bluuebunny/crossref_metadata_2025_split huggingface dataset:
Assembly and architecture of the type III secretion sorting platform
Authors: Soto J. Eduardo , Galán Jorge E. , Lara-Tejero María 
Published: September 2022

SUMMARYType III secretion systems are bacterial nanomachines specialized in protein delivery into target eukaryotic cells. The structural and functional complexity of these machines demand highly coordinated mechanisms for their assembly and operation. The sorting platform is a critical component of type III secretion machines that ensures the timely engagement and secretion of proteins destined to travel this export pathway. However, the mechanisms that lead to the assembly of this multi-component structure have not been elucidated. Herein, employing structure modeling and an extensive in vivo crosslinking strategy, we provide a detailed inter-subunit-contact survey of the entire sorting platform complex. Using the identified crosslinks as signatures for pairwise inter-subunit interactions in combination with systematic genetic deletions, we mapped the assembly process of this unique bacterial structure. Insights generated by this study could serve as the bases for the development of anti-virulence strategies to combat several medically important bacterial pathogens.
Metallothionein-3 attenuates the effect of Cu2+ ions on actin filaments
Authors: Lakha Rabina , Hachicho Carla , Mehlenbacher Matthew R. , Wilcox Dean E. , Austin Rachel N. , Vizcarra Christina L. 
Published: September 2022

AbstractMetallothionein 3 (MT-3) is a cysteine-rich metal-binding protein that is expressed in the mammalian central nervous system and kidney. Various reports have posited a role for MT-3 in regulating the actin cytoskeleton by promoting the assembly of actin filaments. We generated purified, recombinant mouse MT-3 of known metal compositions, either with zinc (Zn), lead (Pb), or copper/zinc (Cu/Zn) bound. None of these forms of MT-3 accelerated actin filament polymerization in vitro, either with or without the actin binding protein profilin. Furthermore, using a co-sedimentation assay, we did not observe Zn-bound MT-3 in complex with actin filaments. Cu2+ ions on their own induced rapid actin polymerization, an effect that we attribute to filament fragmentation. This effect of Cu2+ is reversed by adding either EGTA or Zn-bound MT-3, indicating that either molecule can chelate Cu2+ from actin. Altogether, our data indicate that recombinant MT-3 does not directly bind actin but it does attenuate the Cu-induced fragmentation of actin filaments.
Estradiol (E2) concentration shapes the chromatin binding landscape of the estrogen receptor
Authors: Han Amy L. , Vinod-Paul Kiran , Rao Satyanarayan , Brechbuhl Heather M. , Sartorius Carol A. , Ramachandran Srinivas , Kabos Peter 
Published: September 2022

AbstractHow transcription factors (TF) selectively occupy a minute subset of their binding sites from a sizeable pool of putative sites in large mammalian genomes remains an important unanswered question. In part, nucleosomes help by creating formidable barriers to TF binding. TF concentration itself plays a crucial role in the competition between TFs and nucleosomes. In the case of nuclear receptors, the ligand adds another layer of complexity. Estrogen receptor alpha (ER) is a classic example where its main ligand estradiol (E2) can modulate ER binding on chromatin. Here we show a complete rewiring of ER binding as a function of E2 concentration. As E2 concentration increases by two orders of magnitude, ER levels decrease, and ER binding localizes to promoter-distal sites with strong ER motifs. At low E2 levels, abundant levels of ER are present in the nucleus, and ER binding occurs mostly at sites without an identifiable ER binding motif, potentially in cooperation with other TFs like STAT1. We propose that E2’s effect on ER activity plays a major role in defining genome-wide ER binding profiles. Thus, variations in E2 concentrations in ER-positive breast tumors could be a significant factor driving heterogeneity in tumor phenotype, treatment response, and potentially drug resistance.
Reevaluation of Piezo1 as a gut RNA sensor
Authors: Nickolls AR , O’Brien GS , Shnayder S , Zhang Y , Nagel M , Patapoutian A , Chesler AT 
Published: September 2022

ABSTRACTPiezo1 is a stretch-gated ion channel required for mechanosensation in many organ systems. Recent provocative findings describe a new role for Piezo1 in the gut, suggesting that it is a sensor of microbial single-stranded RNA (ssRNA) rather than mechanical force. If true, this would redefine the scope of Piezo biology. Here, we sought to replicate the central finding that fecal ssRNA is a natural agonist of Piezo1. While we observed that fecal extracts and ssRNA stimulate calcium influx in certain cell lines, this response was independent of Piezo1. Additionally, sterilized dietary extracts devoid of gut biome RNA showed similar cell line-specific stimulatory activity to fecal extracts. Together, our data highlight potential confounds inherent to gut-derived extracts, exclude Piezo1 as a receptor for ssRNA in the gut, and support a dedicated role for Piezo channels in mechanosensing.
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

AbstractBasigin 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

ABSTRACTThe 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

AbstractA 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

ABSTRACTThe 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

AbstractPTCHD1 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

SummaryCancer 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

AbstractMotivationConstraint-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.ResultsWe 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.Contactjon.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

AbstractSynaptic 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

AbstractHepatic 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

AbstractEvolution 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.SignificanceEvolution 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

AbstractSevere 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.