bluuebunny/crossref_metadata_2025 huggingface dataset:
Biochemical analysis of anthocyanin and proanthocyanidin and their regulation in determining chickpea flower and seed coat colours
Authors: Pal Lalita , Dwivedi Vikas , Gupta Santosh Kumar , Saxena Samiksha , Pandey Ashutosh , Chattopadhyay Debasis
Published: August 2022

Flower and seed coat colour are important agronomic traits in chickpea (Cicer arietinum L.). Cultivated chickpeas are of two types namely, desi (dark seeded, purple flower) and kabuli (light colour seeded, white flower). There has been limited information about the molecular mechanism underlying the colour variation of flower and seed coats in desi and kabuli chickpea. We profiled the anthocyanin and proanthocyanidin (PA) contents in chickpea flowers and seed coats. Tissue-specific silencing of two genes encoding a basic helix-loop-helix (CabHLH) protein and a tonoplast-localized multidrug and toxic compound extrusion (CaMATE1) transporter in a desi genotype resulted in the reduction in expressions of anthocyanin and PA biosynthetic genes and anthocyanin and PA contents in the flower and seed coat and produced flowers and seeds with kabuli characteristics. Transcriptional regulation of a subset of anthocyanin and PA biosynthetic genes by a natural CabHLH variant and transport assay of a natural CaMATE1 variant explained the association of these alleles with the kabuli phenotype. We carried out a detailed molecular characterization of these genes, and provided evidences that kabuli chickpea flower and seed colour phenotype can be derived by manipulation of single genes in a desi chickpea background.

Highlight

In this study, we have defined the molecular link between flower and seed color in chickpea and identified CaMATE1 and CabHLH as the regulators of both the traits.
A highly polymorphic effector protein promotes fungal virulence through suppression of plant-associated Actinobacteria
Authors: Snelders Nick C. , Boshoven Jordi C. , Song Yin , Schmitz Natalie , Fiorin Gabriel L. , Rovenich Hanna , van den Berg Grardy C.M. , Torres David E. , Faino Luigi , Seidl Michael F. , Thomma Bart P.H.J.
Published: August 2022

Plant pathogens secrete effector proteins to support host colonization through a wide range of molecular mechanisms, while plant immune systems evolved receptors to recognize effectors or their activities to mount immune responses to halt pathogens. Importantly, plants do not act as single organisms, but rather as holobionts that actively shape their microbiota as a determinant of health, and may thus be targeted by pathogen effectors as such. The soil-borne fungal pathogen Verticillium dahliae was recently demonstrated to exploit the VdAve1 effector to manipulate the host microbiota to promote vascular wilt disease in absence of the corresponding immune receptor Ve1. We now identified a multiallelic V. dahliae gene displaying ~65% sequence similarity to VdAve1, named VdAve1-like (VdAve1L). Interestingly, VdAve1L shows extreme sequence variation, including alleles that encode dysfunctional proteins, indicative of selection pressure to overcome host recognition. We show that the orphan cell surface receptor Ve2, encoded at the Ve1 locus, does not recognize VdAve1L. Furthermore, we show that the full-length variant VdAve1L2 possesses antimicrobial activity, like VdAve1, yet with a divergent activity spectrum. Altogether, VdAve1L2 is exploited by V. dahliae to mediate tomato colonization through the direct suppression of antagonistic Actinobacteria in the host microbiota. Our findings open up strategies for more targeted biocontrol against microbial plant pathogens.
Evolutionary determinants of reproductive seasonality: a theoretical approach
Authors: Burtschell Lugdiwine , Dezeure Jules , Huchard Elise , Godelle Bernard
Published: August 2022

Reproductive seasonality is a major adaptation to seasonal cycles and varies substantially among organisms. This variation, which was long thought to reflect a simple latitudinal gradient, remains poorly understood for many species, in part due to a lacunary theoretical framework. Because seasonal cycles are increasingly disrupted by climate change, a better understanding of the ecology of reproductive seasonality could generate important insights on how climate change may affect biodiversity. The goal of this study was to investigate the drivers of evolutionary transitions towards reproductive seasonality using a realistic agent-based optimisation model simulating the life cycle of a female yellow baboon, who typically breeds year-round. Specifically, we tested the influence of three ecological traits (environmental seasonality, productivity and unpredictability) and three life-history traits (daily reproductive energy expenditure, reproductive cycle length and infant extrinsic mortality) on the intensity of reproductive seasonality. To do so, we simulated diverse reproductive phenology strategies (from non-seasonal to highly seasonal), assessed which were optimal and computed, for the set of optimal strategies, the intensity of reproductive seasonality. We then induced variation in each trait of interest and examined how it affected the intensity of reproductive seasonality. We found significant effects of all three environmental traits: high reproductive seasonality was favoured by high environmental seasonality, low environmental productivity and low unpredictability. It was further, and most strongly, favoured by high daily reproductive energy expenditure. In contrast, there was no significant effect of reproductive cycle length and infant extrinsic mortality. Our modelling approach successfully disentangled the effects of environmental seasonality, productivity and unpredictability on the intensity of reproductive seasonality, which likely all contribute to generate the well-known association between latitude and reproductive seasonality. Our results further highlight the critical importance of life history pace on the evolution of reproductive seasonality. Overall, this study contributes a powerful theoretical framework and modelling tool that may apply across the life-history space, as well as sheds new light on the emergence and maintenance of non-seasonal breeding in slow-living species, including humans.
Targeted genome editing in grape using multiple CRISPR-guided editing systems
Authors: Ren Chong , Lin Yanping , Li Huayang , Li Shaohua , Liang Zhenchang
Published: August 2022

