bluuebunny/crossref_metadata_2025 huggingface dataset:
Erythrocyte invasion-neutralising antibodies prevent Plasmodium falciparum RH5 from binding to basigin-containing membrane protein complexes
Authors: Jamwal Abhishek , Constantin Cristina E. , Henrich Sebastian , Bildl Wolfgang , Fakler Bernd , Draper Simon J. , Schulte Uwe , Higgins Matthew K.
Published: September 2022

Basigin is an essential host receptor for invasion of Plasmodium falciparum into human erythrocytes, interacting with parasite surface protein PfRH5. PfRH5 is a leading blood-stage malaria vaccine candidate and a target of growth-inhibitory antibodies. However, basigin is not alone on the erythrocyte surface. Instead, we show that it is exclusively found in one of two macromolecular complexes, bound predominantly to either plasma membrane Ca2+-ATPase 1/4, PMCA1/4, or monocarboxylate transporter 1, MCT1. PfRH5 binds to either of these complexes with a higher affinity than to isolated basigin ectodomain, making it likely that these are the physiological targets of PfRH5. PMCA-mediated Ca2+ export is not affected by PfRH5, ruling this out as the mechanism underlying changes in calcium flux at the interface between an erythrocyte and the invading parasite. However, our studies rationalise the function of the most effective growth inhibitory antibodies targeting PfRH5. While these antibodies do not reduce the binding of PfRH5 to monomeric basigin, they do reduce its binding to basigin-PMCA and basigin-MCT complexes. This indicates that the most effective PfRH5-targeting antibodies inhibit growth by sterically blocking the essential interaction of PfRH5 with basigin in its physiological context.
Microprobe-XRF assessment of nutrient distribution in soybean, cowpea, and kidney bean seeds: a Fabaceae family case study
Authors: Montanha Gabriel Sgarbiero , Zachi Romeu Sara Luiza , Marques João Paulo Rodrigues , Rohr Lívia Araújo , de Almeida Eduardo , dos Reis André Rodrigues , Linhares Francisco Scaglia , Sabatini Sabrina , Pereira de Carvalho Hudson Wallace
Published: September 2022

The present study explored the microprobe X-ray fluorescence spectroscopy (µ-XRF) for quantitative and space-resolved distribution of macro, i.e., K, P, S, and Ca, and micronutrients, i.e., Fe, Zn, and Mn elemental composition in the cross-sectioned seeds of cowpea (Vigna unguiculata L.), kidney bean (Phaseolus vulgaris L.), and soybean (Glycine max L.) seeds, important agricultural species within the Fabaceae family. It unveils that both macro and micronutrients were heterogeneously distributed across seed tissues. Most of the P and S, Fe, Zn, and Mn were mainly found at the embryo axis tissues in all three Fabaceae species, whereas K was spread along the cotyledon and Ca was mostly observed trapped at the seed coat region. Furthermore, the Pearson correlation coefficient revealed a strong spatial correlation between P and S, and K and S in cowpea and soybean seed tissues, whereas Zn and Mn association was also recorded. Therefore, the µ-XRF technique demonstrates to be an important tool for assessing seed nutrient distribution, thus subsidizing the understanding of the physiological role of nutrients in seeds and fostering innovative approaches for nutrient supply and biofortification.
Tunable dynamics in a multi-strain transcriptional pulse generator
Authors: Zong David M. , Sadeghpour Mehdi , Molinari Sara , Alnahhas Razan N. , Hirning Andrew J. , Giannitsis Charilaos , Ott William , Josić Krešimir , Bennett Matthew R.
Published: September 2022

A major challenge in synthetic biology is the manipulation of engineered gene circuits toward a specified behavior. This challenge becomes more difficult as synthetic systems become more complex by incorporating additional genes or strains. Here we demonstrate that circuit dynamics can be tuned in synthetic consortia through the manipulation of strain fractions within the community. To do this, we constructed a microbial consortium comprised of three strains of engineered Escherichia coli that, when co-cultured, use homoserine lactone (HSL) mediated intercellular signaling to create a multi-strain incoherent type-1 feedforward loop (I1-FFL). Like naturally occurring I1-FFL motifs in gene networks, this engineered microbial consortium acts as a pulse generator of gene expression. We demonstrated that the amplitude of the pulse can be easily tuned by adjusting the relative population fractions of the strains. We created a mathematical model for the temporal dynamics of the microbial consortium and, using this model, identified population fractions that produced desired pulse characteristics. Our work demonstrates that intercellular gene circuits can be effectively tuned simply by adjusting the starting fractions of each strain type.
Enhancing Sox/Oct cooperativity induces higher-grade developmental reset
Authors: MacCarthy Caitlin M. , Malik Vikas , Wu Guangming , Velychko Taras , Keshet Gal , Jauch Ralf , Cojocaru Vlad , Schöler Hans R. , Velychko Sergiy
Published: September 2022

The discovery of induced pluripotent stem cell (iPSC) technology by Shinya Yamanaka has truly enabled the stem cell field. After 16 years of intense research, the delivery methods and culture media have improved but the original factors—Oct4, Sox2, Klf4, and Myc (OSKM)—remain central for driving reprogramming.

