bluuebunny/crossref_metadata_2025_split huggingface dataset:
Alternative catalytic residues in the active site of Esco acetyltransferases
Authors: Ajam Tahereh , De Inessa , Petkau Nikolai , Whelan Gabriela , Pena Vladimir , Eichele Gregor 
Published: September 2019

AbstractCohesin is a protein complex encircles the DNA and regulates the separation of sister chromatids during cell division. Following a catalytic mechanism that is insufficiently understood, Esco1 and Esco2 acetyltransferases acetylate Smc3 subunit of cohesin, thereby inducing a stabilization of cohesin on DNA. As a prerequisite for structure-guided investigation of enzymatic activity, we determine here the crystal structure of the mouse Esco2/CoA complex at 1.8 Å resolution. We reconstitute the entire cohesin as a tetrameric assembly and use it as a physiologically-relevant substrate for enzymatic assays in vitro. Furthermore, we employ cell-based complementation studies in mouse embryonic fibroblast deficient for Esco1 and Esco2, as a means to identify catalytically-important residues in vivo. These analyses demonstrate that D567/S566 and E491/S527, located on opposite sides of the MmEsco2 active site cleft, are critical for catalysis. Our experiments supports a catalytic mechanism of acetylation where residues D567 and E491 are general bases that deprotonate the ε-amino group of lysine substrate, via two nearby serine residues - S566 and S527-that possess a proton relay function.
Deletion of a non-canonical regulatory sequence causes loss of Scn1a expression and epileptic phenotypes in mice
Authors: Haigh Jessica L. , Adhikari Anna , Copping Nycole A. , Stradleigh Tyler , Wade A. Ayanna , Catta-Preta Rinaldo , Su-Feher Linda , Zdilar Iva , Morse Sarah , Fenton Timothy A , Nguyen Anh , Quintero Diana , Agezew Samrawit , Sramek Michael , Kreun Ellie J , Carter Jasmine , Gompers Andrea , Lambert Jason , Canales Cesar P. , Pennacchio Len A. , Visel Axel , Dickel Diane E. , Silverman Jill L. , Nord Alex S. 
Published: September 2019

AbstractGenes with multiple co-active promoters appear common in brain, yet little is known about functional requirements for these potentially redundant genomic regulatory elements. SCN1A, which encodes the NaV1.1 sodium channel alpha subunit, is one such gene with two co-active promoters. Mutations in SCN1A are associated with epilepsy, including Dravet Syndrome (DS). The majority of DS patients harbor coding mutations causing SCN1A haploinsufficiency, however putative causal non-coding promoter mutations have been identified. To determine the functional role of one of these potentially redundant Scn1a promoters, we focused on the non-coding Scn1a 1b regulatory region, previously described as a non-canonical alternative transcriptional start site. Mice harboring a deletion of the extended evolutionarily-conserved 1b non-coding interval exhibited surprisingly severe reductions of Scn1a and NaV1.1 expression in brain with accompanying electroencephalographic seizures and behavioral deficits. This work identified the 1b region as a critical disease-relevant regulatory element and provides evidence that non-canonical and seemingly redundant promoters can have essential function.
Linked optical and gene expression profiling of single cells at high throughput
Authors: Zhang Jesse Q. , Siltanen Christian A. , Liu Leqian , Chang Kai-Chun , Gartner Zev J. , Abate Adam R. 
Published: September 2019

AbstractSingle cell RNA sequencing has emerged as a powerful tool for characterizing cells, but not all phenotypes of interest can be observed through gene expression alone. Linking sequencing with optical analysis has provided insight into the molecular basis behind cellular function, but current approaches have limited throughput. Here, we present a high throughput platform for linked optical and gene expression profiling of single cells. We demonstrate accurate fluorescence and gene expression measurements from thousands of cells in a single experiment and use the platform to characterize DNA and RNA changes in Jurkat cells through the cell cycle. In addition to its scalability, our integration of microfluidics and array-based molecular biology holds promise for comprehensive multi-omics profiling of single cells.
A combined RNA-seq and whole genome sequencing approach for identification of non-coding pathogenic variants in single families
Authors: Bronstein Revital , Capowski Elizabeth E. , Mehrotra Sudeep , Jansen Alex D. , Navarro-Gomez Daniel , Maher Mathew , Place Emily , Sangermano Riccardo , Bujakowska Kinga M. , Gamm David M. , Pierce Eric A. 
Published: September 2019

