition of single-exon genes, along with the unique features present in the viral 5’UTRs, ensure robust expression of viral mRNAs. These features can be exploited as potential therapeutic targets.
Using expert-elicitation to deliver biodiversity monitoring priorities on a Mediterranean island
Authors: Peyton J. , Hadjistylli M. , Tziortzis I. , Erotokritou E. , Demetriou M. , Samuel-Rhoads Y. , Anastasi V. , Fyttis G. , Hadjioannou L. , Ieronymidou C. , Kassinis N. , Kleitou P. , Kletou D. , Mandoulaki A. , Michailidis N. , Papatheodoulou A. , Payiattas G. , Sparrow D. , Sparrow R. , Turvey K. , Tzirkalli E. , Varnava A.I. , Pescott O.L.
Published: August 2021

Biodiversity monitoring plays an essential role in tracking changes in ecosystems, species distributions and abundances across the globe. Data collected through both structured and unstructured biodiversity recording can inform conservation measures designed to reduce, prevent, and reverse declines in valued biodiversity of many types. However, resources for biodiversity monitoring are limited, it is therefore important that funding bodies prioritise actions relative to the requirements in any given region. We addressed this prioritisation requirement through a three-stage process of expert-elicitation, resulting in a prioritised list of twenty biodiversity monitoring needs for Cyprus. Equal priority was assigned to the twenty monitoring needs within three categories: a top nine, a middle five, and a bottom six. The most highly prioritised biodiversity monitoring needs were those related to the development of robust methodologies, and those ensuring a geographic spread of sufficiently skilled and informed contributors. We suggest ways that the results of our expert-elicitation process could be used to support current and future biodiversity monitoring in Cyprus.
Ether anesthetics prevents touch-induced trigger hair calcium-electrical signals excite the Venus flytrap
Authors: Scherzer Sönke , Huang Shouguang , Iosip Anda , Fuchs Ines , Yokawa Ken , AL-Rasheid Khaled A. S. , Heckmann Manfred , Hedrich Rainer
Published: August 2021

Plants do not have neurons but operate transmembrane ion channels and can get electrical excited by physical and chemical clues. Among them the Venus flytrap is characterized by its peculiar hapto-electric signaling. When insects collide with trigger hairs emerging the trap inner surface, the mechanical stimulus within the mechanosensory organ is translated into a calcium signal and an action potential (AP). Here we asked how the Ca2+ wave and AP is initiated in the trigger hair and how it is feed into systemic trap calcium-electrical networks. When Dionaea muscipula trigger hairs matures and develop hapto-electric excitability the mechanosensitive anion channel DmMSL10/FLYC1 and voltage dependent SKOR type Shaker K+ channel are expressed in the sheering stress sensitive podium. The podium of the trigger hair is interface to the flytrap’s prey capture and processing networks. In the excitable state touch stimulation of the trigger hair evokes a rise in the podium Ca2+ first and before the calcium signal together with an action potential travel all over the trap surface. In search for podium ion channels and pumps mediating touch induced Ca2+ transients, we, in mature trigger hairs firing fast Ca2+ signals and APs, found OSCA1.7 and GLR3.6 type Ca2+ channels and ACA2/10 Ca2+ pumps specifically expressed in the podium. Like trigger hair stimulation, glutamate application to the trap directly evoked a propagating Ca2+ and electrical event. Given that anesthetics affect K+ channels and glutamate receptors in the animal system we exposed flytraps to an ether atmosphere. As result propagation of touch and glutamate induced Ca2+ and AP long-distance signaling got suppressed, while the trap completely recovered excitability when ether was replaced by fresh air. In line with ether targeting a calcium channel addressing a Ca2+ activated anion channel the AP amplitude declined before the electrical signal ceased completely. Ether in the mechanosensory organ did neither prevent the touch induction of a calcium signal nor this post stimulus decay. This finding indicates that ether prevents the touch activated, glr3.6 expressing base of the trigger hair to excite the capture organ.
The relationship between benthic nutrient fluxes and bacterial community in Aquaculture Tail-water Treatment Systems
Authors: Nicholaus Regan , Lukwambe Betina , Yanga Wen , Zhenga Zhongming
Published: August 2021