Here we define structural elements in chimeric Sox2/Sox17 transcription factors that rescued the ability of nonfunctional Oct factors to induce pluripotency. Most importantly, we discovered a single amino acid swap in the DNA-binding domain of Sox2, A61V, that stabilizes the Sox/Oct heterodimer on DNA through hydrophobic interaction with Oct. The highly cooperative Sox2AV mutant enables iPSC generation with Oct4 orthologs, such as Oct2 and Oct6, as well as rescues otherwise detrimental Oct4 mutants and domain deletions. Sox2AV has a dramatic effect on the cell fate reset, significantly improving the developmental potential of OSKM iPSCs. Moreover, by swapping multiple beneficial elements of Sox17 into Sox2 we have built a chimeric super-SOX factor—Sox2-17—that delivers unprecedented reprogramming efficiency and kinetics in five tested species. Sox2-17 enhances five-, four-, and three-factor reprogramming up to hundreds of times, enables two-factor generation of human iPSCs, and allows integration-free reprogramming of otherwise non-permissive aged human, non-human primate, and cattle fibroblasts.

Our study demonstrates that a complete developmental reset requires both robust activation of regulatory elements controlled by the canonical SoxOct motif and limiting cellular proliferation driven by Oct4 and Myc. A high level of Sox2 expression and Sox2/Oct4 heterodimerization emerge as the key determinants of high-grade pluripotency that fades along the naïve-to-primed continuum. Transient expression of SK cocktail can restore the naivety, providing a powerful technology to induce more complete developmental reset in pluripotent cells across species.
Nonsynonymous Mutations in Intellectual Disability and Autism Spectrum Disorder Gene PTCHD1 Disrupt N-Glycosylation and Reduce Protein Stability
Authors: Xie Connie T. Y. , Pastore Stephen F. , Vincent John B. , Frankland Paul W. , Hamel Paul A.
Published: September 2022

PTCHD1 has been implicated in Autism Spectrum Disorders (ASD) and/or intellectual disability, where copy number variant losses or loss-of-function coding mutations segregate with disease in an X-linked recessive fashion. Missense variants of PTCHD1 have also been reported in patients. However, the significance of these mutations remains undetermined since the activities, subcellular localization and regulation of the PTCHD1 protein are currently unknown. This paucity of data concerning PTCHD1 prevents the effective evaluation of sequence variants identified during diagnostic screening. Here, we characterize PTCHD1 protein binding partners, extending previously reported interactions with postsynaptic scaffolding protein, SAP102. Six rare missense variants of PTCHD1 were also identified from patients with neurodevelopmental disorders. After modelling these variants on a hypothetical three-dimensional structure of PTCHD1, based on the solved structure of NPC1, PTCHD1 variants harboring these mutations were assessed for protein stability, post-translational processing and protein trafficking. We show here that wild-type PTCHD1 post-translational modification includes complex N-glycosylation and that specific mutant proteins disrupt normal N-link glycosylation processing. However, regardless of their processing, these mutants still localized to PSD95-containing dendritic processes and remained competent for complexing SAP102.
Reprogramming of Iron Metabolism Confers Ferroptosis Resistance in ECM-Detached Cells
Authors: He Jianping , Abikoye Abigail M. , McLaughlin Brett P. , Middleton Ryan S. , Sheldon Ryan , Jones Russell G. , Schafer Zachary T.
Published: September 2022

Cancer cells often acquire resistance to cell death programs induced by loss of integrin-mediated attachment to extracellular matrix (ECM). Given that adaptation to ECM-detached conditions can facilitate tumor progression and metastasis, there is significant interest in effective elimination of ECM-detached cancer cells. Here, we find that ECM-detached cells are remarkably resistant to the induction of ferroptosis. While alterations in membrane lipid content are observed during ECM-detachment, it is instead fundamental changes in iron metabolism that underlie resistance of ECM-detached cells to ferroptosis. More specifically, our data demonstrate that levels of free iron are low during ECM-detachment due to changes in both iron uptake and iron storage. In addition, we establish that lowering the levels of iron storage proteins sensitizes ECM-detached cells to death by ferroptosis. Taken together, our data suggest that therapeutics designed to kill cancer cells by ferroptosis may be hindered by lack of efficacy towards ECM-detached cells.
Transcriptomics data integration for context-specific modeling of Atlantic salmon metabolism: functional evaluation of methods based on metabolic tasks
Authors: Molversmyr Håvard , Øyås Ove , Rotnes Filip , Vik Jon Olav
Published: September 2022

Motivation

Constraint-based models (CBMs) are used to study the metabolic networks of organisms ranging from microbes to multicellular eukaryotes. Published CBMs are usually generic rather than context-specific, meaning that they do not capture metabolic differences between cell types, tissues, environments, or other conditions. However, only a subset of reactions in a model are likely to be active in any given context, and several methods have therefore been developed to extract context-specific models from generic CBMs through integration of omics data.

Results

We tested the ability of six model extraction methods (MEMs) to create functionally accurate context-specific models of Atlantic salmon using a generic CBM (SALARECON) and liver transcriptomics data from contexts differing in water salinity (life stage) and dietary lipids. Reaction contents and metabolic task feasibility predictions of context-specific CBMs were mainly determined by the MEM that was used, but life stage explained significant variance in both contents and predictions for some MEMs. Three MEMs clearly outperformed the others in terms of their ability to capture context-specific metabolic activities inferred directly from the data, and one of these (GIMME) was much faster than the others. Context-specific versions of SALARECON consistently outperformed the generic version, showing that context-specific modeling captures more realistic representations of Atlantic salmon metabolism.