AbstractInherited retinal degenerations (IRDs) are at the focus of current genetic therapeutic advancements. For a genetic treatment such as gene therapy to be successful an accurate genetic diagnostic is required. Genetic diagnostics relies on the assessment of the probability that a given DNA variant is pathogenic. Non-coding variants present a unique challenge for such assessments as compared to coding variants. For one, non-coding variants are present at much higher number in the genome than coding variants. In addition, our understanding of the rules that govern the non-coding regions of the genome is less complete than our understanding of the coding regions. Methods that allow for both the identification of candidate non-coding pathogenic variants and their functional validation may help overcome these caveats allowing for a greater number of patients to benefit from advancements in genetic therapeutics. We present here an unbiased approach combining whole genome sequencing (WGS) with patient induced pluripotent stem cell (iPSC) derived retinal organoids (ROs) transcriptome analysis. With this approach we identified and functionally validated a novel pathogenic non-coding variant in a small family with a previously unresolved genetic diagnosis.
E. coliTraR allosterically regulates transcription initiation by altering RNA polymerase conformation and dynamics
Authors: Chen James , Gopalkrishnan Saumya , Chiu Courtney , Chen Albert Y. , Campbell Elizabeth A. , Gourse Richard L. , Ross Wilma , Darst Seth A. 
Published: September 2019

AbstractTraR and its homolog DksA are bacterial proteins that regulate transcription initiation by binding directly to RNA polymerase (RNAP) rather than to promoter DNA. Effects of TraR mimic the combined effects of DksA and its cofactor ppGpp. How TraR and its homologs regulate transcription is unclear. Here, we use cryo-electron microscopy to determine structures ofEscherichia coliRNAP, with or without TraR, and of an RNAP-promoter complex. TraR binding induced RNAP conformational changes not seen in previous crystallographic analyses, and a quantitative analysis of RNAP conformational heterogeneity revealed TraR-induced changes in RNAP dynamics. These changes involve mobile regions of RNAP affecting promoter DNA interactions, including the βlobe, the clamp, the bridge helix, and several lineage-specific insertions. Using mutational approaches, we show that these structural changes, as well as effects on σ70region 1.1, are critical for transcription activation or inhibition, depending on the kinetic features of regulated promoters.
Transcriptomic analysis of native versus cultured human and mouse dorsal root ganglia focused on pharmacological targets
Authors: Wangzhou Andi , McIlvried Lisa A. , Paige Candler , Barragan-Iglesias Paulino , Guzman Carolyn A. , Dussor Gregory , Ray Pradipta R. , Gereau Robert W. , Price Theodore J. 
Published: September 2019

