bluuebunny/crossref_metadata_2025_split huggingface dataset:
A thermostable phospholipase C obtained by consensus design
Authors: Val Diego S. , Nardo Luisina Di , Marchisio Fiorela , Peiru Salvador , Castelli María Eugenia , Abriata Luciano , Menzella Hugo G. , Rasia Rodolfo M. 
Published: October 2022

AbstractProteins’ extraordinary performance in recognition and catalysis have led their use in a range of applications. But proteins obtained from natural sources are oftentimes not suitable for direct use in industrial or diagnostic setups. Natural proteins, evolved to optimally perform a task in physiological conditions, usually lack the stability required to be used in harsher conditions. Therefore, the alteration of the stability of proteins is commonly pursued in protein engineering studies.Here we achieved a substantial thermal stabilization of a bacterial Zn(II) dependent phospholipase C by consensus sequence design. We retrieved and analyzed sequenced homologs from different sources, selecting a subset of examples for expression and characterization. A non-natural consensus sequence showed the highest stability and activity among those tested. Comparison of activity and stability parameters of this stabilized mutant and other natural variants bearing similar mutations allow us to pinpoint the sites most likely to be responsible for the enhancement. Point mutations in these sites alter the unfolding process of the consensus sequence. We show that the stabilized version of the protein retains full activity even in the harsh oil degumming conditions, making it suitable for industrial applications.
Simultaneous Membrane and RNA Binding by Tick-Borne Encephalitis Virus Capsid Protein
Authors: Pulkkinen L. I. A. , Barrass S. V. , Lindgren M. , Pace H. , Överby A. K. , Anastasina M. , Bally M. , Lundmark R. , Butcher S. J. 
Published: October 2022

AbstractTick-borne encephalitis virus is an enveloped, pathogenic, RNA virus in the family Flaviviridae, genus Flavivirus. Viral particles are formed when the nucleocapsid, consisting of an RNA genome and multiple copies of the capsid protein, buds through the endoplasmic reticulum membrane and acquires the viral envelope and the associated proteins. The coordination of the nucleocapsid components to the sites of assembly and budding are poorly understood. Here, we investigate nucleocapsid assembly by characterizing the interactions of the wild-type and truncated capsid proteins with membranes by using biophysical methods and model membrane systems. We show that capsid protein initially binds membranes via electrostatic interactions with negatively-charged lipids which is followed by membrane insertion. Additionally, we show that membrane-bound capsid protein can recruit viral genomic RNA. We confirm the biological relevance of the biophysical findings by using mass spectrometry to show that purified virions contain negatively-charged lipids. Our results suggest that nucleocapsid assembly is coordinated by negatively-charged membrane patches on the endoplasmic reticulum and that the capsid protein mediates direct contacts between the nucleocapsid and the membrane.
Introducing anrbcL andtrnL reference library to aid in the metabarcoding analysis of foraged plants from semi-arid eastern South African savannas
Authors: Botha Danielle , du Plessis Mornè , Siebert Frances , Barnard Sandra 
Published: October 2022

AbstractThe success of a metabarcoding study is determined by the extent of taxonomic coverage and the quality of records available in the DNA barcode reference database used. This study aimed to create anrbcLa andtrnL (UAA) DNA barcode sequence reference database of plant species that are potential herbivore foraging targets and commonly found in semi-arid savannas of eastern South Africa. A study-area-specific species list of 755 species was compiled. Thereafter, reference libraries forrbcLa andtrnL (UAA) sequences were created mined from sequence databases according to specific quality criteria to ensure accurate taxonomic coverage and resolution. The taxonomic reliability of these reference libraries was evaluated by testing for the presence of a barcode gap, identifying a data-appropriate identification threshold, and determining the identification accuracy of reference sequences via primary distance-based criteria. The finalrbcLa reference dataset consisted of 1238 sequences representing 318 genera and 562 species. The finaltrnL dataset consisted of 921 sequences representing 270 genera and 461 species. Barcode gaps were found for 76% of the taxa in therbcL barcode reference dataset and 68% of the taxa in thetrnL barcode reference dataset. The identification success rate, calculated with thek-nn criterion was 85.86% for therbcL dataset and 73.72% for thetrnL dataset. The datasets forrbcL andtrnL combined during this study are not presented as a complete DNA reference library, but rather as two datasets that should be used in unison to identify plants present in the semi-arid eastern savannas of South Africa.
Electron microscopy reveals toroidal shape of master neuronal cell differentiator REST – RE1-Silencing Transcription factor
Authors: Veverka Pavel , Brom Tomáš , Janovič Tomáš , Stojaspal Martin , Pinkas Matyáš , Nováček Jiří , Hofr Ctirad 
Published: October 2022

