bluuebunny/crossref_metadata_2025_split huggingface dataset:
A large-scale genome-based survey of acidophilic Bacteria suggests that genome streamlining is an adaption for life at low pH
Authors: Cortez Diego , Neira Gonzalo , González Carolina , Vergara Eva , Holmes David S. 
Published: November 2021

AbstractGenome streamlining theory suggests that reduction of microbial genome size optimizes energy utilization in stressful environments. Although this hypothesis has been explored in several cases of low nutrient (oligotrophic) and high temperature environments, little work has been carried out on microorganisms from low pH environments and what has been reported is inconclusive. In this study, we performed a large-scale comparative genomics investigation of more than 260 bacterial high-quality genome sequences of acidophiles, together with genomes of their closest phylogenetic relatives that live at circum-neutral pH. A statistically supported correlation is reported between reduction of genome size and decreasing pH that we demonstrate is due to gene loss and reduced gene sizes. This trend is independent from other genome size constraints such as temperature and G+C content. Genome streamlining in the evolution of acidophilic Bacteria is thus supported by our results. Analyses of predicted COG categories and subcellular location predictions indicate that acidophiles have a lower representation of genes encoding extra-cellular proteins, signal transduction mechanisms and proteins with unknown function, but are enriched in inner membrane proteins, chaperones, basic metabolism, and core cellular functions. Contrary to other reports for genome streamlining, there was no significant change in paralog frequencies across pH. However, a detailed analysis of COG categories revealed a higher proportion of genes in acidophiles in the following categories: “Replication and repair”, “Amino acid transport” and “Intracellular trafficking”. This study brings increasing clarity regarding genomic adaptations of acidophiles to life at low pH while putting elements such as the reduction of average gene size under the spotlight of streamlining theory.
Accelerated Bone Healing in Calvarial and Femoral Defects with Injectable Microcarriers that Mimic the Osteogenic Niche
Authors: Haase Candice , Jaligama Sravani , Mondragon Eli , Pan Simin , McNeill Eoin P. , Co Cynthia , Tahan Daniel , Clough Bret H. , Sears Nick , Dominic Abhishai , Kameoka Jun , Gregory Carl A. , Kaunas Roland 
Published: November 2021

AbstractOsteo-enhanced human mesenchymal stem cells (OEhMSCs) secrete an osteogenic cell matrix (OCM) that mimics the composition of anabolic bone tissue and strongly enhances OEhMSC retention and subsequent bone repair in vivo. Here we demonstrate a system for rapid production of gelatin methacrylate microcarriers coated with decellularized OCM (OCM-GelMA) to serve as an injectable bone graft material with high osteogenic potential comparable to a clinically utilized gold standard, bone morphogenic protein 2 (BMP-2). OEhMSCs seeded onto OCM-GelMA secreted high levels of osteogenic and angiogenic cytokines and expressed higher levels of BMP-2 relative to OEhMSCs on bare GelMA microcarriers. OEhMSCs co-administered with OCM-GelMA microcarriers resulted in enhanced healing of murine critical-sized calvarial defects, which was comparable to that achieved with a BMP-2-laden gelatin sponge control. When tested in a murine femoral defect model, OCM-GelMA co-administered with OEhMSCs also induced profound bone growth within the defect. We submit that OCM-GelMA promotes OEhMSC paracrine release to accelerate bone repair, indicating their potential as a bone graft for use in minimally invasive surgery.
Bacterial chemotaxis to saccharides is governed by a trade-off between sensing and uptake
Authors: Norris Noele , Alcolombri Uria , Keegstra Johannes M. , Yawata Yutaka , Menolascina Filippo , Frazzoli Emilio , Levine Naomi M. , Fernandez Vicente I. , Stocker Roman 
Published: November 2021