Constructed-wetlands, Biofilms, and sedimentation are potential aquaculture tail-water treatments however their roles on the distribution of benthic microbial community and the way they affect the interaction between microbial community and inorganic nutrient fluxes have not been fully explored. This study applied 16S rRNA high-throughput sequencing technology to investigate the microbial community distribution and their link with nutrient fluxes in an aquaculture tail-water bioremediation system. Results showed that bacterial community compositions were significantly different in constructed-wetland and biofilm treatments (p<0.05) relative to sedimentation. The composition of the 16S rRNA genes among all the treatments was enriched with Proteobacteria, Bacteroidetes, Firmicutes, and Flavobacteria. NMDS analysis showed that the bacterial composition in constructed-wetland and biofilm samples clustered separately compared to those in sedimentation. The Functional-Annotation-of-Prokaryotic-Taxa analysis indicated that the proportions of sediment-microbial-functional groups (aerobic-chemoheterophy, chemoheterotrophy, and nitrate-ammonification combined) in the constructed-wetland treatment were 47%, 32% in biofilm and 13% in sedimentation system. Benthic-nutrient fluxes for phosphate, ammonium, nitrite, nitrate and sediment oxygen consumption differed markedly among the treatments (p<0.05). Canonical correspondence analysis indicated constructed-wetland had the strongest association between biogeochemical contents and the bacterial community relative to other treatments. This study suggests that the microbial community distributions and their interactions nutrient fluxes were most improved in the constructed-wetland followed by the area under biofilm and sedimentation treatment.
Cascading effects of elephant-human interactions in a savanna ecosystem and the implications for ecology and conservation
Authors: Western David , Mose Victor N.
Published: August 2021

Our study monitored the changes in elephant numbers, distribution and ecological impact over a fifty-year period as the free-ranging intermingled movements of wildlife and traditional subsistence pastoralists across the Amboseli ecosystem were disrupted by a national park, livestock ranches, farms, settlements and changing lifestyles and economies.

Elephants compressed into the national park by poaching and settlement turned woodlands to grassland and shrublands and swamps into short grazing lawns, causing a reduction of plant and herbivore diversity and resilience to extreme events. The results echo the ecological findings of high-density elephant populations in protected areas across eastern and southern Africa. The impact has led to the view of elephants in parks being incompatible with biodiversity and to population control measures.

In contrast to Amboseli National Park, we found woody vegetation grew and plant diversity fell in areas abandoned by elephants. We therefore used naturalist and exclosure experiments to determine the density-dependent response of vegetation to elephants. We found plant richness to peak at the park boundary where elephants and livestock jostled spatially, setting up a creative browsing-grazing tension and a patchwork of habitats explaining the plant richness.

A review of prehistorical and historical literature lends support to the Amboseli findings that elephants and people, the two dominant keystone species in the savannas, have been intimately entangled prior to the global ivory trade and colonialism. The findings point to the inadequacy of parks for conserving mega herbivores and as ecological baselines.

The Amboseli study underlines the significance of space and mobility in expressing the keystone role of elephants, and to community-based conservation as a way to win space and mobility for elephants, alleviate the ecological disruption of compressed populations and minimize population management.
Drug resistance mechanisms create targetable proteostatic vulnerabilities in Her2+ breast cancers
Authors: Singh Navneet , Romick-Rosendale Lindsey , Watanabe-Chailland Miki , Privette Vinnedge Lisa M. , Komurov Kakajan
Published: August 2021