AbstractThe RE1-Silencing Transcription factor (REST) is essential for neuronal differentiation. Here, we report the first 18.5-angstrom electron microscopy structure of human REST. The refined electron map suggests that REST forms a torus that can accommodate DNA double-helix in the central hole. Additionally, we quantitatively described REST binding to the canonical DNA sequence of the neuron-restrictive silencer element. We developed protocols for the expression and purification of full-length REST and the shortened variant REST-N62 produced by alternative splicing. We tested the mutual interaction of full-length REST and the splicing variant REST-N62. Revealed structure-function relationships of master neuronal repressor REST will allow finding new biological ways of prevention and treatment of neurodegenerative disorders and diseases.
Setd2 inactivation sensitizes lung adenocarcinoma to inhibitors of oxidative respiration and mTORC1 signaling
Authors: Walter David M. , Gladstein Amy C. , Doerig Katherine R. , Natesan Ramakrishnan , Baskaran Saravana G. , Gudiel A. Andrea , Adler Keren M. , Acosta Jonuelle O. , Wallace Douglas C. , Asangani Irfan A. , Feldser David M. 
Published: October 2022

AbstractSETD2 is a tumor suppressor that is frequently inactivated in several cancer types. The mechanisms through which SETD2 inactivation promotes cancer are unclear, and whether targetable vulnerabilities exist in these tumors is unknown. Here we identify heightened mTORC1-associated gene expression programs and functionally higher levels of oxidative metabolism and protein synthesis as prominent consequences of Setd2 inactivation in KRAS-driven mouse models of lung adenocarcinoma. Blocking oxidative respiration and mTORC1 signaling abrogates the high rates of tumor cell proliferation and tumor growth specifically in SETD2-deficient tumors. Our data nominate SETD2 deficiency as a functional marker of sensitivity to clinically actionable therapeutics targeting oxidative respiration and mTORC1 signaling.
Inversion polymorphism in a complete human genome assembly
Authors: Porubsky David , Harvey William T. , Rozanski Allison N. , Ebler Jana , Höps Wolfram , Ashraf Hufsah , Hasenfeld Patrick ,   Human Pangenome Reference Consortium (HPRC),   Human Genome Structural Variation Consortium (HGSVC), Paten Benedict , Sanders Ashley D. , Marschall Tobias , Korbel Jan O. , Eichler Evan E. 
Published: October 2022

AbstractThe completion of the human genome significantly improved our ability to discover and interpret genome copy number variation. In order to understand its impact on the characterization of inversion polymorphisms, we remapped data from 41 human genomes and 10 new samples against the telomere-to-telomere (T2T) reference genome as compared to the standard GRCh38 reference. Our analysis shows a ~21% increase in sensitivity identifying and improving mapping of 63 inversions. We further identify 26 misorientations within GRCh38, and show that the T2T reference is three times more likely to represent the correct orientation of the major human allele. As a result, we report a significant bias for inversions accumulating within the pericentromeric regions of specific chromosomes and show that functional annotations around inverted regions, such as topological-associated domains, can be better interpreted.
A novel nuclease is the executing part of a bacterial plasmid defense system
Authors: Weiß Manuela , Giacomelli Giacomo , Ben Assaya Mathilde , Grundt Finja , Haouz Ahmed , Peng Feng , Petrella Stéphanie , Wehenkel Anne Marie , Bramkamp Marc 
Published: October 2022

AbstractCells are continuously facing the risk of taking up foreign DNA that can compromise genomic and cellular integrity. Therefore, bacteria are in a constant arms race with mobile genetic elements such as phages, transposons and plasmids. They have developed several active strategies against invading DNA molecules that can be seen as a bacterial ‘innate immune system’. Anti-phage systems are usually organized in ‘defense islands’ and can consist of restriction-modification (R-M) systems, CRISPR-Cas, and abortive infection (Abi) systems. Despite recent advances in the field, much less is known about plasmid defense systems. We have recently identified the MksBEFG system inCorynebacterium glutamicumas a novel plasmid defense system which comprises homologues of the condensin system MukFEB. Here, we investigated the molecular arrangement of the MksBEFG complex. Importantly, we identified MksG as a novel nuclease that degrades plasmid DNA and is, thus, the executing part of the system. The crystal structure of MksG revealed a dimeric assembly through its DUF2220 C-terminal domains. This domain is homologous to the TOPRIM domain of the topoisomerase II family of enzymes and contains the corresponding divalent ion binding site that is essential for DNA cleavage in topoisomerases, explaining thein vitronuclease activity of MksG. We further show that the MksBEF subunits exhibit an ATPase cycle similar to MukBEFin vitroand we reason that this reaction cycle, in combination with the nuclease activity provided by MksG, allows for processive degradation of invading plasmids. Super-resolution localization microscopy revealed that the Mks system is spatially regulated via to the polar scaffold protein DivIVA. Introduction of plasmids increases the diffusion rate and alters the localization of MksG, indicating an activation of the systemin vivo.
Phosphorylation chemistry of the Bordetella PlrSR TCS and its contribution to bacterial persistence in the lower respiratory tract
Authors: Barr Sarah A. , Kennedy Emily N. , McKay Liliana S. , Johnson Richard M. , Ohr Ryan J. , Cotter Peggy A. , Bourret Robert B. 
Published: October 2022