AbstractTo swim up gradients of nutrients, E. coli senses nutrient concentrations within its periplasm. For small nutrient molecules, periplasmic concentrations typically match extracellular concentrations. However, this is not necessarily the case for saccharides, such as maltose, which is transported into the periplasm via a specific porin. Previous observations have shown that under various conditions E. coli limits maltoporin abundance so that, for extracellular micromolar concentrations of maltose, there are predicted to be only nanomolar concentrations of free maltose in the periplasm. Thus, in the micromolar regime, the total uptake of maltose from the external environment into the cytoplasm is limited not by the abundance of cytoplasmic transport proteins but by the abundance of maltoporins. Here we present results from experiments and modeling showing that this porin-limited transport enables E. coli to sense micromolar gradients of maltose despite having a high-affinity ABC transport system that is saturated at these micromolar levels. We used microfluidic assays to study chemotaxis of E. coli in various gradients of maltose and methyl-aspartate and leveraged our experimental observations to develop a mechanistic transport-and-sensing chemotaxis model. Incorporating this model into agent-based simulations, we discover a trade-off between uptake and sensing: although high-affinity transport enables higher uptake rates at low nutrient concentrations, it severely limits dynamic sensing range. We thus propose that E. coli may limit periplasmic uptake to increase its chemotactic sensitivity, enabling it to use maltose as an environmental cue.Statement of SignificanceBacterial chemotaxis is among the best-studied systems in biology and is paradigmatic of the mechanisms used by cells to link sensory inputs with regulated responses, thus providing insight into the ecological basis of cellular physiology. Here we present a mechanistic chemotaxis model that describes how the regulation of the transport of a sugar into and out of the cell’s periplasm affects the cell’s motile response to that sugar. Based on observations from population-level chemotaxis assays, we uncover an ecologically relevant trade-off between sensing and uptake. The general finding of this work is that, while high-affinity transport allows for higher uptake rates, it can severely limit the cell’s dynamic sensing range.
Protamine lacking piscine spermatozoa are transcriptionally active
Authors: Castro-Arnau Júlia , Chauvigné François , Gómez-Garrido Jessica , Esteve-Codina Anna , Dabad Marc , Alioto Tyler , Finn Roderick Nigel , Cerdà Joan 
Published: November 2021

AbstractTranscriptional quiescence of post-meiotic spermatozoa associated with protamine-mediated chromatin condensation is widely recognized in animals. How sperm acquire the extratesticular maturational competence to move and fertilize the egg is therefore thought to occur via non-transcriptional mechanisms. Here, using transcriptional profiling during spermatozoon differentiation in a fish that does not condense chromatin with protamines, we uncover spatially distinct roles of the GnRH receptor and PDGF signaling pathways between the somatic epithelia of the extratesticular ducts and the maturing spermatozoa. In vitro induction and inhibition experiments demonstrate that the endocrine signaling pathways are conserved in different lineages of fish and activate de novo transcription of spermatozoon genes required for the acquisition of full motility. These experiments further confirmed that mitochondrial translation is important for sperm maturation in anamniotes as in amniotes, but that transcriptional quiescence of post-meiotic spermatozoa is not a pan vertebrate phenomenon. On the contrary, the data show that the identified signal transduction pathways between the soma and the sperm upregulate effector genes essential for maturational competence and male fertility.
Biochemical analysis of deacetylase activity of rice sirtuin OsSRT1, a class IV member in plants
Authors: Mitra Nilabhra , Dey Sanghamitra 
Published: November 2021

AbstractThe role of plant sirtuins is slowly unwinding. There are only reports of H3K9Ac deacetylation by OsSRT1. This belongs to class IV sirtuin family with a longer C-terminus. Here C-terminus is required for ligand binding and catalysis. OsSRT1 can deacetylate the lys residues at the N terminal tail of both H3 and H4. It can also target the non-histone target, OsPARP1 playing a role in DNA damage repair pathway. Changes in the extent of different histone deacetylation by OsSRT1 is also related with different abiotic stress conditions. NAM and ADP-ribose has negative effect on OsSRT1 deacetylation.HighlightsOsSRT1 is capable of deacetylating various lysine residues of histone H3 and H4 as well as OsPARP1.The extra long C-terminus of OsSRT1 is required for its substrate binding and thus, its catalysis.On plant’s exposure to H2O2 and Arsenic toxicity, there is a relationship between increased expression of OsSRT1 and increased deacetylation of H3 and H4.
Natural variation in root exudation of GABA and DIMBOA impacts the maize root endosphere and rhizosphere microbiomes
Authors: Wang Peng , Lopes Lucas Dantas , Lopez-Guerrero Martha G. , van Dijk Karin , Alvarez Sophie , Riethoven Jean-Jack , Schachtman Daniel P. 
Published: November 2021