Contact

jon.vik@nmbu.no
Cell-type specific deletions of Neuroligin 2 reveal a vital role of synaptic excitation-inhibition balance
Authors: Longley Colleen M. , Xu Xize , Messier Jessica E. , Cai Zhao-Lin , Park Jung Woo , Chen Hongmei , Reznik Derek L. , Jadi Monika P. , Xue Mingshan
Published: September 2022

Synaptic excitation (E) and inhibition (I) stay relatively proportional to each other over different spatiotemporal scales, orchestrating neuronal activity in the brain. This proportionality, referred to as E-I balance, is thought to be critical for neuronal functions because its disruption was observed in many neurological disorders. However, the causal evidence demonstrating its significance is scarce. Here we show that deleting Neuroligin-2 (Nlgn2), a postsynaptic adhesion molecule at inhibitory synapses, from mouse glutamatergic or GABAergic neurons reduces inhibition cell-autonomously without affecting excitation, thereby disrupting E-I balance and causing lethality. In contrast, deleting Nlgn2 constitutively or simultaneously from both glutamatergic and GABAergic neurons results in viable mice. A neural network model shows that reducing inhibition in either neuronal type is detrimental to network activity, but in both types partially re-establishes E-I balance and activity. Together, our results provide evidence for an essential role of E-I balance in brain functions and organism survival.
A spatial atlas of Wnt and Frizzled receptor expression in adult mouse liver
Authors: Gayden Jenesis , Hu Shikai , Joseph Paul N. , Delgado Evan , Liu Silvia , Bell Aaron , Puig Stephanie , Monga Satdarshan P. , Freyberg Zachary
Published: September 2022

Hepatic zonation is critical for most metabolic functions in liver. Wnt signaling plays an important role in establishing and maintaining liver zonation. Yet, the anatomic expression of Wnt signaling components, including all 10 Frizzled receptors (Fzds), has not been characterized in adult liver. To address this, we quantitatively mapped the spatial expression of Wnt/Fzd pathway components in adult mouse liver via multiplex fluorescentin situhybridization. While all 10 Fzds are expressed within a metabolic unit, Fzds 1, 4, and 6 are the highest expressed. Though the majority of Wnt signaling occurs in zone 3, expression of most Fzds is not zonated. In contrast, Fzd6 is preferentially expressed in zone 1. We also discovered that Wnt2 and Wnt9b expression is highly zonated and primarily found in zone 3. Therefore, our results suggest that zonated Wnt expression is critical for zonation maintenance in healthy adult liver. Finally, we showed that Fzds and Wnts are not uniformly expressed by all hepatic cell types. Rather, there is broad distribution among both hepatocytes and non-parenchymal cells, including endothelial cells. Overall, our establishment of a definitive mRNA expression atlas of Wnt/Fzd pathway components opens the door to future functional characterization in healthy and disease states.
Compensatory Relationship between Low Complexity Regions and Gene Paralogy in the Evolution of Prokaryotes
Authors: Persi Erez , Wolf Yuri I. , Karamycheva Svetlana , Makarova Kira S , Koonin Eugene V.
Published: September 2022

Evolution of genomes in all life forms involves two distinct, dynamic types of genomic changes: gene duplication (and loss) that shape families of paralogous genes and extension (and contraction) of low complexity regions (LCR), which occurs through dynamics of short repeats in protein-coding genes. Although the roles of each of these types of events in genome evolution have been studied, their co-evolutionary dynamics is not thoroughly understood. Here, by analyzing a wide range of genomes from diverse bacteria and archaea, we show that LCR and paralogy represent two distinct routes of evolution that are inversely correlated. Emergence of LCR is a prominent evolutionary mechanism in fast evolving, young protein families, whereas paralogy dominates the comparatively slow evolution of old protein families. Analysis of multiple prokaryotic genomes shows that the formation of LCR is likely a widespread, transient evolutionary mechanism that temporally and locally affects also ancestral functions, but apparently, fades away with time, under mutational and selective pressures, yielding to gene paralogy. We propose that compensatory relationships between short-term and longer-term evolutionary mechanisms are universal in the evolution of life.

Significance

Evolution of genomes in all organisms involves a variety of changes occurring on different spatial and temporal scales, from point mutations to whole genome duplication. Here we demonstrate that during the evolution of bacterial and archaeal genomes, there is a universal inverse relationship between the formation of low complexity regions in protein sequences through proliferation of short repeats and gene duplication. The former process apparently is a route of short-term adaptation whereas the latter one dominates evolution on longer temporal scales. We propose that compensatory relationships between evolutionary mechanisms acting at different spatial and temporal scales are a general feature of the process of evolution.
Decoding a cryptic mechanism of metronidazole resistance among globally disseminated fluoroquinolone-resistant Clostridioides difficile
Authors: Olaitan Abiola O. , Dureja Chetna , Youngblom Madison A. , Topf Madeline A. , Shen Wan-Jou , Gonzales-Luna Anne J. , Deshpande Aditi , Hevener Kirk E. , Freeman Jane , Wilcox Mark H. , Palmer Kelli L. , Garey Kevin W. , Pepperell Caitlin S. , Hurdle Julian G.
Published: September 2022