2SUMMARYBordetella species cause lower respiratory tract infections in mammals. B. pertussis and B. bronchiseptica are the causative agents of whooping cough and kennel cough, respectively. The current acellular vaccine for B. pertussis protects against the pertussis toxin but does not prevent transmission or colonization. Cases of B. pertussis infections are on the rise even in areas of high vaccination. The PlrSR two-component system, is required for persistence in the mouse lung. A partial plrS deletion strain and a plrS H521Q strain cannot survive past three days in the lung, suggesting PlrSR works in a phosphorylation dependent mechanism. We characterized the biochemistry of B. bronchiseptica PlrSR and found that both proteins function as a canonical two-component system. His521 and Glu522 were essential for PlrS autophosphorylation. Asn525 was essential for phosphatase activity. The PAS domain was critical for both PlrS autophosphorylation and phosphatase activities. PlrS can both phosphotransfer to and exert phosphatase activity towards PlrR. Unexpectedly, PlrR forms a tetramer when unphosphorylated and a dimer upon phosphorylation. Finally, we demonstrated the importance of PlrS phosphatase activity for persistence within the murine lung. By characterizing PlrSR we hope to guide future in vivo investigation for development of new vaccines and therapeutics.10GRAPHICAL ABSTRACT
Growth performance and nutritional composition of Clarias gariepinus in flood-and-drain aquaponics coupled with three species of leafy vegetable
Authors: Olanrewaju Gbolaga O. , Sarpong David D. , Aremu Abiola O. , Ade-Ademilua Elizabeth O. 
Published: October 2022

AbstractRecirculating aquaponics integrate aquaculture and hydroponics with the aid of microorganisms to ensure a sustainable supply of fish and vegetables. In this study, we designed and constructed a flood-and-drain aquaponic system with Clarias gariepinus as the aquaculture and Celosia argentea, Corchorus olitorius, and Ocimum gratissimum as plant components. Nitrogenous waste from the aquaculture unit was circulated to the plant growth beds, which served as bio-oxidizers of toxic ammonia to nitrate and returned less toxic water back to the aquaculture unit. An evaluation of the growth parameters of C. gariepinus in aquaponics and control fish tanks revealed that aquaponics-raised C. gariepinus gained an additional 205.6% of their initial mean weight, whereas those grown in the control fish tank gained an additional 182.2% of their initial mean weight. The majority (37.5%) of aquaponics-raised C. gariepinus weighed 750 g -1 kg, whereas the highest percentage of C. gariepinus raised in the control fish tank (23.4%) weighed 500 g - 700 g. C. gariepinus raised in aquaponics had significantly higher gross feed conversion efficiency and protein efficiency ratio than those raised in the control tank. The mortality rate in the aquaponic fish tank was 0% compared to the 11.43% mortality rate in the control fish tank. There was no significant difference in the nutritional composition of C. gariepinus raised in either tank; however, the aquaponic fish tank had a higher nitrogen retention rate. This study showed that C. gariepinus raised in aquaponics had better biomass accumulation than those raised in conventional fishponds.
Incipient parallel evolution of SARS-CoV-2 Deltacron variant in South Brazil
Authors: Sant’Anna Fernando Hayashi , Andreis Tiago Finger , Salvato Richard Steiner , Muterle Ana Paula , Comerlato Juliana , Gregianini Tatiana Schaffer , Barcellos Regina Bones , Godinho Fernanda Marques , Resende Paola Cristina , da Luz Wallau Gabriel , Castro Thaís Regina y , Casarin Bruna Campestrini , de Almeida Vieira Andressa , Schwarzbold Alexandre Vargas , de Arruda Trindade Priscila , Giannini Gabriela Luchiari Tumioto , Freese Luana , Bristot Giovana , Brasil Carolina Serpa , de Oliveira Rocha Bruna , Martins Paloma Bortolini , de Oliveira Francine Hehn , van Oosterhout Cock , Wendland Eliana 
Published: October 2022