AbstractRoot exudates are important for shaping root-associated microbiomes. However, studies on a wider range of metabolites in exudates are required for a comprehensive understanding about their influence on microbial communities. We identified maize inbred lines that differ in exudate concentrations of DIMBOA and GABA using a semi-hydroponic system. These lines were grown in the field to determine the changes in microbial diversity and gene expression due to varying concentrations of DIMBOA and GABA in exudates using 16S rRNA amplicon sequencing and metatranscriptomics. Results showed individual and interaction effects of DIMBOA and GABA on the rhizosphere and root endosphere β-diversity, most strongly at the V10 growth stage. The main bacterial families affected by both compounds were Ktedonobacteraceae and Xanthomonadaceae. Higher concentrations of DIMBOA in exudates affected the rhizosphere metatranscriptome, enriching for KEGG pathways associated with plant disease. This study validated the use of natural variation within plant species as a powerful approach for understanding the role of root exudates on microbiome selection. We also showed that a semi-hydroponic system can be used to identify maize genotypes that differ in GABA and DIMBOA exudate concentrations under field conditions. The impact of GABA exudation on root-associated microbiomes was shown for the first time.
Single-Cell Multiomics Defines Tolerogenic Extrathymic Aire-Expressing Populations with Unique Homology to Thymic Epithelium
Authors: Wang Jiaxi , Lareau Caleb A. , Bautista Jhoanne , Gupta Alexander , Sandor Katalin , Germino Joe , Yin Yajie , Arvedson Matt , Reeder Gabriella C. , Cramer Nathan T. , Xie Fang , Ntranos Vasilis , Satpathy Ansuman T. , Anderson Mark S. , Gardner James M. 
Published: November 2021

AbstractThe Autoimmune Regulator (Aire) gene, well defined for its role in medullary thymic epithelial cells (mTECs) and immune self-tolerance, is also expressed in extrathymic Aire-expressing cells (eTACs) in the secondary lymphoid organs. eTACs have been shown to be hematopoietic antigen presenting cells (APCs) and potent inducers of immune tolerance (1–3). However, the precise identity and function of these cells remain unclear. Here, we use high-dimensional single-cell multiomics and functional approaches to define eTACs at the transcriptional, genomic, and proteomic level. We find that eTACs consist of two similar cell types: CCR7+ Aire-expressing migratory dendritic cells (AmDCs) and a unique Aire-hi population co-expressing Aire and RAR-related orphan receptor gamma-t (RORγt). The latter, which have significant transcriptional and genomic homology to migratory dendritic cells (migDCs) and mTECs, we term Janus cells (JCs). All eTACs, and JCs in particular, have a highly accessible chromatin structure and high levels of broad gene expression, including tissue-specific antigens, as well as remarkable transcriptional and genomic homology to thymic medullary epithelium. As in the thymus, Aire expression in eTACs is also dependent on RANK-RANK-ligand interactions. Furthermore, lineage-tracing shows that JCs are not precursors to the majority of AmDCs. Finally, self-antigen expression by eTACs is sufficient to mediate negative selection of T cells escaping thymic selection and can prevent autoimmune diabetes in non-obese diabetic mice. This transcriptional, genomic, and functional symmetry between a hematopoietic Aire-expressing population in the periphery and an epithelial Aire-expressing population in the thymus suggests that a core biological program may influence self-tolerance and self-representation across the spectrum of immune development.
Detection of Selenium/Sulphur substitution in the heterologous expression of Thioredoxin isoform 1 (Trx1) fromSaccharomyces cerevisiae
Authors: de Almeida Filho Humberto Antunes , Lery Leticia Miranda Santos , Valente Ana Paula , Netto Luis Eduardo Soares , de Almeida Fabio Ceneviva Lacerda 
Published: November 2021