AbstractDorsal root ganglion (DRG) neurons detect sensory inputs and are crucial for pain processing. They are often studied in vitro as dissociated cell cultures with the assumption that this reasonably represents in vivo conditions. However, to our knowledge, no study has ever directly compared genome-wide transcriptomes of DRG tissue in vivo versus in vitro, or between different labs and culturing protocols. We extracted bilateral lumbar DRG from C57BL6/J mice and human organ donors, and acutely froze one side and processed the other side as a dissociated cell culture, which was then maintained in vitro for 4 days. RNA was extracted and sequenced using the NextSeq Illumina platform. Comparing native to cultured human or mouse DRG, we found that the overall expression level of many ion channels and GPCRs specifically expressed in neurons is markedly lower in culture, but still expressed. This suggests that most pharmacological targets expressed in vivo are present in culture conditions. However, there are changes in expression levels for these genes. The reduced relative expression for neuronal genes in human DRG cultures is likely accounted for by increased expression of genes in fibroblast-like and other proliferating cells, consistent with the mitotic status of many cells in these cultures. We did find a subset of genes that are typically neuronally expressed, increased in human and mouse DRG cultures, including genes associated with nerve injury and/or inflammation in preclinical models such as BDNF, MMP9, GAL, and ATF3. We also found a striking upregulation of a number of inflammation-associated genes in DRG cultures, although many were different between mouse and human. Our findings suggest an injury-like phenotype in DRG cultures that has important implications for the use of this model system for pain drug discovery.
Alzheimer’s genetic risk factorFERMT2(Kindlin-2) controls axonal growth and synaptic plasticity in an APP-dependent manner
Authors: Eysert Fanny , Coulon Audrey , Boscher Emmanuelle , Vreulx Anaїs-Camille , Flaig Amandine , Mendes Tiago , Hughes Sandrine , Grenier-Boley Benjamin , Hanoulle Xavier , Demiautte Florie , Bauer Charlotte , Marttinen Mikael , Takalo Mari , Amouyel Philippe , Desai Shruti , Pike Ian , Hiltunen Mikko , Chécler Frédéric , Farinelli Mélissa , Delay Charlotte , Malmanche Nicolas , Hébert Sébastien , Dumont Julie , Kilinc Devrim , Lambert Jean-Charles , Chapuis Julien 
Published: September 2019

ABSTRACTAlthough APP metabolism is being intensively investigated, a large fraction of its modulators are yet to be characterized. In this context, we combined two genome-wide high-content screenings to assess the functional impact of miRNAs and genes on APP metabolism and the signaling pathways involved. This approach highlighted the involvement ofFERMT2(or Kindlin-2), a genetic risk factor of Alzheimer’s disease (AD), as a potential key modulator of axon guidance; a neuronal process that depends on the regulation of APP metabolism. We found that FERMT2 directly interacts with APP to modulate its metabolism and that FERMT2 under-expression impacts axonal growth, synaptic connectivity and long-term potentiation in an APP-dependent manner. Lastly, the rs7143400-T allele, which is associated with an increased AD risk and localized within the 3’UTR of FERMT2, induced a down-regulation of FERMT2 expression through binding of miR-4504 among others. This miRNA is mainly expressed in neurons and significantly overexpressed in AD brains compared to controls. Altogether, our data provide strong evidence for a detrimental effect of FERMT2 under-expression in neurons and insight on how this may influence AD pathogenesis.
Interhemispheric connections between olfactory bulbs improve odor detection
Authors: Kermen Florence , Yaksi Emre 
Published: September 2019

SUMMARYInterhemispheric connections enable interaction and integration of sensory information in bilaterian nervous systems and are thought to optimize sensory computations. However, the cellular and spatial organization of interhemispheric networks as well as the computational properties they mediate in vertebrates are still poorly understood. Thus, it remains unclear to which extent the connectivity between left and right brain hemispheres participates in sensory processing. Here, we show that the zebrafish olfactory bulbs (OBs) receive direct interhemispheric projections from their contralateral counterparts in addition to top-down inputs from the contralateral zebrafish homolog of olfactory cortex. The direct interhemispheric projections between the OBs reach peripheral layers of the contralateral OB and retain a fine-grained topographic organization, which directly connects similarly tuned olfactory glomeruli across hemispheres. In contrast, interhemispheric top-down inputs consist of diffuse projections that broadly innervate the inhibitory granule cell layer. Jointly, these interhemispheric connections elicit a balance of topographically organized excitation and non-topographic inhibition on the contralateral OB and modulate odor responses. We show that the interhemispheric connections in the olfactory system enable the modulation of odor response and improve the detection of a reproductive pheromone, when presented together with competing complex olfactory cues, by boosting the response of the pheromone selective neurons. Taken together, our data shows a previously unknown function for an interhemispheric connection between chemosensory maps of the olfactory system.
TGFβ signaling is required for tenocyte recruitment and functional neonatal tendon regeneration
Authors: Kaji Deepak A. , Howell Kristen L. , Huang Alice H. 
Published: September 2019