The CRISPR/Cas9 system, together with newly developed CRISPR technologies such as CRISPR/LbCpf1 and base editors, have expanded the scope of targeted genome editing in plants. However, in grape, applications of these novel CRISPR-guided tools have not been reported. Here, we employed EGFP (enhanced green fluorescent protein) and RUBY to help to screen transformed grape cells based on fluorescence and red betalain and tested the activities of CRISPR/LbCpf1, CRISPR/xCas9 and cytidine base editor (CBE) in grape, respectively. The grape TMT1 (tonoplastic monosaccharide transporter1) and TMT2 genes were simultaneously edited by using LbCpf1, resulting in an efficiency of 16-48%. Furthermore, high temperature (34°C) could enhance editing efficiencies at most of the designed targets. The CRISPR/xCas9 could induce targeted mutagenesis at the target with NGG PAM, but the efficiencies were very low (< 1.9%). The targets with GAA and GAT PAMs that are reported in mammalian cells and rice were not recognized by xCas9 in our study. Moreover, successful C-to-T substitutions were achieved in GAI1 (gibberellin insensitive1) gene by using CBE. The editing efficiencies ranged from 2.4 to 15% at the two targets in GAI1 in grape cells. Analysis of independent embryos revealed a C-to-T efficiency of 12.5% at the first target of GAI1. Taken together, our results demonstrate the efficacy of these CRISPR-guided tools in grape and provide evidence for further application of these editing tools in this economically important species.
The regulatory role of anti-sigma factor, RsbW, in Clostridioides difficile stress response, persistence and infection
Authors: Cheng Jeffrey K. J. , Đapa Tanja , Chan Ivan Y. L. , MacCreath Thomas , Slater Ross , Unnikrishnan Meera
Published: August 2022

The anaerobic pathogen Clostridioides difficile, a primary cause of antibiotic-associated diarrhoea, faces a variety of stresses in the environment and in the mammalian gut. To cope with environmental stresses, it uses the alternative sigma factor B (σB) to modulate gene transcription, which is regulated by an anti-sigma factor, RsbW. To understand the role of RsbW in C. difficile physiology, a rsbW mutant (ΔrsbW) where σB is ‘always on’, was generated. ΔrsbW did not have deleterious fitness defects but tolerated acidic environments and detoxified reactive oxygen and nitrogen species better. ΔrsbW was defective in spore and biofilm formation, adhered better to human gut epithelia and was less virulent in a Galleria mellonella infection model. A transcriptomic analysis to understand this unique phenotype showed a change in expression of some σB-controlled genes along with several non-σB controlled genes. Interestingly, sinRR’ locus that encodes a pleiotropic regulator, was highly upregulated in ΔrsbW indicating a potential indirect role for σB or RsbW in control of sinRR’. Furthermore, the unexpected lower intracellular levels of σB observed suggest post translational control mechanisms. Our study thus provides new insight into the regulatory role of RsbW and the complexity of regulatory networks in C. difficile.

Importance

Pathogens, like C. difficile, face a range of stresses in the environment and within the host. Alternate transcriptional factors such as sigma factor B (σB) enables the bacterium to respond quickly to different types of stresses and are conserved across bacteria. Anti-sigma factors like RsbW control the activation of genes via these pathways. Such transcriptional control systems provide pathogens like C. difficile a route to tolerance and detoxification of harmful compounds. In this study we investigate the role of RsbW in C. difficile physiology. We demonstrate distinctive phenotypes for a rsbW mutant in growth, persistence and virulence. Our data suggest new σB regulatory circuits in C. difficile. Understanding bacterial responses to external stress is key to designing better strategies to combat this highly resilient bacterial pathogen.
Human conflict with wild carnivores and herbivores: a comparison of socio-economics, damage and prey preference of wild carnivores
Authors: Bayani Abhijeet , Dandekar Nikhil
Published: August 2022

Human-wildlife conflict (HWC) at the fringes of protected areas is a major concern in the conservation biology. Although damage caused by carnivores and herbivores may vary in their magnitude, more attention has been given to carnivores due to various reasons. In Tadoba-Andhari Tiger Reserve (TATR) we compared economic dependence of locals on agriculture and livestock independently and found that income from livestock is only additional, whereas agriculture is the major source of livelihood. We also studied the relative abundance of wild herbivores and diet of tiger, leopard and sloth bear and found that these carnivores have largely been dependent on chital, nilgai and wild pig despite livestock population being higher. We found HWC mitigation in TATR effective but biased towards big cats while the damage in main livelihood (agriculture) being higher. We recommend higher attention to crop-raiding problem for the successful wildlife conservation in central India.
A Peronophythora litchii RXLR effector PlAvh202 enhances plant susceptibility by destabilizing host SAMS
Authors: Li Peng , Li Wen , Zhou Xiaofan , Situ Junjian , Xie Lizhu , Xi Pinggen , Yang Bo , Kong Guanghui , Jiang Zide
Published: August 2022