AbstractThioredoxins are ubiquous proteins with 2 cysteines at the active site. The isoform 1 of Thioredoxin fromSaccharomyces cerevisae(Trx1) has six sulphur aminoacids, two cysteines and four methionines. In this work we performed the replacement of cysteines by selenocysteines by growth of a transformed celular expression vectorE. coli BL21-DE3in selenocysteine containing culture medium. The Maldi-TOF spectra of Seleno/Sulphur substituted Trx1 revealed six component peaks with 46-48 Da range between them, that is the isotopic Seleno-Sulfur difference, showing the replacement of the Cysteines and Methionines to Selenocysteines and Selenomethionines. The Maldi-TOF spectra of the peptides derived from Trypsin digestion of the purified Thioredoxin (peptide mass fingerprint) show Selenocysteine and Selenomethionine containing peptides. Therefore we are demonstrating that cystein can be replaced by selenocystein and be metabolically converted to selenomethionine during Trx1 heterologous translation. Furthermore, the Maldi-TOF spectra are showing the presence of the most abundant isotopes of selenium inserted in the peptides containing cysteine and methionine, derived from the Trx1 digestion. The one dimensional77Se–1Hheteronuclear multiple quantum coherence NMR spectroscopy (1D-HMQC) for reduced Seleno substituted Trx1 (Se-Trx1), revealed three ressonance lines for1Hβ1from Selenocysteines 30 and 33, between 1.6 and 2,0 ppm. The bidimensional HMQC spectra (2D-HMQC) of the reduced Se-Trx1 show the77Se ressonance signal in 178 ppm, coupled with1Hβ1and1Hβ2lines between 2.1 and 1.8 ppm. The 1D-HMQC for oxidized Trx1 revealed the only one broad resonance in 2.6 ppm probably relative to the1Hβ1prótons. The 2D-HMQC spectrum of oxidized protein shows a higher chemical shift of selenocysteine77Se (832 ppm) if compared to reduced state (178 ppm). Together these data are showing that the protocol ofSe – Ssubstitution developed here is a efficient method to label the active site of Thioredoxin 1 with a broad band chemical shift atom77Se. Furthermore the large spectral window of the77SeNMR detected between reduced and oxidized states of the Thioredoxin 1 shows that this atom is an excellent probe for accessing oxidative states and probably the conformational dynamics of the active site of the Se-Trx1.
Large-scale discovery of recombinases for integrating DNA into the human genome
Authors: Durrant Matthew G. , Fanton Alison , Tycko Josh , Hinks Michaela , Chandrasekaran Sita S. , Perry Nicholas T. , Schaepe Julia , Du Peter P. , Lotfy Peter , Bassik Michael C. , Bintu Lacramioara , Bhatt Ami S. , Hsu Patrick D. 
Published: November 2021

SUMMARYRecent microbial genome sequencing efforts have revealed a vast reservoir of mobile genetic elements containing integrases that could be useful genome engineering tools. Large serine recombinases (LSRs), such as Bxb1 and PhiC31, are bacteriophage-encoded integrases that can facilitate the insertion of phage DNA into bacterial genomes. However, only a few LSRs have been previously characterized and they have limited efficiency in human cells. Here, we developed a systematic computational discovery workflow that identifies thousands of new LSRs and their cognate DNA attachment sites by. We validate this approach via experimental characterization of LSRs in human cells, leading to three classes of LSRs distinguished from one another by their efficiency and specificity. We identify landing pad LSRs that efficiently integrate into synthetically installed attachment sites orthogonal to the human genome, human genome-targeting LSRs with computationally predictable pseudosites, and multi-targeting LSRs that can unidirectionally integrate cargos at with similar efficiency and superior specificity to commonly used transposases. LSRs from each category were functionally characterized in human cells, overall achieving up to 7-fold higher plasmid recombination than Bxb1 and genome insertion efficiencies of 40-70% with cargo sizes over 7 kb. Overall, we establish a paradigm for large-scale discovery of microbial recombinases and reconstruction of their target sites directly from microbial sequencing data. This strategy provides a rich resource of over 60 experimentally characterized LSRs that can function in human cells and thousands of additional candidates for large-payload genome editing without exposed DNA double-stranded breaks.
A Retrotranslocation Assay that Predicts Defective VCP/p97-Mediated Trafficking of a Retroviral Signal Peptide
Authors: Das Poulami , Xu Wendy Kaichun , Singh Gautam Amit Kumar , Lozano Mary M. , Dudley Jaquelin P. 
Published: November 2021