Oncogenic kinase inhibitors show short-lived responses in the clinic due to high rate of acquired resistance. We previously showed that pharmacologically exploiting oncogene-induced proteotoxic stress can be a viable alternative to oncogene-targeted therapy. Here, we performed extensive analyses of the transcriptomic, metabolomic and proteostatic perturbations during the course of treatment of Her2+ breast cancer cells with a Her2 inhibitor covering the drug response, resistance, relapse and drug withdrawal phases. We found that acute Her2 inhibition, in addition to blocking mitogenic signaling, leads to significant decline in the glucose uptake, and shutdown of glycolysis and of global protein synthesis. During prolonged therapy, compensatory overexpression of Her3 allows for the reactivation of mitogenic signaling pathways, but fails to re-engage the glucose uptake and glycolysis, resulting in proteotoxic ER stress, which maintains the protein synthesis block and growth inhibition. Her3-mediated cell proliferation under ER stress during prolonged Her2 inhibition is enabled due to the overexpression of the eIF2 phosphatase GADD34, which uncouples protein synthesis block from the ER stress response to allow for active cell growth. We show that this imbalance in the mitogenic and proteostatic signaling created during the acquired resistance to anti-Her2 therapy imposes a specific vulnerability to the inhibition of the endoplasmic reticulum quality control machinery. The latter is more pronounced in the drug withdrawal phase, where the de-inhibition of Her2 creates an acute surge in the downstream signaling pathways and exacerbates the proteostatic imbalance. Therefore, the acquired resistance mechanisms to oncogenic kinase inhibitors may create secondary vulnerabilities that could be exploited in the clinic.
CREB5 reprograms nuclear interactions to promote resistance to androgen receptor targeting therapies
Authors: Hwang Justin , Arafeh Rand , Seo Ji-Heui , Baca Sylvan C. , Ludwig Megan , Arnoff Taylor E. , Richter Camden , Bergom Hannah E. , McSweeney Sean , Rennhack Jonathan P. , Klingenberg Sarah A. , Cheung Alexander TM. , Kwon Jason , So Jonathan , Kregel Steven , Van Allen Eliezer M. , Drake Justin M. , Freedman Mathew L. , Hahn William C.
Published: August 2021

Metastatic castration resistant prostate cancers (mCRPC) are treated with therapies that antagonize the androgen receptor (AR). Nearly all patients develop resistance to AR-targeted therapies (ART). Our previous work identified CREB5 as an upregulated target gene in human mCRPC that promoted resistance to all clinically-approved ART. The mechanisms by which CREB5 promotes progression of mCRPC or other cancers remains elusive. Integrating ChIP-seq and rapid immunoprecipitation and mass spectroscopy of endogenous proteins (RIME), we report that cells overexpressing CREB5 demonstrate extensive reprogramming of nuclear protein-protein interactions in response to the ART agent enzalutamide. Specifically, CREB5 physically interacts with AR, the pioneering actor FOXA1, and other known co-factors of AR and FOXA1 at transcription regulatory elements recently found to be active in mCRPC patients. We identified a subset of CREB5/FOXA1 co-interacting nuclear factors that have critical functions for AR transcription (GRHL2, HOXB13) while others (TBX3, NFIC) regulated cell viability and ART resistance and were amplified or overexpressed in mCRPC. Upon examining the nuclear protein interactions and the impact of CREB5 expression on the mCRPC patient transcriptome, we found CREB5 was associated with TGFβ and Wnt signaling and epithelial to mesenchymal transitions, implicating these pathways in ART resistance. Overall, these observations define the molecular interactions among CREB5, FOXA1, and pathways that promote ART resistance.
Chlamydomonas ATX1 is essential for Cu distribution towards the secretory pathway and maintenance of biomass in conditions demanding cupro-enzyme dependent metabolic pathways
Authors: Pham Keegan L. J. , Schmollinger Stefan , Merchant Sabeeha S. , Strenkert Daniela
Published: August 2021