ABSTRACTTendon injuries are common with poor healing potential. The paucity of therapies for tendon is due to our limited understanding of the cells and molecules that drive tendon regeneration. Using a model of neonatal mouse tendon regeneration, we determined the molecular basis for regeneration and identify TGFβ signaling as a major pathway. Through targeted gene deletion, small molecule inhibition, and lineage tracing, we elucidated TGFβ-dependent and –independent mechanisms underyling tendon regeneration. Importantly, functional recovery depended on TGFβ signaling and loss of function is due to impaired tenogenic cell recruitment from bothScxlinand non-Scxlinsources. We show that TGFβ signaling is required directly in neonatal tenocytes for recruitment and that TGFβ is positively regulated in tendons. Collectively, these results are the first to show a functional role for TGFβ signaling in tendon regeneration and offer new insights toward the divergent cellular activities that may lead to regenerative vs fibrotic healing.
Peptide-based quorum sensing systems inPaenibacillus polymyxa
Authors: Voichek Maya , Maaß Sandra , Kroniger Tobias , Becher Dörte , Sorek Rotem 
Published: September 2019

AbstractPaenibacillus polymyxais an agriculturally important plant growth-promoting rhizobacterium. ManyPaenibacillusspecies are known to be engaged in complex bacteria-bacteria and bacteria-host interactions, which in other species were shown to necessitate quorum sensing communication. However, to date no quorum sensing systems have been described inPaenibacillus. Here we show that the type strainP. polymyxaATCC 842 encodes at least 16 peptide-based communication systems. Each of these systems is comprised of a pro-peptide that is secreted to the growth medium and processed to generate a mature short peptide. Each peptide has a cognate intracellular receptor of the RRNPP family, and we show that external addition ofP. polymyxacommunication peptides leads to reprogramming of the transcriptional response. We found that these quorum sensing systems are conserved across hundreds of species belonging to thePaenibacillaceaefamily, with some species encoding more than 25 different peptide-receptor pairs, representing a record number of quorum sensing systems encoded in a single genome.
Atoh1 is repurposed from neuronal to hair cell determinant by Gfi1 acting as a coactivator without redistribution of its genomic binding sites
Authors: Costa Aida , Powell Lynn M. , Soufi Abdenour , Lowell Sally , Jarman Andrew P. 
Published: September 2019

AbstractAlthough the lineage-determining ability of transcription factors is often modulated according to cellular context, the mechanisms by which such switching occurs are not well known. Using a transcriptional programming model, we found that Atoh1 is repurposed from a neuronal to an inner ear hair cell (HC) determinant by the combined activities of Gfi1 and Pou4f3. In this process, Atoh1 maintains its regulation of neuronal genes but gains ability to regulate HC genes. Pou4f3 enables Atoh1 access to genomic locations controlling the expression of sensory (including HC) genes, but Atoh1+Pou4f3 are not sufficient for HC differentiation. Gfi1 is key to the Atoh1-induced lineage switch, but surprisingly does not alter Atoh1’s binding profile. Gfi1 acts in two divergent ways. It represses the induction by Atoh1 of genes that antagonise HC differentiation, a function in keeping with its well-known repressor role in haematopoiesis. Remarkably, we find that Gfi1 also acts as a co-activator: it binds directly to Atoh1 at existing target genes to enhance its activity. These findings highlight the diversity of mechanisms by which one TF can redirect the activity of another to enable combinatorial control of cell identity.
Mapping Cell Membrane Organization and Dynamics Using Soft Nano-Imprint Lithography
Authors: Sansen T. , Sanchez-Fuentes D. , Rathar R. , Colom-Diego A. , El Alaoui F. , Viaud J. , Macchione M. , de Rossi S. , Matile S. , Gaudin R. , Bäcker V. , Carretero-Genevrier A. , Picas L. 
Published: September 2019