AbstractWith the coexistence of multiple lineages and increased international travel, recombination and gene flow are likely to become increasingly important in the adaptive evolution of SARS-CoV-2. This could result in the incipient parallel evolution of multiple recombinant lineages. However, identifying recombinant lineages is challenging, and the true extent of recombinant evolution in SARS-CoV-2 may be underestimated. This study describes the first SARS-CoV-2 Deltacron recombinant case identified in Brazil. We demonstrate that the recombination breakpoint is at the beginning of Spike gene (S). The 5′ genome portion (circa 22 kb) resembles the AY.101 lineage (VOC Delta), and the 3′ genome portion (circa 8 kb nucleotides) is most similar to the BA.1.1 lineage (VOC Omicron). Furthermore, evolutionary genomic analyses indicate that the new strain emerged after a single recombination event between lineages of diverse geographical locations in December 2021 in South Brazil. This Deltacron, named AYBA-RS, is one out of almost 30 recombinants described this year. The submission of only four sequences in the GISAID database suggests that this Brazilian lineage had a minor epidemiological impact. On the other hand, the recent emergence of this and various other Deltacron recombinant lineages (i.e., XD, XF, and XS) suggests that gene flow and recombination may play an increasingly important role in the COVID-19 pandemic. We explain the evolutionary and population genetic theory that support this assertion, and we conclude that this stresses the need for continued genomic and epidemiological surveillance. This is particularly important for countries where multiple variants are present, as well as for countries that receive significant inbound international travel.
Type-specific EV-D68 real-time RT-PCR assay for the detection of all extant enterovirus D68 strains
Authors: Ng Terry Fei Fan , Nix W. Allan , Rogers Shannon L. , Emery Brian , Chern Shur-Wern , Butler Kiantra , Oberste M. Steven 
Published: October 2022

AbstractEnterovirus D68 (EV-D68) has caused recurring respiratory disease outbreaks in the United States since 2014. The dominant circulating EV-D68 strain has evolved from clade B1 to the more recent B2 and B3 clades. As recurrent outbreaks and continued virus evolution are expected for EV-D68, a robust real-time PCR assay that detects known strains as well as potential emerging strains is critical for national surveillance and clinical diagnostics. We describe a type-specific EV-D68 real-time RT-PCR (rRT-PCR) assay termed CDC2022, which targets sequences encoding conserved amino acid regions of all extant EV-D68 strains. We targeted three motifs conserved among all strains in the last 60 years. The assay achieved 100% (270/270) sensitivity and 100% (344/344) specificity when tested with a collection of 613 respiratory specimens, compared to the gold-standard EV semi-nested VP1 PCR and sequencing assay (snPCR/Seq). CDC2022 gave negative results with 289/289 non-target viruses, including 104 EV A-D isolates, 165 rhinovirus (RV) isolates or clinical specimens, and 14 other common respiratory viruses. The assay can detect as few as 0.28 CCID50per reaction. Anin silico“phylo-primer-mismatch” analysis was performed to visualize primer/probe mismatches and to compare CDC2022 with other EV-D68 rRT-PCR assays, including the previous CDC assay (CDC2015) developed in 2014 for clade B1 strains. It showed that CDC2022 has the fewest primer/probe mismatches among all assays analyzed and is suitable for all clades. We additionally tested 11 EV-D68-positive clinical specimens from 2022 that were confirmed by snPCR/Seq, and all were detected. CDC2022 assay could provide a critical tool for molecular surveillance of EV-D68.
The voltage-gated sodium channel NaV1.7 underlies endometriosis-associated chronic pelvic pain
Authors: Castro Joel , Maddern Jessica , Chow Chuen Yuen , Tran Poanna , Vetter Irina , King Glenn F. , Brierley Stuart M. 
Published: October 2022