Copper (Cu) chaperones, of which yeast ATX1 is a prototype, are small proteins with a Cu(I) binding Mx-CxxC motif, and are responsible for directing intracellular Cu towards specific client protein targets that use Cu as a cofactor. The Chlamydomonas reinhardtii ATX1 (CrATX1) was identified because of its high sequence similarity with yeast ATX1. Like the yeast homologue, CrATX1 accumulates in iron-deficient cells (but is not impacted by other metal-deficiencies), and YFP-ATX1 is distributed in the cytoplasm. Reverse genetic analysis using artificial microRNA (amiRNA) to generate lines with reduced CrATX1 abundance and CRISPR/CPF1 to generate ATX1 knock out lines validated a function for ATX1 in iron-poor cells, most likely because of an impact on metalation of the multicopper oxidase FOX1, which is an important component in high-affinity iron uptake. A more general impact on the secretory pathway is indicated by reduced growth of ATX1 mutant lines on guanine as a sole nitrogen source, which we attribute to loss of function of UOX1, a urate oxidase involved in guanine assimilation. The block of Cu trafficking towards the secretory pathway in ATX1 mutants is strikingly evident by a reduced amount of intracellular Cu in all conditions probed in this work.
Impact of Human Serum Proteins on Susceptibility ofAcinetobacter baumanniito Cefiderocol: role of iron transport
Authors: Le Casin , Pimentel Camila , Pasteran Fernando , Tuttobene Marisel R. , Subils Tomas , Escalante Jenny , Nishimura Brent , Arriaga Susana , Carranza Aimee , Vila Alejandro J. , Corso Alejandra , Actis Luis A. , Tolmasky Marcelo E. , Bonomo Robert A. , Ramírez María Soledad
Published: August 2021

Cefiderocol is a siderophore antibiotic that co-opts iron transporters to facilitate cell entry. We show that genes related to iron uptake systems and resistance to β-lactams inAcinetobacter baumanniihave altered expression levels in the presence of human serum, human serum albumin, or human pleural fluid. Cefiderocol MICs are also raised in the presence of the mentioned fluids. Clinical response inA. baumanniiinfections may be related to the interplay of these human factors.
The evolution of fluoroquinolone resistance in Salmonella under exposure to sub-inhibitory concentration of enrofloxacin
Authors: Gu Yufeng , Huang Lulu , Wu Cuirong , Huang Junhong , Hao Haihong , Yuan Zonghui , Cheng Guyue
Published: August 2021

The evolution of resistance in Salmonella to fluoroquinolones (FQs) under a broad range of sub-inhibitory concentrations (sub-MICs) has not been systematically studied. This study investigated the mechanism of resistance development in Salmonella enterica serovar Enteritidis (S. Enteritidis) under sub-MICs of 1/128×MIC to 1/2×MIC of enrofloxacin (ENR), a widely used veterinary FQ. It was shown that the resistance rate and resistance level of S. Enteritidis varied with the increase of ENR concentration and duration of selection. qRT-PCR results demonstrated that the expression of outer membrane porin (OMP) genes, ompF, ompC and ompD, were down-regulated first to rapidly adapt and develop resistance of ≤ 4×MIC, and as the resistance level increased (≥8×MIC), the up-regulated expression of efflux pump genes, acrB, emrB amd mdfA, along with mutations in quinolone resistance-determining region (QRDR) gradually played a decisive role. Cytohubba analysis based on transcriptomic profiles demonstrated that purB, purC, purD, purF, purH, purK, purL, purM, purN and purT were the hub genes for the FQs resistance. ‘de novo’ IMP biosynthetic process, purine ribonucleoside monophosphate biosynthetic process and purine ribonucleotide biosynthetic process were the top three biological processes screened by MCODE. This study first described the dynamics of FQ resistance evolution in Salmonella under a long-term selection of sub-MICs of ENR in vitro. In addition, this work offers greater insight into the transcriptome changes of S. Enteritidis under the selection of ENR and provides a framework for FQs resistance of Salmonella for further studies.
Deep learning-based algorithm for predicting the live birth potential of mouse embryos
Authors: Tokuoka Yuta , Yamada Takahiro G. , Mashiko Daisuke , Ikeda Zenki , Kobayashi Tetsuya J. , Yamagata Kazuo , Funahashi Akira
Published: August 2021