Oomycete pathogens can secret hundreds of effectors into plant cells to interfere with plant immune system during infection. Here we identified a cytoplasmic RXLR effector from the most destructive pathogen of litchi, Peronophythora litchii, and named it as PlAvh202. PlAvh202 could suppress cell death triggered by INF1 and Avr3a/R3a in Nicotiana benthamiana, and was essential for P. litchii virulence. In addition, PlAvh202 also suppressed plant immune responses and promoted the susceptibility of N. benthamiana to Phytophthora capsici. Further research revealed that PlAvh202 could suppress ethylene (ET) production by targeting and destabilizing plant S-adenosyl-L-methionine synthetase (SAMS), a key enzyme in ET biosynthesis pathway, in a 26S proteasome-dependent manner. Transient expression of LcSAMS3 induced ET production and enhanced plant resistance, whereas inhibition of ET biosynthesis promoted P. litchii infection, supporting that LcSAMS and ET positively regulate litchi immunity towards P. litchii. Overall, these findings highlight that SAMS can be targeted by oomycete RXLR effector to manipulate ET-mediated plant immunity.
Development of molecular markers of the Kunitz trypsin inhibitor mutant alleles generated by CRISPR/Cas9-mediated mutagenesis in soybean
Authors: Wang Zhibo , Shea Zachary , Rosso Luciana , Shang Chao , Li Jianyong , Bewick Patrick , Li Qi , Zhao Bingyu , Zhang Bo
Published: August 2022

The digestibility of soybean meal can be severely impacted by trypsin inhibitor (TI), one of the most abundant anti-nutritional factors present in soybean seeds. TI can restrain the function of trypsin, a critical enzyme that breaks down proteins in the digestive tract. Soybean accessions with low TI content have been identified. However, it is challenging to breed the low TI trait into elite cultivars due to a lack of molecular markers associated with low TI traits. We identified Kunitz trypsin inhibitor 1 (KTI1, Glyma01g095000) and KTI3 (Glyma08g341500) as two seed-specific TI genes. Mutant kti1 and kti3 alleles carrying small deletions or insertions within the gene open reading frames were created in the soybean cultivar Glycine max cv. Williams 82 (WM82) using the CRISPR/Cas9-mediated genome editing approach. The KTI content and TI activity both remarkably reduced in kti1/3 mutants compared to the WM82 seeds. There was no significant difference in terms of plant growth or maturity days of kti1/3 transgenic and WM82 plants in greenhouse condition. We further identified a T1 line, #5-26, that carried double homozygous kti1/3 mutant alleles, but not the Cas9 transgene. Based on the sequences of kti1/3 mutant alleles in #5-26, we developed markers to co-select for these mutant alleles by using a gel-electrophoresis-free method. The kti1/3 mutant soybean line and associated selection markers will assist in accelerating the introduction of low TI trait into elite soybean cultivars in the future.
Local adaptation of host-species specific gut microbiota
Authors: Sprockett Daniel D. , Price Jeffrey D. , Juritsch Anthony F. , Schmaltz Robert J. , Real Madalena V. F. , Goldman Samantha , Sheehan Michael , Ramer-Tait Amanda E. , Moeller Andrew H.
Published: August 2022

Mammalian species harbor compositionally distinct gut microbial communities, but the mechanisms that maintain specificity of symbionts to host species remain unclear. Here we show that natural selection within house mice (Mus musculus domesticus) drives deterministic assembly of the house-mouse gut microbiota from mixtures of native and non-native microbiotas. Competing microbiotas from wild-derived lines of house mice and other mouse species (Mus and Peromyscus spp.) within germ-free wildtype (WT) and Rag1-knockout (Rag1−/−) house mice revealed widespread fitness advantages for native gut bacteria. Certain native Bacteriodetes and Firmicutes favored by selection in WT hosts were not favored or disfavored in Rag1−/− hosts, which lack adaptive immunity, indicating that Rag1 mediates fitness advantages of these strains. This study demonstrates local adaptation of gut microbiota to a mammalian species.

One-Sentence Summary

Adaptive advantages for native bacteria underlie the assembly of the mouse gut microbiota.
Direct control of translational elongation by TAOK2β highlights altered protein synthesis as a fundamental underlying component of autism
Authors: Henis Melad , Rücker Tabitha , Scharrenberg Robin , Richter Melanie , Baltussen Lucas , Meka Durga Praveen , Schwanke Birgit , Neelagandan Nagammal , Daaboul Danie , Murtaza Nadeem , Krisp Christoph , Harder Sönke , Schlüter Hartmut , Kneussel Matthias , Hermans-Borgmeyer Irm , de Wit Joris , Singh Karun K. , Duncan Kent E. , de Anda Froylan Calderón
Published: August 2022