AbstractChronic pelvic pain (CPP) is the primary symptom of endometriosis patients, but adequate treatments are lacking. Modulation of ion channels expressed by sensory nerves innervating the viscera have shown promise for the treatment of irritable bowel syndrome and overactive bladder. However, similar therapies have not been explored for endometriosis-associated CPP. Here we examined the role of the voltage-gated sodium (NaV) channel NaV1.7 in the sensitivity of vagina-innervating sensory afferents and investigated whether NaV1.7 inhibition reduces nociceptive signals from the vagina and ameliorates endometriosis-associated CPP. The mechanical responsiveness of vagina-innervating sensory afferents was assessed withex vivosingle unit recording preparations. Pain evoked by vaginal distension (VD) was quantified by the visceromotor response (VMR)in vivo. In control mice, pharmacological activation of NaV1.7 with OD1 sensitised vagina-innervating pelvic afferents to mechanical stimuli. Using a syngeneic mouse model of endometriosis, we established that endometriosis sensitized vagina-innervating pelvic afferents to mechanical stimuli. The highly selective NaV1.7 inhibitor Tsp1a revealed that this afferent hypersensitivity occurred in a NaV1.7-dependent manner. Moreover,in vivointra-vaginal treatment with Tsp1a reduced the exaggerated VMRs to VD that is characteristic of mice with endometriosis. Conversely, Tsp1a did not alterex vivoafferent mechanosensitivity orin vivoVMRs to VD in Sham control mice. Collectively, these findings suggest that NaV1.7 plays a crucial role in endometriosis-induced vaginal hyperalgesia. Importantly, NaV1.7 inhibition selectively alleviated endometriosis-associated CPP without the loss of normal sensation, suggesting that selective targeting of NaV1.7 could improve the quality of life of women with endometriosis.
Upregulation of CD55 complement regulator in distinct PBMC subpopulations of COVID-19 patients is associated with suppression of interferon responses
Authors: Detsika M. G. , Sakkou M. , Triantafillidou V. , Konstantopoulos D. , Grigoriou E. , Psarra K. , Jahaj E. , Dimopoulou I , Orfanos S. E. , Tsirogianni A. , Kollias G. , Kotanidou A. 
Published: October 2022

AbstractComplement activation has been verified in COVID-19 patients by both increased serum levels of complement factors C3a and C5b-9 and increased complement deposition at the tissue levels. Complement regulatory proteins (CRPs) CD55, CD46, CD59 and CR1 act to control complement overactivation and eliminate complement deposition and cell lysis. The aim of the study was to investigate the expression of CRPs in COVID-19 in order to identify potential dysregulated expression patterns of CRPs and address whether these may contribute to disease pathogenesis.Single cell RNA-sequencing (scRNA-seq) analysis performed on isolated PBMCs revealed an increase of CD55 expression in severe and critical COVID-19 patients compared to healthy controls. This increase was also detected upon integrated subclustering analysis of the monocyte, T cell and B cell populations. Flow cytometric analysis verified the distinct pattern of upregulated CD55 expression in monocyte and T cell sub populations of severe COVID-19 patients. This upregulation was associated with decreased expression of interferon stimulated genes (ISGs) in patients with severe COVID-19 suggesting a potential suppressor effect of CD55 on interferon responses. The present study identifies a COVID-19 specific CD55 expression pattern in PBMC subpopulations that coincides with reduced interferon responses thus indicating that the complement regulator CD55 may contribute to COVID-19 pathogenesis.
The FIGNL1-interacting protein C1orf112 is synthetic lethal with PICH and mediates RAD51 retention on chromatin
Authors: Stok Colin , van den Tempel Nathalie , Everts Marieke , Wierenga Elles , Bakker Femke , Kok Yannick , Alves Inês Teles , Jae Lucas T. , Bhattacharya Arkajyoti , Karanika Elefteria , Perepelkina Polina , Bergink Steven , Chan Kok-Lung , de Boer H. Rolf , Fehrmann Rudolf S.N. , van Vugt Marcel A.T.M. 
Published: October 2022

AbstractJoint DNA molecules are natural by-products of DNA replication and repair. Persistent joint molecules give rise to ultrafine DNA bridges (UFBs) in mitosis, which compromise sister chromatid separation. The DNA translocase PICH (ERCC6L) plays a central role in UFB resolution. A genome-wide loss-of-function screen was performed to identify the genetic contexts in which cells become dependent on PICH. In addition to genes involved in DNA condensation, centromere stability and DNA damage repair, we identified the uncharacterized protein C1orf112. We find that C1orf112 interacts with and stabilizes the AAA+ ATPase FIGNL1. Inactivation of either C1orf112 or FIGNL1 resulted in UFB formation, prolonged retention of RAD51 on chromatin, impaired replication fork dynamics, and consequently impaired genome maintenance. Combined, our data reveal that inactivation of C1orf112 and FIGNL1 dysregulates RAD51 dynamics at replication forks, resulting in DNA replication defects, and a dependency on PICH to preserve cell viability.