Severe outbreaks and deaths have been linked to the emergence and global spread of fluoroquinolone-resistant Clostridioides difficile over the past two decades. At the same time, metronidazole, a nitro-containing antibiotic, has shown decreasing clinical efficacy in treating C. difficile infection (CDI). Most metronidazole-resistant C. difficile exhibit an unusual resistance phenotype that can only be detected in susceptibility tests utilizing molecularly intact heme. Here we describe the mechanism underlying this trait, which we discovered using molecular genetics, phylogenetics, and population analyses. Most metronidazole-resistant strains evolved a T to G mutation, we term PnimBG, in the -10 regulatory promoter of the 5-nitroimidazole reductase nimB, resulting in the gene being constitutively transcribed. Silencing or deleting nimB eliminated metronidazole resistance. We identified the protein as a heme-dependent nitroreductase that degraded nitro-drugs to an amine lacking antimicrobial activity. We further discovered that the metronidazole-resistant PnimBGmutation was strongly associated with the Thr82Ile substitution conferring fluoroquinolone resistance in epidemic strains. Re-analysis of published genomes from global isolates confirmed that all but one encoding PnimBG also carried the Thr82Ile mutation. Our findings suggest that fluoroquinolone and metronidazole resistance co-mediated the pandemic of healthcare-associated C. difficile that are associated with poorer treatment outcomes in CDI patients receiving metronidazole.
The ancestral ESCRT protein TOM1L2 selects ubiquitinated cargoes for retrieval from cilia
Authors: Shinde Swapnil Rohidas , Mick David U. , Aoki Erika , Rodrigues Rachel B. , Gygi Steven P. , Nachury Maxence V.
Published: September 2022

Many G protein-coupled receptors (GPCRs) reside within cilia of mammalian cells and must undergo regulated exit from cilia for the appropriate transduction of signals such as Hedgehog morphogens. Lysine 63-linked ubiquitin (K63Ub) chains mark GPCRs for regulated removal from cilia, but the molecular basis of K63Ub recognition inside cilia remains elusive. Here we show that the BBSome –the trafficking complex in charge of retrieving GPCRs from cilia– engages the ancestral endosomal sorting factor TOM1L2 (Target of Myb1-Like 2) to recognize UbK63 chains within cilia. TOM1L2 directly binds to UbK63 chains and to the BBSome and targeted disruption of the TOM1L2/BBSome interaction results in the accumulation of TOM1L2, ubiquitin and the GPCRs SSTR3, Smoothened and GPR161 inside cilia. Strikingly, the single cell algaChlamydomonasalso requires its TOM1L2 orthologue to clear ubiquitinated proteins from cilia. We conclude that TOM1L2 broadly enables the retrieval of UbK63-tagged proteins by the ciliary trafficking machinery.
Human olfactory neuronal cells through nasal biopsy: molecular characterization and utility in brain science
Authors: Yang Kun , Ishizuka Koko , Lane Andrew P. , Narita Zui , Hayashida Arisa , Lema Yukiko Y. , Heffron Emma , Loudd Haydn , Schumacher Maeve , Kano Shin-Ichi , Tomoda Toshifumi , Kamiya Atsushi , Ma Minghong , Geman Donald , Younes Laurent , Sawa Akira
Published: September 2022

Biopsy is crucial in clinical medicine to obtain tissues and cells that directly reflect the pathological changes of each disease. However, the brain is an exception due to ethical and practical challenges. Nasal biopsy, which captures the olfactory neuronal epithelium, has been considered as an alternative method of obtaining neuronal cells from living patients. Multiple groups have enriched olfactory neuronal cells (ONCs) from biopsied nasal tissue. ONCs can be obtained from repeated biopsies in a longitudinal study, providing mechanistic insight associated with dynamic changes along the disease trajectory and treatment response. Nevertheless, molecular characterization of biopsied nasal cells/tissue has been insufficient. Taking advantage of recent advances in next-generation sequencing technologies at the single-cell resolution and related rich public databases, we aimed to define the neuronal characteristics, homogeneity, and utility of ONCs. We applied single-cell and bulk RNA sequencing for ONCs, analyzing and comparing the data with multiple public datasets. We observed that the molecular signatures of ONCs are similar to those of neurons, distinct from major glial cells. The signatures of ONCs resemble those of developing neurons and share features of excitatory neurons in the prefrontal and cingulate cortex. The high homogeneity of ONCs is advantageous in pharmacological, functional, and protein studies. Accordingly, we provide two proof-of-concept examples for functional and protein studies, solidifying the utility of ONCs in studying objective biomarkers and molecular mechanisms for brain disorders. The ONCs may also be useful in the studies for the olfactory epithelium impairment and the resultant mental dysfunction elicited by SARS-CoV-2.

SIGNIFICANCE STATEMENT

To study dynamic changes and underlying mechanisms along disease trajectory and treatment response in neuropsychiatric disorders, olfactory neuronal cells (ONCs) enriched from biopsied nasal tissue may provide a crucial tool. Because ONCs can be obtained from repeated biopsies in a longitudinal study, this tool has been believed to be useful and complementary to postmortem brains and induced pluripotent stem cell-derived neurons. Nevertheless, molecular characterization of biopsied nasal cells/tissue has been insufficient, which hampers a broader use of this resource. Taking advantage of recent advances in next-generation sequencing technologies at the single-cell resolution and related rich public databases, the present study defines ONCs’ neuronal characteristics, homogeneity, and unique utility for the first time.
Molecular characters and phylogenetic analysis of Clostridium perfringens from different regions in China, from 2013 to 2021
Authors: Zhong Jia xin , Zheng Hao ran , Wang Yuan yuan , Bai Lu lu , Du Xiao li , Wu Yuan , Lu Jin xing
Published: September 2022