In assisted reproductive technology (ART), embryos produced by in vitro fertilization (IVF) are graded according to their live birth potential, and high-grade embryos are preferentially transplanted. However, the rate of live birth following clinical ART remains low worldwide, suggesting that grading is inaccurate. One explanation is that grading is classically based on the characteristic shape of embryos at a limited number of developmental stages and does not consider the shape of embryos and intracellular structures, e.g., nuclei, at various stages important for normal embryogenesis. Therefore, here we developed a Normalized Multi-View Attention Network (NVAN) that directly predicts live birth potential from nuclear structural features in live-cell fluorescence images taken of mouse embryos across a wide range of stages. The classification accuracy of our method was 83.87%, which greatly exceeded that of existing machine-learning methods and that of visual inspection by embryo culture specialists. By visualizing the features that contributed most to the prediction of live birth potential, we found that the size and shape of the cell nucleus at the morula stage and at the time of cell division were important for live birth prediction. We anticipate that our method will help ART and developmental engineering as a new basic technology for IVF embryo selection.
Optimized detection of allelic imbalances specific for homologous recombination deficiency improves the prediction of clinical outcomes in cancer
Authors: Perez-Villatoro Fernando , Oikkonen Jaana , Casado Julia , Chernenko Anastasiya , Gulhan Doga C. , Tumiati Manuela , Li Yilin , Lavikka Kari , Hietanen Sakari , Hynninen Johanna , Haltia Ulla-Maija , Tyrmi Jaakko S. , Laivuori Hannele , Konstantinopoulos Panagiotis A. , Hautaniemi Sampsa , Kauppi Liisa , Färkkilä Anniina
Published: August 2021

Homologous recombination DNA-repair deficiency (HRD) is a common driver of genomic instability and confers a therapeutic vulnerability in cancer. The accurate detection of somatic allelic imbalances (AIs) has been limited by methods focused onBRCA1/2mutations and using mixtures of cancer types. Using pan-cancer data, we revealed distinct patterns of AIs in high-grade serous ovarian cancer (HGSC). We used machine learning and statistics to generate improved criteria to identify HRD in HGSC (ovaHRDscar). ovaHRDscar significantly predicted clinical outcomes in three independent patient cohorts with higher precision than previous methods. Characterization of 98 spatiotemporally distinct metastatic samples revealed low intra-patient variation and indicated the primary tumor as the preferred site for clinical sampling in HGSC. Further, our approach improved the prediction of clinical outcomes in triple-negative breast cancer (tnbcHRDscar), validated in two independent patient cohorts. In conclusion, our tumor-specific, systematic approach has the potential to improve patient selection for HR-targeted therapies.
Uncovering non-random binary patterns within sequences of intrinsically disordered proteins
Authors: Cohan Megan C. , Shinn Min Kyung , Lalmansingh Jared M. , Pappu Rohit V.
Published: August 2021

Sequence-ensemble relationships of intrinsically disordered proteins (IDPs) are governed by binary patterns such as the linear clustering or mixing of specific residues or residue types with respect to one another. To enable the discovery of potentially important, shared patterns across sequence families, we describe a computational method referred to as NARDINI for Non-random Arrangement of Residues in Disordered Regions Inferred using Numerical Intermixing. This work was partially motivated by the observation that parameters that are currently in use for describing different binary patterns are not interoperable across IDPs of different amino acid compositions and lengths. In NARDINI, we generate an ensemble of scrambled sequences to set up a composition-specific null model for the patterning parameters of interest. We then compute a series of pattern-specific z-scores to quantify how each pattern deviates from a null model for the IDP of interest. The z-scores help in identifying putative non-random linear sequence patterns within an IDP. We demonstrate the use of NARDINI derived z-scores by identifying sequence patterns in three well-studied IDP systems. We also demonstrate how NARDINI can be deployed to study archetypal IDPs across homologs and orthologs. Overall, NARDINI is likely to aid in designing novel IDPs with a view toward engineering new sequence-function relationships or uncovering cryptic ones. We further propose that the z-scores introduced here are likely to be useful for theoretical and computational descriptions of sequence-ensemble relationships across IDPs of different compositions and lengths.
Spherical micro-hole grid for high-resolution retarding field analyzer
Authors: Muro Takayuki , Matsushita Tomohiro , Sawamura Kazumi , Mizuno Jun
Published: August 2021