Microdeletions in the 16p11.2 region of the human genome are frequently associated with autism spectrum disorders (ASDs), but how these genomic rearrangements cause ASD remains unclear. Here, we reveal that TAOK2β, a protein isoform encoded by the human TAOK2 gene located in the 16p11.2 locus, regulates mRNA translation. To identify key functional interaction partners of TAOK2β, we performed proteomic screening from Neuro-2a (N2a) cells, mouse cortices, and cultured neurons. This revealed translation factors as a major class of enriched interacting proteins. Consistently, TAOK2β is present in mouse cortical polyribosomes and cortices from Taok2 knockout mice show increased ribosome density on mRNAs and enhanced protein synthesis. Several lines of evidence support an effect of TAOK2β on translation elongation via phosphorylation of eukaryotic elongation factor (eEF2). TAOK2 can directly phosphorylate eEF2 on Threonine 56 and this phosphorylation is reduced in cortices from Taok2 knockout mice. TAOK2β WT overexpression increased eEF2 phosphorylation levels and reduced protein synthesis, whereas a kinase-dead allele of TAOK2β showed opposite effects. Finally, we show that cortices from the mouse model of the human 16p11.2 microdeletion have increased polysome/monosome (P/M) ratios and protein synthesis, phenocopying Taok2 loss of function. Importantly, defective translation phenotypes observed in the mouse 16p11.2 microdeletion model of ASD could be normalized either by reintroducing Taok2 in vivo or by delivering TAOK2β to cortical neurons derived from 16p11.2 microdeletion mice. Our results uncover a critical role of TAOK2β as a regulator of protein synthesis and support the idea that translational control is a common endpoint of ASD-associated signaling pathways.
Infrared videography of a subcutaneous knee tattoo as a simple and inexpensive method to overcome skin motion artifact in rodent kinematics
Authors: Moukarzel George , Rauscher Bradley C. , Patil Chetan A. , Spence Andrew J.
Published: August 2022

Kinematic analyses of rodent behavior are frequently used in neuroscience research, and commonly in spinal cord injury (SCI) studies. Unfortunately, skin motion artifact introduces significant errors into these data, because the skin is only loosely coupled to the underlying skeleton by connective tissue. In rats, these errors can be as large as 50-75%,as quantified by past work using x-ray fluoroscopy. Here we show that infrared videography of a subcutaneous tattoo can overcome skin motion artifact in rodent kinematics. The method yields data similar to gold standard x-ray fluoroscopy systems at a fraction of the cost, does not affect the animals’ locomotion, and results in markers that persist for at least 10 weeks. We found that, compared to a gold-standard x-ray fluoroscopy study that directly tracked the skeleton, our method reduced the error in mean hip angle from 17 ± 6.0 to 3.1 ± 2.4 degrees (mean ± SEM), and the root-mean-square (RMS) error across the mean hip angle waveform from 20 to 5.3 degrees (n=4 rats). The knee joint angle waveform derived from infra-red imaging tightly matched the shape of the x-ray waveform after allowing for a constant offset, having RMS error reduced from 8.1 to 1.2 degrees. The method stands to significantly reduce between-animal errors, and hence between laboratory errors, in these ubiquitous model systems, especially important in SCI studies where individuals are assigned to different treatments.
Direction mutation pressure of SARS-CoV-2 helps to understand the past and predict the future evolution: C>U and G>U biased mutagenesis forces the majority of amino-acid substitutions to be from CG-rich losers to U-rich gainers
Authors: Voronka Alexandr , Efimenko Bogdan , Oreshkov Sergey , Franco Melissa , Fleischmann Zoe , Yurov Valerian , Trufanova Arina , Timonina Valeria , Ree Natalia , Zalevsky Arthur , Penfrat Emma , Junier Thomas , Agranovsky Alexey , Khrapko Konstantin , Gunbin Konstantin , Fellay Jacques , Popadin Konstantin
Published: August 2022

Evolution is a function of mutagenesis and selection. To analyse the role of mutagenesis on the structure of the SARS-CoV-2 genome, we reconstructed the mutational spectrum, which was highly C>U and G>U biased. This bias forces the SARS-CoV-2 genome to become increasingly U-rich unless selection cancels it. We analysed the consequences of this bias on the composition of the most neutral (four-fold degenerate synonymous substitutions) and the least neutral positions (nonsynonymous substitutions). The neutral nucleotide composition is already highly saturated by U and, according to our model, it is at equilibrium, suggesting that in the future, we don’t expect any more increase in U. However, nonsynonymous changes continue slowly evolve towards equilibrium substituting CG-rich amino-acids (“losers”) with U-rich ones (“gainers”). This process is universal for all genes of SARS-CoV-2 as well as for other coronaviridae species. In line with the direction mutation pressure hypothesis, we show that viral-specific amino acid content is associated with the viral-specific mutational spectrum due to the accumulation of effectively neutral slightly deleterious variants (losers to gainers) during the molecular evolution. The tuning of a protein space by the mutational process is expected to be typical for species with relaxed purifying selection, suggesting that the purging of slightly-deleterious variants in the SARS-CoV-2 population is not very effective, probably due to the fast expansion of the viral population during the pandemic. Understanding the mutational process can help to design more robust vaccines, based on gainer-rich motifs, close to the mutation-selection equilibrium.
Endogenous RNAi silences a burgeoning sex chromosome arms race
Authors: Vedanayagam Jeffrey , Lin Ching-Jung , Papareddy Ranjith , Nodine Michael , Flynt Alex S. , Wen Jiayu , Lai Eric C.
Published: August 2022