Clostridium perfringens (C. perfringens) is a significant foodborne pathogen and a common cause of intestinal diseases in both animals and humans. Altogether, 186 isolates were obtained from humans (n = 147), animals (n = 25), and food (n = 14), comprising 174 type A strains (93.55%), 11 type F strains (5.91%), and one type D strain (0.54%); and were analyzed by multilocus sequence typing (MLST) and antimicrobial susceptibility testing. Additionally, some specific ST complexes were analyzed by cgMLST and cgSNP to investigate genetic relatedness. MLST indicated the most prevalent STs of C. perfringens of human and animal origin were as follows: ST221 (5/147), ST62 (4/147), ST408 (4/147), and ST493 (4/147) were predominant in humans, while ST479 (5/25) was the major type in animals. Within the same ST complex, genetically unrelated relationships or potential clustering/transmission events were further recognized by cgMLST and cgSNP, illustrating that these two methods are valuable in defining outbreaks and transmission events. All tested isolates were susceptible to vancomycin and meropenem. The rates of resistance to metronidazole, penicillin, cefoxitin, moxifloxacin, and clindamycin were low (metronidazole: 1.08%; penicillin: 9.68%; cefoxitin: 0.54%; moxifloxacin: 6.45%; and chloramphenicol: 3.76%). Interestingly, 49.66% of human origin were clindamycin-resistant, and 18.2% were penicillin-insensitive. Importantly, the portion of MDR isolates was significantly lower than in previous reports. The study provides an overview of the epidemiological characteristics of C. perfringens with different origins and hosts in China. C. perfringens demonstrated remarkable genetic diversity and distinct molecular features compared to antibiotic-resistance profiles from other studies.

IMPORTANCE

C. perfringens is one of the most common bacterial causes of foodborne illness globally, responsible for several food-poisoning outbreaks each year. This study provides an overview of C. perfringens isolates from different hosts and regions in China according to MLST, antibiotic-resistance characters, cgMLST, and cgSNP analyses, showing high genetic diversity and identifying potential clustering and transmission events. The antimicrobial profile in this study was distinct from that of a previous report with a much lower MDR rate, indicating that C. perfringens in China needs further investigation.
Compounds derived from N,N-dimethyldithiocarbamate are effective copper-dependent antimicrobials against Streptococcus pneumoniae
Authors: Menghani Sanjay V. , Sanchez-Rosario Yamil , Pok Chansorena , Liu Renshuai , Gao Feng , O’Brien Henrik , Neubert Miranda J. , Ochoa Klariza , Hellinger Riley D. , Wang Wei , Johnson Michael D. L.
Published: September 2022

N,N-dimethyldithiocarbamate (DMDC) is a potent copper-dependent antimicrobial against several pathogens, including Streptococcus pneumoniae. Despite the availability of several vaccines against multiple disease-causing strains of S. pneumoniae, the rise of antimicrobial resistance and pneumococcal disease caused by strains not covered by the vaccine creates a need for developing novel antimicrobial strategies. We derived novel compounds from DMDC and tested their effectiveness as copper-dependent antimicrobials against S. pneumoniae through in vitro growth and killing curves. Compounds that caused a growth defect and were bactericidal in vitro were tested against other strains of S. pneumoniae and in complex with different transition metals. We found two compounds, sodium N-benzyl-N-methyldithiocarbamate and sodium N-allyl-N-methyldithiocarbamate (herein “Compound 3” and “Compound 4”), were effective against TIGR4, D39, and ATCC® 6303™ (a type 3 capsular strain) and further increased the internal concentrations of copper to the same previously reported levels as with DMDC and copper treatment. We found that both Compound 3 and Compound 4 were bacteriostatic in combination with zinc. We tested Compound 3 and Compound 4 in vivo against a murine pneumonia model, finding that Compound 3, and not Compound 4, was effective in significantly decreasing the bacterial burden in the blood and lungs of S. pneumoniae-infected mice. We found that the combination of Compound 3 and copper made the pneumococcus more susceptible to activated macrophage mediated killing via an in vitro macrophage killing assay. Collectively, we demonstrate that derivatizing DMDC holds promise as potent bactericidal antibiotics against S. pneumoniae.
Reduced ribosome activity influences the non-uniform evolution of 16S rRNA hypervariable regions
Authors: Bose Nikhil , Moore Sean D.
Published: September 2022

16S rRNA gene sequences are commonly analyzed for taxonomic and phylogenetic studies because they contain hypervariable regions that can help distinguish different genera. However, intra-genus distinction is often difficult due to high sequence identities among closely related species. Although common tools for 16S sequence taxonomic classification weight residue variations equally during comparisons, specific residues within hypervariable regions have not drifted evenly through evolution, suggesting that portions of them may be biologically important. We developed an in vivo test system where 16S variants coexisted among natural ribosome populations which allowed their fitness to be evaluated. We found that versions with evolutionarily disparate hypervariable regions were underpopulated in ribosomes and active translation pools, even for a single nucleotide polymorphism (SNP), which indicates functional constraints to the free evolutionary drift of hypervariable regions. Using an in silico method (positional relative entropy), we analyzed over 12,000 16S V3-V4 sequences across Escherichia and Shigella strains and identified species that can be distinguished by position-specific SNPs present in multiple 16S alleles in a genome. When we evaluated these informative SNPs with our in vivo system, we discovered that ribosomes harboring them were compromised, suggesting that their evolution is indeed biologically constrained. Overall, this study demonstrates that SNPs within hypervariable regions are not necessarily inconsequential and that common computational approaches for taxonomic 16S rRNA sequence classification should not assume an even probability of residues at each position.