AbstractMembrane shape is a key feature of many cellular processes, including cell differentiation, division, migration, and trafficking. The development of nanostructured surfaces allowing for the in situ manipulation of membranes in living cells is crucial to understand these processes, but this requires complicated and limited-access technologies. Here, we investigate the self-organization of cellular membranes by using a customizable and bench top method allowing to engineer 1D SiO2 nanopillar arrays of defined sizes and shapes on high-performance glass compatible with advanced microscopies. As a result of this original combination, we provide a mapping of the morphology-induced modulation of the cell membrane mechanics, dynamics and steady-state organization of key protein complexes implicated in cellular trafficking and signal transduction.
High Resolution Structural Insights into Heliorhodopsin Family
Authors: Kovalev K. , Volkov D. , Astashkin R. , Alekseev A. , Gushchin I. , Haro-Moreno J. M. , Rogachev A. , Balandin T. , Borshchevskiy V. , Popov A. , Bourenkov G. , Bamberg E. , Rodriguez-Valera F. , Bueldt G. , Gordeliy V. 
Published: September 2019

AbstractRhodopsins are the most abundant light-harvesting proteins. A new family of rhodopsins, heliorhodopsins (HeRs), was recently discovered. In opposite to the known rhodopsins their N-termini face the cytoplasm. HeRs structure and function remain unknown. We present structures of two HeR-48C12 states at 1.5 Å showing its remarkable difference from all known rhodopsins. Its internal extracellular part is completely hydrophobic, while the cytoplasmic part comprises a cavity (’active site’), surrounded by charged amino acids and containing a cluster of water molecules, presumably being a primary proton acceptor from the Schiff base. At acidic pH a planar triangle molecule (acetate) is present in the ‘active site’ which demonstrated its ability to maintain such anions as carbonate or nitrate. Structure-based bioinformatic analysis identified 10 subfamilies of HeRs suggesting their diverse biological functions. The structures and available data suggest an enzymatic activity of HeR-48C12 subfamily and their possible involvement into fundamental redox biological processes.
Coordination of humoral immune factors dictates compatibility betweenSchistosoma mansoniandBiomphalaria glabrata
Authors: Li Hongyu , Hambrook Jacob R. , Pila Emmanuel A. , Gharamah Abdullah A. , Fang Jing , Wu Xinzhong , Hanington Patrick C. 
Published: September 2019

AbstractImmune factors in snails of the genusBiomphalariaare critical for combatingSchistosoma mansoni, the predominant cause of human intestinal schistosomiasis. Independently, many of these factors are known to play an important role in, but not fully define, the compatibility between the model snailB. glabrata,andS. mansoni. Here, we demonstrate association between four, previously characterized humoral immune molecules;BgFREP3,BgTEP1,BgFREP2 and Biomphalysin. We also identify unique immune determinants in the plasma ofS. mansoni-resistantB. glabratathat explain the incompatible phenotype. These factors coordinate to initiate haemocyte-mediated deestruction ofS. mansonisporocysts via production of reactive oxygen species. The inclusion ofBgFREP2 in aBgFREP3-initiated complex that also includesBgTEP1 almost completely explains resistance toS. mansoniin this model. Our study unifies many independent lines of investigation to provide a more comprehensive understanding of the snail immune system in the context of infection by this important human parasite.
Modeling the metabolic interplay between a parasitic worm and its bacterial endosymbiont allows the identification of novel drug targets
Authors: Curran DM , Grote A , Nursimulu N , Geber A , Voronin D , Jones Drew , Ghedin E , Parkinson J 
Published: September 2019

AbstractThe filarial nematodeBrugia malayirepresents a leading cause of disability in the developing world, causing lymphatic filariasis in nearly 40 million people. Currently available drugs are not well-suited to mass drug administration efforts, so new treatments are urgently required. One potential vulnerability is the endosymbiotic bacteriaWolbachia—present in many filariae—which is vital to the worm.Genome scale metabolic networks have been used to study prokaryotes and protists and have proven valuable in identifying therapeutic targets, but only recently have been applied to eukaryotic organisms. Here, we presentiDC625, the first compartmentalized metabolic model of a parasitic worm. We used this model to show how metabolic pathway usage allows the worm to adapt to different environments, and predict a set of 99 reactions essential to the survival ofB. malayi. We validated three of those reactions with drug tests and demonstrated novel antifilarial properties for all three compounds.