Although the biological utilities of endogenous RNAi (endo-RNAi) have been largely elusive, recent studies reveal its critical role in the non-model fruitfly Drosophila simulans to suppress selfish genes, whose unchecked activities can severely impair spermatogenesis. In particular, hairpin RNA (hpRNA) loci generate endo-siRNAs that suppress evolutionary novel, X-linked, meiotic drive loci. The consequences of deleting even a single hpRNA (Nmy) in males are profound, as such individuals are nearly incapable of siring male progeny. Here, comparative genomic analyses of D. simulans and D. melanogaster mutants of the core RNAi factor dcr-2 reveal a substantially expanded network of recently-emerged hpRNA-target interactions in the former species. The de novo hpRNA regulatory network in D. simulans bears compelling signatures of sex chromosome conflict and provides insight into molecular strategies that underlie hpRNA emergence. In particular, our data support the existence of ongoing rapid evolution of Nmy/Dox-related networks, recurrent targeting of testis HMG Box loci by hpRNAs, and connections to the piRNA pathway. Importantly, the impact of the endo-RNAi network on gene expression flips the convention for regulatory networks, since we observe strong derepression of targets of the youngest hpRNAs, but only mild effects on the targets of the oldest hpRNAs. These data suggest that endo-RNAi are especially critical during incipient stages of intrinsic sex chromosome conflicts, and that continual cycles of distortion and resolution may contribute to the segregation of species.
Targeting Plasmodium falciparum calcium dependent protein kinase-1: Computational modelling approach towards the discovery of antimalarial drug from medicinal herbs and clinically approved anti-malarial drugs pharmacophores
Authors: Adelusi Temitope Isaac , Ojo Taiwo Ooreoluwa , Quadri Bolaji Olawale , Oyewole Precious Moyosoluwa , Oladipo Elijah Kolawole
Published: August 2022

The Key participants in the calcium signaling process in the malaria parasite are calcium dependent protein kinases. It has been established that Plasmodium falciparum calcium dependent protein kinase 1 (PfCDPK1) is essential for the invasion of host erythrocytes as well as for the parasite’s asexual development. Due to malaria’s resistance to currently available anti-malarial medications, it has become necessary to create effective inhibitory agents that target invasion-dependent proteins. In this study, libraries of 856 chemicals were screened using a variety of computational methods. Physicochemical property testing, molecular docking analysis, and pharmacokinetic ADMET profiling were used to determine these drugs’ capacity to inhibit PfCDPK1. 88 of the 856 compounds derived from known antimalarial botanicals and clinically approved antimalarial drug pharmacophores have demonstrated a potent inhibitory effect on PfCDPK1. Additionally, based on research done on the purported inhibitory standard of PfCDPK1 (Purfalcamine), which had a binding affinity of −8.8 KJ/mol, twelve compounds, which include 17-hydroxyazadiradione, Nimbaflavone, Dehydrocarpaine-II, Picracin, Epicatechin gallate, and Naringin, ZINC000012504271, ZINCC000100029905, ZINC000004149268, ZINC000078832372, ZINC000102739374 and ZINC000198562314 had exhibited better binding affinities (−8.8, −8.8, −9.2, −9.1, −9.3, −10.0, −8.1, −7.9, −7.9, −8.2, −8.3, −7.9 KJ/mol respectively), pharmacokinectics, and physicochemical properties. According to our findings, 88 compounds are likely pharmacological targets with domiciled PfCDPK1 inhibitory effects. Nevertheless, our findings highlight these twelve stand-out compounds, suggesting that they may be the best candidates with regard to therapeutic efficacy.
Loss of chromosome Y in primary tumors
Authors: Qi Meifang , Pang Jiali , Mitsiades Irene , Lane Andrew A. , Rheinbay Esther
Published: August 2022

It has long been recognized that certain cancer types afflict female and male patients disproportionately. The reasons for this disparity include differences in male/female physiology, effect of sex hormones, risk behavior and environmental exposures, and different copy number of the sex chromosomes X and Y. Loss of Y (LOY) is common in peripheral blood cells in aging men, and this phenomenon is associated with several diseases. However, the frequency and role of LOY in tumors is little understood. Here, we present a comprehensive catalog of LOY in >5,000 primary tumors from male patients in the TCGA. We show that LOY rates vary by tumor type, and provide evidence for LOY being either a passenger or driver event depending on context. LOY in uveal melanoma specifically is associated with age, survival and is an independent predictor of poor outcome. LOY creates common dependencies on DDX3X and EIF1AX in male cell lines, suggesting that LOY generates unique vulnerabilities that could be therapeutically exploited.
Robust total X-ray scattering workflow to study correlated motion of proteins in crystals
Authors: Meisburger Steve P. , Case David A. , Ando Nozomi
Published: August 2022

The breathing motions of proteins are thought to play a critical role in function. However, current techniques to study key collective motions are limited to spectroscopy and computation. We present a high-resolution experimental approach based on the total scattering from protein crystals at room temperature (TS/RT-MX) that captures both structure and collective motions. To reveal the scattering signal from protein motions, we developed a general workflow that enables robust subtraction of lattice disorder. The workflow introduces two new methods: GOODVIBES, a detailed and refinable lattice disorder model based on the rigid-body vibrations of a crystalline elastic network; and DISCOBALL, an independent method of validation that estimates the displacement covariance between proteins in the lattice in real space. We demonstrate the robustness of this workflow and further demonstrate how it can be interfaced with MD simulations towards obtaining high-resolution insight into functionally important protein motions.
Autocrine Proteinase Activated Receptor (PAR) mediated signaling in prostate cancer cells
Authors: Chandrabalan Arundhasa , Ramachandran Rithwik
Published: August 2022