A wide-acceptance-angle spherical grid composed of numerous micro cylindrical holes was developed to be used for the retarding grid of a display-type retarding field analyzer (RFA) and to enhance the energy resolution (E/ΔE). Each cylindrical hole with a diameter of 50 µm and a depth of 80 µm is directed to the spherical center. The inner radius of the spherical grid is 40 mm. The holed area corresponds to an acceptance angle of ±52°. The E/ΔE of an RFA equipped with the developed holed grid was estimated to be 2000 from a measured Au 4f photoemission spectrum. A clear photoelectron hologram was observed in the Mo 4p core-level region of MoS2, indicating that the RFA with the holed grid is effective for photoelectron holography.
UnitCell Tools, a package to determine unit-cell parameters from a single electron diffraction pattern
Authors: Shi Hong-Long , Li Zi-An
Published: August 2021

Electron diffraction techniques in transmission electron microscopy (TEM) have been successfully employed for determining the unit-cell parameters of crystal phases, albeit they exhibit a limited accuracy compared with X-ray or neutron diffraction, and they often involve a tedious measurement procedure. Here, a new package for determining unit-cell parameters from a single electron diffraction pattern has been developed. The essence of the package is to reconstruct a 3D reciprocal primitive cell from a single electron diffraction pattern containing both zero-order Laue zone and high-order Laue zone reflections. Subsequently, the primitive cell can be reduced to the Niggli cell which, in turn, can be converted into the unit cell. Using both simulated and experimental patterns, we detail the working procedure and address some effects of experimental conditions (diffraction distortions, misorientation of the zone axis and the use of high-index zone axis) on the robustness and accuracy of the software developed. The feasibility of unit-cell determination of the TiO2nanorod using this package is also demonstrated. Should the parallel-beam, nano-beam and convergent-beam modes of the TEM be used flexibly, the software can determine unit-cell parameters of unknown-structure crystallites (typically >50 nm).
An experiential learning-integrated framework to improve problem-solving skills of engineering graduates
Authors: Sangwan Kuldip Singh , Singh Rajni
Published: August 2021

Purpose

Problem solving skills (PSS), an important component of learning outcomes, is one of the desirable skills in engineering graduates as stated by many employers, researchers and government bodies in India for a strong foothold in professional world. There is a need to develop comprehensive understanding and integration of theory (concept) and practice (process) of PSS in the context of experiential learning (EL).

Design/methodology/approach

The present study is qualitative in nature using a conceptual research design focussing on synthesis and model building framework. The key elements of the study are PSS, EL and their integration. The study seeks to develop conceptual integration of PSS across multiple theories and perspectives. It offers an enhanced view of a concept of PSS by summarising and integrating extant knowledge. It presents the complete and comprehensive meaning/definition of PSS. Subsequently, it also explores EL and synthesises the different variants of EL that can be used to develop PSS. Finally, the study builds a theoretical framework that proposes integration and interplay between PSS and EL.

Findings

Problem-solving operates at three levels: problem concept (nature and context), process (stages with strategies) and solution (open-ended). EL can be used as a tool to develop PSS in an integrated manner. It is found that EL and problem-solving interplay with each other as both are cyclic in nature and have commonalities strengthening each other.

Practical implications

The proposed framework can be adopted in engineering education for making the engineering graduates job ready.

Originality/value

The study proposes a framework based on integration of EL and problem-solving focusing on specific aims and goals of the course, learning approaches, learning strategies and authentic learning (learning environment). This integration would bridge the gap between engineering education and industry requirements. EL integrated problem-solving focus on pedagogical knowledge (knowing how to facilitate discussion among learners and curricular knowledge) and instructional knowledge (knowing how to introduce, organise different methods and assess).