Importance

Hypervariable regions within 16S rRNA genes are commonly analyzed to determine microbial diversity. However, because sequences within a genus are highly similar, strain- or species-specific identification is often uncertain. Because there are no established functions of hypervariable regions, residue variations within them are often evenly weighted when making taxonomic comparisons. This study established that 16S rRNAs with naturally occurring variations in hypervariable regions can affect ribosome quality, indicating that their residues should not be weighted equally during taxonomic sequence classifications.
Continuous directed evolution of a feedback-resistant Arabidopsis arogenate dehydratase in plantized E. coli
Authors: Leong Bryan J. , Hanson Andrew D.
Published: September 2022

Continuous directed evolution (CDE) is a powerful tool for enzyme engineering due to the depth and scale of evolutionary search that it enables. If suitably controlled and calibrated, CDE could be widely applied in plant breeding and biotechnology to improve plant enzymes ex planta. We tested this concept by evolving Arabidopsis arogenate dehydratase (AtADT2) for resistance to feedback inhibition. We used an Escherichia coli platform with a phenylalanine biosynthesis pathway reconfigured (‘plantized’) to mimic the plant pathway, a T7RNA polymerase-base deaminase hypermutation system (eMutaT7), and 4-fluorophenylalanine as selective agent. Selection schemes were pre-validated using a known feedback-resistant AtADT2 variant. We obtained variants that had 4-fluorophenylalanine resistance at least matching the known variant and that carried mutations in the ACT domain responsible for feedback inhibition. We conclude that ex planta CDE of plant enzymes in a microbial platform is a viable way to tailor characteristics that involve interaction with small molecules.
Global mosquito virome profiling and mosquito spatial diffusion pathways revealed by marker-viruses
Authors: Zhao Lu , Yu Ping , Shi Chenyan , Jia Lijia , Evans Atoni , Wang Xiaoyu , Wu Qun , Xiong Guodian , Ming Zhaoyan , Salazar Ferdinand , Agwanda Bernard , Bente Dennis , Wang Fei , Liu Di , Yuan Zhiming , Xia Han
Published: September 2022

Mosquitoes are vectors of numerous emergence and reemergence of mosquito-borne diseases, leading to an overwhelming global challenge. The boom in metagenomic studies promoted the increase of mosquito viruses being described, and studies from different regions of the world showed that mosquitoes harbor abundant and diverse viromes. However, there is still a lack of large-scale systematic comparison of viromes among various ecological factors such as the mosquito species/genera, location, etc, and consistent patterns associated with these factors are not clear. Here, we provide an overview of virome profiling that integrated the perspective of mosquito genus, locations, and climates based on the global scale, and redefined the ‘core-virome’. Our results also further implicate some mosquito-associated viruses strongly associated with the ecological factors and highlighted the evidence of mosquitoes’ cross-regional movements by the candidate marker viruses. The study may be helpful in gaining new insights into strategies to prevent arbovirus epidemics.
Ribosomal RNA 2’-O-methylation dynamics impact cell fate decisions
Authors: Häfner SJ , Jansson MD , Altinel K , Kraushar ML , Andersen KL , Abay-Nørgaard Z Caldwell , Fontenas M , Sørensen DM , Gay DM , Arendrup FS , Tehler D , Krogh N , Nielsen H , Kirkeby A , Lund AH
Published: September 2022

Translational regulation impacts both pluripotency maintenance and cell differentiation. To what degree the ribosome itself exerts control over this process remains unanswered. Accumulating evidence has demonstrated heterogeneity in ribosome composition in various organisms. 2’-O-methylation of rRNA represents an important source of heterogeneity, where site-specific alteration of methylation levels can modulate translation. Here we explore changes in rRNA 2’-O-methylation during mouse brain development and during tri-lineage differentiation of human embryonic stem cells. We find distinct alterations between brain regions, as well as clear dynamics during cortex development and germ layer differentiation. We identify a methylation site which impacts neuronal differentiation. Modulation of its methylation levels affects ribosome association of the Fragile X Mental Retardation Protein and translation of WNT pathway-related mRNAs. Together, the data reveals ribosome heterogeneity through rRNA 2’-O-methylation during early development and differentiation and suggests a direct role for ribosomes in regulating translation during cell fate acquisition.
The role of the γ subunit in the photosystem of the lowest-energy phototrophs
Authors: Namoon Dowrung , Rudling Nicola M , Canniffe Daniel P
Published: September 2022

Purple phototrophic bacteria use a core ‘photosystem’ consisting of light harvesting antenna complex 1 (LH1) surrounding the reaction centre (RC), which primarily absorbs far-red–near-infrared light and converts it to chemical energy. Species in the Blastochloris genus, which are able to use light >1000nm for photosynthesis, use bacteriochlorophyll (BChl) b rather than the more common BChl a as their major photopigment, and also uniquely assemble LH1 with an additional polypeptide subunit, LH1γ, encoded by multiple open reading frames in their genomes. In order to assign a role to this subunit, we deleted the four LH1γ-encoding genes in the model Blastochloris viridis. Interestingly, growth under halogen bulbs routinely used for cultivation of anoxygenic phototrophs yielded cells displaying an absorption maximum of 825 nm, similar to that of the RC complex without LH1, but growth under white light from fluorescent bulbs yielded cells with an absorption maximum at 972 nm. HPLC analysis of pigment composition and sucrose density gradient fractionation demonstrate that the mutant grown in white light assembles RC–LH1, albeit with an absorption maximum blue-shifted by 46 nm relative to the WT complex. Wavelengths between 900–1000 nm transmit poorly through the atmosphere due to strong absorption by water, thus our results provide an evolutionary rationale for the incorporation of the γ subunit into the LH1 ring; this polypeptide red-shifts the absorption maximum of the complex to a range of the spectrum where the photons are of lower energy but are more abundant. Finally, we transformed the mutant with plasmids carrying genes encoding natural LH1γ variants and demonstrate that the polypeptide found in the WT complex red-shifts absorption back to 1018 nm, but incorporation of a distantly-related variant results in only a moderate red-shift. This result suggests that tuning the absorption maximum of this organism is possible, and may permit light capture past the current low-energy limit of natural photosynthesis.
An approach towards identifying the site of interaction between the Streptomyces phage ØC31 protein Gp3 and the coiled-coil domain of its serine integrase
Authors: Halablab Monalissa , Colloms Sean , Kane Steven
Published: September 2022