Proteinase activated receptors (PARs) are G protein-coupled receptors (GPCRs) activated by limited N-terminal proteolysis. A variety of proteolytic enzymes derived from the coagulation cascade and inflammatory milieu activate PARs, however specific activators in different physiological and pathophysiological contexts remain poorly defined. PARs are highly expressed in many cancer cells and regulate various aspects of tumor growth and metastasis. Endogenous proteinases that regulate PARs in the setting of various tumors however remains unresolved. Prostate cancer (PCa) remains a major cause of mortality in men despite advances in early detection and clinical intervention. PAR expression has been reported in PCa, however, their role here remains poorly defined. In androgen independent PC3 cells, we find functional expression of PAR1 and PAR2 but not PAR4. Using genetically encoded PAR cleavage biosensors, we find that PCa cells secrete proteolytic enzymes that cleave PARs and trigger autocrine signaling. Deletion of PAR1 and PAR2 using CRISPR/Cas9 combined with microarray analysis revealed genes that are differentially regulated by this autocrine signalling mechanism. Interestingly, several genes that are known PCa prognostic factors or biomarker were differentially expressed in PAR1-KO and PAR2-KO PC3 cells. We also examined PAR1 and PAR2 regulation of PCa cell proliferation and migration using PAR1 and PAR2-KO PC3 cells, as well as PAR1 and PAR2 specific agonists and antagonists. We find that PAR1 and PAR2 have opposite effects on PC3 cell proliferation and migration. In summary, we have identified an autocrine signaling mechanism through PARs as a regulator of PCa cell function.
CRISPRa screen identifies a role for c-KIT signaling in tamoxifen resistance, potentially through upregulation of ABC transporters
Authors: Marks Brooke A. , Choate Lauren A. , Sams Kelly , Zhu Lina , Waidyaratne Gavisha , Danko Charles G. , Coonrod Scott A.
Published: August 2022

Resistance to endocrine therapy is a common problem in patients with estrogen receptor alpha (ERα) positive breast cancer. In this study, we took a non-biased genome-wide approach to identify novel mechanisms of endocrine resistance using a clustered regularly interspaced short palindromic repeats (CRISPR) activating (CRISPRa) screen. Results from the screen identified 109 candidate resistance-associated genes, with several of these genes, such as EGFR and SRC, having been previously associated with endocrine resistance. One candidate gene that has not been previously associated with endocrine resistance is the tyrosine kinase receptor, c-KIT. We further tested for associations between c-KIT and endocrine resistance and found that c-KIT overexpressing cells proliferate more rapidly in the presence of tamoxifen compared to control cell lines. To gain deeper insight into the potential role of c-KIT signaling in tamoxifen resistance, we next performed precision nuclear run-on and sequencing (PRO-seq) analysis of c-KIT overexpressing cells to identify downstream factors that may mediate the c-KIT response. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of the overexpressed genes found that the only class of factors that was significantly induced by c-KIT was the ATP-binding cassette (ABC) transporters; specifically, ABCA1, ABCA4, and ABCG1. Interestingly, overexpression of two of these ABC transporters, ABCA1 and ABCG1, significantly correlated with worse prognosis in ERα+ breast cancer patients following endocrine therapy. We then tested for potential therapeutic effects of c-KIT inhibition on endocrine resistance and found that the c-KIT inhibitor Gleevec appears to synergize with tamoxifen to suppress MCF-7-S cell growth. Together, our findings support the hypothesis that c-KIT signaling promotes endocrine resistance via the induction of ABC transporter activity. Additionally, our studies suggest that inhibition of c-KIT signaling may represent a novel strategy for preventing or overcoming endocrine resistance in ERα+ patients.
Accurate prediction of transition metal ion location via deep learning
Authors: Dürr Simon L. , Levy Andrea , Rothlisberger Ursula
Published: August 2022

Metal ions are essential cofactors for many proteins. In fact, currently, about half of the structurally characterized proteins contain a metal ion. Metal ions play a crucial role for many applications such as enzyme design or design of protein-protein interactions because they are biologically abundant, tether to the protein using strong interactions, and have favorable catalytic properties e.g. as Lewis acid. Computational design of metalloproteins is however hampered by the complex electronic structure of many biologically relevant metals such as zinc that can often not be accurately described using a classical force field. In this work, we develop two tools - Metal3D (based on 3D convolutional neural networks) and Metal1D (solely based on geometric criteria) to improve the identification and localization of zinc and other metal ions in experimental and computationally predicted protein structures. Comparison with other currently available tools shows that Metal3D is the most accurate metal ion location predictor to date outperforming geometric predictors including Metal1D by a wide margin using a single structure as input. Metal3D outputs a confidence metric for each predicted site and works on proteins with few homologes in the protein data bank. The predicted metal ion locations for Metal3D are within 0.70 ± 0.64 Å of the experimental locations with half of the sites below 0.5 Å. Metal3D predicts a global metal density that can be used for annotation of structures predicted using e.g. AlphaFold2 and a per residue metal density that can be used in protein design workflows for the location of suitable metal binding sites and rotamer sampling to create novel metalloproteins. Metal3D is available as easy to use webapp, notebook or commandline interface.
Evidence for a travel direction signal in humans that is independent of head direction
Authors: Cheng You , Ling Sam , Stern Chantal E. , Huang Andrew , Chrastil Elizabeth R.
Published: August 2022