ABSTRACTStudies of viral replication have provided critical insights into host processes, including protein trafficking and turnover. Mouse mammary tumor virus (MMTV) is a betaretrovirus that encodes a functional 98-amino acid signal peptide (SP). MMTV SP is generated from both Rem and envelope precursor proteins by signal peptidase cleavage in the endoplasmic reticulum (ER) membrane. We previously showed that SP functions as an HIV-1 Rev-like protein that is dependent on the AAA ATPase VCP/p97 to subvert ER-associated degradation (ERAD). SP contains a nuclear/nucleolar localization sequence (NLS/NoLS) within the N-terminal 45 amino acids. To directly determine the SP regions needed for membrane extraction and trafficking, we developed a quantitative retrotranslocation assay with biotin acceptor peptide (BAP)-tagged SP proteins. Use of alanine substitution mutants of BAP-tagged MMTV SP in retrotranslocation assays revealed that mutation of amino acids 57 and 58 (M57-58) interfered with ER membrane extraction, whereas adjacent mutations did not. The M57-58 mutant also showed reduced interaction with VCP/p97 in co-immunoprecipitation experiments. Using transfection and reporter assays to measure activity of BAP-tagged proteins, both M57-58 and an adjacent mutant (M59-61) were functionally defective compared to wild-type SP. Confocal microscopy revealed defects in SP nuclear trafficking and abnormal localization of both M57-58 and M59-61. Furthermore, purified GST-tagged M57-58 and M59-61 demonstrated reduced ability to oligomerize compared to tagged wild-type SP. These experiments suggest that SP amino acids 57-58 are critical for VCP/p97 interaction and retrotranslocation, whereas residues 57-61 are critical for oligomerization and nuclear trafficking independent of the NLS/NoLS. Our results emphasize the complex host interactions with long signal peptides.IMPORTANCEEndoplasmic reticulum-associated degradation (ERAD) is a form of cellular protein quality control that is manipulated by viruses, including the betaretrovirus, mouse mammary tumor virus (MMTV). MMTV-encoded signal peptide (SP) has been shown to interact with an essential ERAD factor, VCP/p97 ATPase, to mediate its extraction from the ER membrane, also known as retrotranslocation, for RNA-binding and nuclear function. In this manuscript, we developed a quantitative retrotranslocation assay that identified an SP substitution mutant, which is defective for VCP interaction as well as nuclear trafficking, oligomer formation, and function. An adjacent SP mutant was competent for retrotranslocation and VCP interaction, but shared the other defects. Our results revealed the requirement for VCP during SP trafficking and the complex cellular pathways used by long signal peptides.
Computational characterization of recombinase circuits for periodic behaviors
Authors: Landau Judith , Cuba Samaniego Christian , Giordano Giulia , Franco Elisa 
Published: November 2021

AbstractIn nature, recombinases are site-specific proteins capable of rearranging DNA, and they are expanding the repertoire of gene editing tools used in synthetic biology. The on/off response of recombinases, achieved by inverting the direction of a promoter, makes them suitable for Boolean logic computation; however, recombinase-based logic gate circuits are single-use due to the irreversibility of the DNA rearrangement, and it is still unclear how a dynamical circuit, such as an oscillator, could be engineered using recombinases. Preliminary work has demonstrated that recombinase-based circuits can yield periodic behaviors in a deterministic setting. However, since a few molecules of recombinase are enough to perform the inverting function, it is crucial to assess how the inherent stochasticity at low copy number affects the periodic behavior. Here, we propose six different circuit designs for recombinase-based oscillators. We model them in a stochastic setting, leveraging the Gillespie algorithm for extensive simulations, and we show that they can yield periodic behaviors. To evaluate the incoherence of oscillations, we use a metric based on the statistical properties of auto-correlation functions. The main core of our design consists of two self-inhibitory, recombinase-based modules coupled by a common promoter. Since each recombinase inverts its own promoter, the overall circuit can give rise to switching behavior characterized by a regular period. We introduce different molecular mechanisms (transcriptional regulation, degradation, sequestration) to tighten the control of recombinase levels, which slows down the response timescale of the system and thus improves the coherence of oscillations. Our results support the experimental realization of recombinase-based oscillators and, more generally, the use of recombinases to generate dynamic behaviors in synthetic biology.
Association of cochlear outer hair cell - type II spiral ganglion afferents with protection from noise-induced hearing loss
Authors: Cederholm Jennie M.E. , Parley Kristina E. , Perera Chamini J. , von Jonquieres Georg , Pinyon Jeremy L. , Julien Jean-Pierre , Ryugo David K. , Ryan Allen F. , Housley Gary D. 
Published: November 2021

AbstractThe medial olivocochlear (MOC) efferent feedback circuit projecting to the cochlear outer hair cells (OHCs) confers protection from noise-induced hearing loss and is generally thought to be driven by inner hair cell (IHC) - type I spiral ganglion afferent (SGN) input. Knockout of the Prph gene (PrphKO) encoding the peripherin type III intermediate filament disrupted the OHC - type II SGN innervation and virtually eliminated MOC – mediated contralateral suppression from noise delivered to the opposite ear, measured as a reduction in cubic distortion product otoacoustic emissions. Electrical stimulation of the MOC pathway elicited contralateral suppression indistinguishable between wildtype (WT) and PrphKO mice, indicating that the loss of contralateral suppression was not due to disruption of the efferent arm of the circuit; IHC – type I SGN input was also normal, based on auditory brainstem responses. High-intensity, broadband noise (108 dB SPL, 1 hour) produced permanent hearing loss in PrphKO mice, but not in WT littermates. These findings associate OHC-type II input with MOC efferent - based otoprotection at loud sound levels.