Streptomyces phage ØC31 encodes a serine integrase which recombines the host (attB) and phage (attP) attachment sites to form new attL and attR sites. For attL and attR to be recombined by the integrase, the presence of its cognate recombination directionality factor (RDF) is required. It is hypothesized that the RDF binds the coiled-coil domain of the integrase to accomplish this switch in recombination directionality. Yet, nothing is known about the site of interaction between the RDF and integrase. Here, we tried to identify the region on the coiled-coil domain of ØC31 integrase to which its cognate RDF (gp3) might bind. Mutant integrases fused to their cognate RDF (gp3) were created and selected for inactivity in attL x attR recombination. It was not possible to characterize the mutants and identify the Int-RDF binding region, due to unforeseen errors that occurred during the construction of the mutant library, but we demonstrated that simple experimental approaches could be used to identify this region. Despite this, an integrase mutant (P398L D595N) fused to gp3 was characterized. This mutant was catalyzing a more unidirectional attL x attR recombination reaction with reduced attP x attB recombination compared to the wild-type integrase. This mutant was selected since it could be used within the field of molecular biology to construct inversion switches which are a key element by which cells can be computerized.

By the time this work was uploaded on bioRxiv, there is now a publication that investigated the Int-RDF interaction interface and several residues at the base of the coiled-coil that affected both the interaction and recombinase activity were identified (Paul C M Fogg, Ellen Younger, Booshini D Fernando, Thanafez Khaleel, W Marshall Stark, Margaret C M Smith, Recombination directionality factor gp3 binds PhiC31 integrase via the zinc domain, potentially affecting the trajectory of the coiled-coil motif, Nucleic Acids Research, Volume 46, Issue 3, 16 February 2018, Pages 1308-1320, https://doi.org/10.1093/nar/gkx1233).
GTP as an enzymatic synchronizer
Authors: Guo Chen-Jun , Zhou Xian , Liu Ji-Long
Published: September 2022

Synchronizer is the key component of modern automobile gearbox, which coordinates the deceleration and acceleration of two groups of gears. In industry, synchronizer is composed of several components with fine structure. Here, we show that guanosine triphosphate (GTP) has the property of enzyme synchronizer. Through cryo-electron microscopy (cryo-EM) and 3D variability analysis, we obtain the full spectrum conformation of CTP synthase at near atomic resolution. In the catalytic process of CTP synthase, we find that GTP synchronizes the reaction of two independent domains, namely, glutamine-dependent aminotransferase (GAT) domain and amidoligase (AL) domain. The wing structure acts as a striker, and then stimulates the intermediate ammonia to complete the catalytic cycle through the internal gas channel. Different from being degraded to regulate tubulins or G-proteins, GTP acts as an enzymatic synchronizer to coordinate the reactions catalyzed by the two independent domains of CTP synthase.
Novel Biosensor Identifies Ruxolitinib as a Potent and Cardioprotective CaMKII Inhibitor
Authors: Gaido Oscar E. Reyes , Granger Jonathan M. , Nkashama Lubika J. , Lin Brian L. , Long Alan , Mesubi Olurotimi O. , Schole Kate L. , Terrilion Chantelle E. , Liu Jun O. , Luczak Elizabeth D. , Anderson Mark E.
Published: September 2022

Ca2+/Calmodulin-dependent protein kinase II (CaMKII) hyperactivity causes heart injury and arrhythmias—two major sources of mortality worldwide. Despite proven benefits of CaMKII inhibition in numerous preclinical models of heart disease, translation of CaMKII antagonists into humans has been stymied by low potency, toxicity, and an enduring concern for adverse effects on cognition due to an established role of CaMKII in learning and memory. To address these challenges, we asked if any clinically approved drugs, developed for other purposes, were potent CaMKII inhibitors. For this, we engineered a novel fluorescent biosensor, CaMKAR (CaMKII Activity Reporter), which features superior sensitivity, kinetics, and tractability for high throughput screening. Using this tool, we carried a drug repurposing screen (4,475 compounds in clinical use) in human cells expressing autonomously active CaMKII. This yielded five previously unrecognized CaMKII inhibitors with clinically relevant potency: ruxolitinib, baricitinib, silmitasertib, crenolanib, and abemaciclib. Standout among these, ruxolitinib, an orally bioavailable and U.S Food and Drug Administration (FDA)-approved medication, inhibited CaMKII in cultured cardiomyocytes and in mice at concentrations equivalent to human doses. 10-minute treatment in mice was sufficient to prevent atrial fibrillation— the most common clinical arrhythmia. At cardioprotective doses, ruxolitinib-treated mice behaved normally in established cognitive assays. Our results suggest that human CaMKII inhibition is feasible and safe, and support prompt clinical investigation of ruxolitinib for cardiac indications.