We often assume that travel direction is redundant with head direction, but from first principles these two factors provide differing spatial information. Although head direction has been found to be a fundamental component of human navigation, it is unclear how self-motion signals for travel direction contribute to forming a travel trajectory. Employing a novel motion adaptation paradigm from visual neuroscience designed to preclude a contribution of head direction, we found high-level aftereffects of perceived travel direction, indicating that travel direction is a fundamental component of human navigation. Interestingly, we discovered a higher frequency of reporting perceived travel toward the adapted direction compared to a no-adapt control – an aftereffect that runs contrary to low-level motion aftereffects. This travel aftereffect was maintained after controlling for possible response biases and approaching effects, and it scaled with adaptation duration. These findings demonstrate the first evidence of how a pure travel direction signal might be represented in humans, independent of head direction.
An expanded phylogeny of Setaria (Poaceae, Panicoideae, Paniceae) and its relationships within the subtribe Cenchrinae
Authors: Delfini Carolina , Acosta Juan M. , Pensiero José F. , Aliscioni Sandra S.
Published: August 2022

Setaria P. Beauv. is the largest genus of the “bristle clade”, including between 115 and 160 species. Previous molecular phylogenetic studies showed Setaria likely to be para- or polyphyletic, retrieving several clades apparently consistent in all analyses and correlated with the geographic origin of species. In this study, we evaluate the phylogeny of the subtribe Cenchrinae using parsimony, likelihood, and Bayesian inference based on the plastid marker ndhF and increasing the number of sampled species. Our main objective was analyze American taxa with inflorescences of the “Paspalidium type” (i.e., subgenera Paurochaetium and Reverchoniae) to test whether they, as traditionally circumscribed, form a natural group. Our findings recovered both subgenera as polyphyletic, with their species distributed in different morphologically distinctive clades and not necessarily correlated with the geographic origin. Additionally, we were able to include a second voucher of species that were imprecisely located in previous studies and define their placements in the tree, as well as confirm that Setaria is polyphyletic as currently delineated. A comparison with the results from other studies, comments on Stenotaphrum Trin. and a brief discussion on conflicting placements in the “Cenchrus clade”, and of Acritochaete Pilg. are also included here.
Golgi-dependent Reactivation and Regeneration of Quiescent Neural Stem Cells
Authors: Gujar Mahekta R. , Gao Yang , Teng Xiang , Deng Qiannan , Tan Ye Sing , Toyama Yusuke , Wang Hongyan
Published: August 2022

The ability of stem cells to switch between quiescent and proliferative states is crucial for maintaining tissue homeostasis and regeneration. InDrosophila, quiescent neural stem cells (qNSCs) extend a primary protrusion, which is removed prior to NSC reactivation. Here, we have unravelled that qNSC protrusions can be regenerated upon injury. This regeneration process relies on the Golgi apparatus which acts as the major acentrosomal microtubule-organizing centre in qNSCs. A Golgi-resident GTPase Arf1 and its guanine-nucleotide exchange factor Sec71 promote NSC reactivation and regeneration via the regulation of microtubule growth. Arf1 physically associates with its new effector Mini Spindles (Msps)/XMAP215, a microtubule polymerase. Finally, Arf1 functions upstream of Msps to target the cell-adhesion molecule E-cadherin to NSC-neuropil contact sites during NSC reactivation. Our findings have establishedDrosophilaqNSCs as a new regeneration model and identified a novel Arf1/Sec71-Msps pathway in the regulation of microtubule growth and NSC reactivation.
Phenotype switching in a global method for agent-based models of biological tissue
Authors: Bergman Daniel , Jackson Trachette L.
Published: August 2022

Agent-based models (ABMs) are an increasingly important tool for understanding the complexities presented by phenotypic and spatial heterogeneity in biological tissue. The resolution a modeler can achieve in these regards is unrivaled by other approaches. However, this comes at a steep computational cost limiting either the scale of such models or the ability to explore, parameterize, analyze, and apply them. When the models involve molecular-level dynamics, especially cell-specific dynamics, the limitations are compounded. We have developed a global method for solving these computationally expensive dynamics significantly decreases the computational time without altering the behavior of the system. Here, we extend this method to the case where cells can switch phenotypes in response to signals in the microenvironment. We find that the global method in this context preserves the temporal population dynamics and the spatial arrangements of the cells while requiring markedly less simulation time. We thus add a tool for efficiently simulating ABMs that captures key facets of the molecular and cellular dynamics in heterogeneous tissue.

Author summary

Agent-based models (ABMs) are an important tool for understanding how cells and molecular compounds interact to produce complex, emergent behavior. The principal feature of ABMs that set them apart from other types of models is their ability to capture the diversity of cells in a tissue. However, this feature comes at the cost of long simulation times, reducing the ability to apply the findings of these models to improve our understanding of living organisms. We present here a means of simulating ABMs using a more efficient method, called the global method, when the cells are undergoing discrete, phenotypic changes in response to molecular cues. We demonstrate that the global method preserves the key features of the ABM while performing simulations much faster. This allows for more efficient testing of biological hypotheses in a mathematical framework that captures key facets of the diversity in biological tissue.
Structure and Neutralization Mechanism of a Human Antibody Targeting a Complex Epitope on Zika Virus
Authors: Adams Cameron , Carbaugh Derek L. , Shu Bo , Ng Thiam-Seng , Castillo Izabella N. , Bowmik Ryan , Segovia-Chumbez Bruno , Puhl Ana C. , Graham Stephen , Diehl Sean A. , Lazear Helen M. , Lok Shee-mei , de Silva Aravinda M. , Premkumar Lakshmanane
Published: August 2022

We currently have an incomplete understanding of why only a fraction of human antibodies that bind to flaviviruses block infection of cells. Here we define the footprint of a strongly neutralizing human monoclonal antibody (mAb G9E) with Zika virus (ZIKV) by both X-ray crystallography and cryo-electron microscopy. Flavivirus envelope (E) glycoproteins are present as homodimers on the virion surface, and G9E bound to a quaternary structure epitope spanning both E protomers forming a homodimer. As G9E mainly neutralized ZIKV by blocking a step after viral attachment to cells, we tested if the neutralization mechanism of G9E was dependent on the mAb cross-linking E molecules and blocking low-pH triggered conformational changes required for viral membrane fusion. We introduced targeted mutations to the G9E paratope to create recombinant antibodies that bound to the ZIKV envelope without cross-linking E protomers. The G9E paratope mutants that bound to a restricted epitope on one protomer poorly neutralized ZIKV compared to the wild-type mAb, demonstrating that the neutralization mechanism depended on the ability of G9E to cross-link E proteins. In cell-free low pH triggered viral fusion assay, both wild-type G9E, and epitope restricted paratope mutant G9E bound to ZIKV but only the wild-type G9E blocked fusion. We propose that, beyond antibody binding strength, the ability of human antibodies to cross-link E-proteins is a critical determinant of flavivirus neutralization potency.
Choroid plexus volume is enlarged in clinically isolated syndrome patients with optic neuritis
Authors: Klistorner Samuel , Van der Walt Anneke , Barnett Michael H , Butzkueven Helmut , Kolbe Scott , Parratt John , Yiannikas Con , Klistorner Alexander
Published: August 2022

Objectives

People with Multiple Sclerosis (MS) have a larger choroid plexus (CP) volume than healthy controls. We investigated CP volume in early MS by quantitatively assessing brain MRI scans in patients presenting with optic neuritis (ON) as a clinically isolated syndrome (CIS), compared to a cohort with established Relapsing Remitting Multiple Sclerosis (RRMS) and healthy controls.

Methods

Pre- and post-gadolinium 3D-T1, 3D FLAIR and diffusion-weighted images were acquired from 44 CIS ON patients at baseline, 1, 3, 6 and 12 months after the onset of ON. Fifty RRMS patients and 50 healthy controls were also included for comparison.

Results

ANOVA revealed significantly larger CP volumes in both ON CIS and RRMS groups compared to healthy controls (p<0.001 for both), but no difference between ON CIS and RRMS patients (p=0.9)

Twenty-three ON CIS patients who converted to CDMS during 10 years of follow-up demonstrated CP volume similar to RRMS patients, but significantly larger compared to healthy controls (p<0.001). Increased CP volume was identified even in a sub-group of patients without MS-like lesions at baseline (p<0.001).

A significant (∼6%) transient increase of CP volume was observed following a new bout of inflammation, which, however, returned to pre-inflammatory state few months later. CP volume was not associated with the severity of acute inflammation of the optic nerve or long-term optic nerve axonal loss, not with brain lesion load or severity of tissue damage within lesions.

Interpretation

Our data demonstrate that enlarged CP can be observe very early in a disease, transiently reacts to acute inflammation, but not associated with the degree of tissue destruction.
Improved cell-free transcription-translation reactions in microfluidic chemostats augmented with hydrogel membranes for continuous small molecule dialysis
Authors: Lavickova Barbora , Grasemann Laura , Maerkl Sebastian J
Published: August 2022

Increasing protein production capacity of the PURE cell-free transcription-translation (TX-TL) system will be key to implementing complex synthetic biological circuits, and to establish a fully self-regenerating system as a basis for the development of a synthetic cell. Under steady-state conditions, the protein synthesis capacity of the PURE system is likely at least one order of magnitude too low to express sufficient quantities of all PURE protein components. This is in part due to the fact that protein synthesis can’t be sustained during the entire dilution cycle, especially at low dilution rates. We developed a microfluidic chemostat augmented with semi-permeable membranes that combines steady-state reactions and continuous dialysis as a possible solution to enhance protein synthesis at steady-state. In batch operation, the continuous dialysis of low molecular weight components via the membranes extended protein synthesis by over an order of magnitude from 2 hours to over 30 hours, leading to a seven-fold increase in protein yield. In chemostat operation, continuous dialysis enabled sustained protein synthesis during the entire dilution cycle even for low dilution rates, leading to six-fold higher protein levels at steady state. The possibility to combine and independently manipulate continuous dialysis and chemostat operation renders our dialysis chemostat a promising technological basis for complex cell-free synthetic biology applications that require enhanced protein synthesis capacity.
Accuracy of non-parametric species richness estimators across taxa and regions
Authors: Soukainen Arttu , Cardoso Pedro
Published: August 2022

Non-parametric species richness estimators are efficient and widely used when sampling is incomplete. There is little consensus on which of the available estimators works best across taxa and regions. Until now no work compared existing algorithms with multiple datasets encompassing contrasting scenarios.

We used data from 62 inventories worldwide at different spatial scales, including 20 vertebrate, 22 invertebrate and 20 plant datasets, and compared the accuracy of the most used non-parametric estimators (Chao and Jackknife) and improvements to their original formulations.

Our results highlight the good performance of the Jackknife estimators for incidence data, especially the P-corrected first order jackknife estimator (Jack1inP). This algorithm ranked most often the best or among the best performing estimators using two measures of accuracy that measure deviation from expectation along the accumulation curve.

We argue that Jack1inP can be considered a universal estimator for species richness, regardless of taxon, temporal and spatial scales, or completeness of the sampling. More research should however be directed towards finding the precise contexts when each estimator might perform best.