One Sentence Summary

We developed a CaMKII biosensor suitable for high throughput screening and identified ruxolitinib as a CaMKII inhibitor capable of rescuing cardiac arrhythmia.
Plant litter chemistry controls coarse‐textured soil carbon dynamics
Authors: Huys Raoul , Poirier Vincent , Bourget Malo Y. , Roumet Catherine , Hättenschwiler Stephan , Fromin Nathalie , Munson Alison D. , Freschet Grégoire T.
Published: September 2022

As soils store more carbon (C) than the Earth's atmosphere and terrestrial biomass together, the balance between soil C uptake in the form of soil organic matter (SOC) and release as CO2 upon its decomposition is a critical determinant in the global C cycle regulating our planet's climate. Although plant litter is the predominant source of C fuelling both soil C build‐up and losses, the issue of how litter chemistry influences this balance remains unresolved.

As a contribution to solving that issue, we traced the fate of C during near‐complete decomposition of 13C‐labelled leaf and root litters from 12 plant species in a coarse‐textured soil. We separated the soil organic carbon into mineral‐associated organic matter (MAOM) and particulate organic matter (POM) pools, and investigated how 14 litter chemical traits affected novel SOC formation and native SOC mineralization (i.e. the priming effect) in these soil fractions.

We observed an overall net increase in SOC due to the addition of litter, which was stronger for root than for leaf litters. The presumed stable MAOM‐C pool underwent both substantial stabilization and mineralization, whereas the presumably less stable POM‐C pool showed substantial stabilization and reduced mineralization. Overall, the initial increase in soil C mineralization was fully counterbalanced by a later decrease in native soil C mineralization. POM‐C formation as well as MAOM‐C formation and mineralization were positively related to the initial litter lignin concentration and negatively to that of the nitrogen leachates, whereas the opposite was observed for POM‐C mineralization.

Synthesis. Our results highlight the importance of litter chemical traits for SOC formation, and stabilization, destabilization and mineralization. In our coarse‐textured soil, the amount of MAOM‐C did not change despite large C fluxes through this pool. The litter chemical traits that drove these processes differed from those frequently reported for fine‐textured soils far from mineral‐associated C saturation. To account for these discrepancies, we propose an integrative perspective in which litter quality and soil texture interactively control soil C fluxes by modulating several SOC stabilization and destabilization mechanisms. Irrespective, our results open new critical perspectives for managing soil C pools globally.
Salvianolic acid B, a new type I IRE1 kinase inhibitor, abrogates AngII‐induced angiogenesis by interacting with IRE1 in its active conformation
Authors: Fan Fangtian , Liu Fang , Shen Peiliang , Tao Li , Zhang Hongjiang , Wu Hongyan
Published: September 2022

Angiotensin II (AngII)‐mediated pathological angiogenesis is one of the important factors promoting the progression of atherosclerosis, tumour metastasis, and diabetic retinopathy. Here, we first demonstrate that salvianolic acid B (Sal B) attenuated AngII‐induced angiogenesis by downregulating the IRE1/ASK1/JNK/p38MAPK signalling pathway and protected vascular endothelial cells from hypoxia‐induced damage. These pharmacological consequences could be ascribed to the unique interactions between Sal B and the ATP‐binding cavity of IREIα, leading to bi‐directional roles of IRE1 kinase and endonuclease activity; this may possibly be one of the essential mechanisms of the bi‐directional regulation of angiogenesis in different conditions. Moreover, our results indicated that IRE1 was a novel anti‐angiogenesis target and type I IRE1 kinase inhibitor (e.g., Sal B, APY29) and might be a potentially eligible low‐toxicity drug for treating AngII‐mediated pathological angiogenesis.
Individual socioeconomic status and risk of out‐of‐hospital cardiac arrest: A nationwide case–control analysis
Authors: Lee Sun Young , Park Jeong Ho , Choi Young Ho , Lee Jungah , Ro Young Sun , Hong Ki Jeong , Song Kyoung Jun , Shin Sang Do
Published: September 2022

Objectives

Area‐level socioeconomic status (SES) is associated with the incidence of out‐of‐hospital cardiac arrest (OHCA); however, the effects of individual‐level SES on OHCA occurrence are unknown. This study investigated whether individual‐level SES is associated with the occurrence of OHCA.

Methods

This case–control study used data from the nationwide OHCA registry and the National Health Information Database (NHID) in Korea. All adult patients with OHCA of a medical etiology from 2013 to 2018 were included. Four controls were matched to each OHCA patient based on age and sex. The exposure was individual‐level SES measured by insurance type and premium, which is based on income in Korea. National Health Insurance (NHI) beneficiaries were divided into four groups (Q1–Q4), and medical aid beneficiaries were separately classified as the lowest SES group. The adjusted odds ratios (aORs) and 95% confidence intervals (CIs) for the outcomes were calculated. Stratified analyses were conducted according to age and sex.

Results

A total of 105,443 cases were matched with 421,772 controls. OHCA occurred more frequently in the lower SES groups. Compared with the highest SES group (Q1), the aORs for OHCA occurrence increased as the SES decreased (aORs [95% CI] were 1.21 [1.19–1.24] for Q2, 1.33 [1.31–1.36] for Q3, 1.32 [1.30–1.35] for Q4, and 2.08 [2.02–2.13] for medical aid). Disparity by individual‐level SES appeared to be greater in males than in females and greater in the young and middle‐aged adults than in older adults.

Conclusions

Low individual‐level SES was associated with a higher probability of OHCA occurrence. Efforts are needed to reduce SES disparities in the occurrence of OHCA.