bluuebunny/crossref_metadata_2025 huggingface dataset:
iPSC-based modeling of THD recapitulates disease phenotypes and reveals neuronal malformation
Authors: Tristán-Noguero Alba , Fernández-Carasa Irene , Calatayud Carles , Bermejo-Casadesús Cristina , Campa Leticia , Artigas Francesc , Domingo-Jiménez Rosario , Ibáñez Salvador , Pineda Mercè , Artuch Rafael , Raya Ángel , García-Cazorla Àngels , Consiglio Antonella
Published: February 2022

Tyrosine hydroxylase deficiency (THD) is a rare genetic disorder leading to dopaminergic depletion and early-onset parkinsonism. Affected children present with either a severe form that does not respond to L-Dopa treatment (THD-B), or a milder L-Dopa responsive form (THD-A). We generated induced pluripotent stem cells (iPSCs) from THD patients that were differentiated into dopaminergic neurons (DAn) and compared with control-DAn from healthy individuals and gene-corrected isogenic controls. Consistent with patients, THD iPSC-DAn displayed lower levels of DA metabolites and reduced TH expression, when compared to controls. Moreover, THD iPSC-DAn showed abnormal morphology, including reduced total neurite length and either an abnormal TH proximodistal gradient (THDA), or neurite arborization defects (THDB). Treatment of THD-iPSC-DAn with L-Dopa rescued the neuronal defects and disease phenotype only in THDA-DAn. Interestingly, L-Dopa treatment at the stage of neuronal precursors could prevent the alterations in THDB-iPSC-DAn, thus suggesting the existence of a critical developmental window in THD. Our iPSC-based model recapitulates THD disease phenotypes and response to treatment, representing a promising tool for investigating pathogenic mechanisms, drug screening, and personalized management.
Copepod functional traits and groups show divergent biogeographies in the global ocean
Authors: Benedetti Fabio , Wydler Jonas , Vogt Meike
Published: February 2022

Aim

The distribution of zooplankton functional traits is a key factor for regulating food web dynamics and carbon cycling in the oceans. Yet, we lack a clear understanding of how many functional groups (FGs) exist in the zooplankton and how their traits are distributed on a global scale. Here, we model and map the environmental habitats of copepod (i.e. the main component of marine zooplankton) FGs to identify regions sharing similar functional trait expression, at the community level.

Taxon

Marine planktonic Neocopepoda.

Location

Global ocean.

Methods

Factor analysis on mixed data and hierarchical clustering were used to identify copepod FGs based on five species-level functional traits. An ensemble of species distribution models was used to estimate the environmental niches of the species modelled and the community weighted mean values of the traits studied. Ocean regions were defined based on their community-level mean trait expression using a principal component analysis and hierarchical clustering.

Results

Eleven global copepod FGs were identified. They displayed contrasting latitudinal patterns in mean annual habitat suitability that could be explained by differences in environmental niche preferences: two FGs were associated with polar conditions, one followed the global temperature gradient, five were associated with tropical oligotrophic gyres, and the remaining three with boundary currents and counter currents. Four main regions of varying community weighted mean trait values emerged: the Southern Ocean, the northern and southern high latitudes, the tropical gyres, and the boundary currents and upwelling systems.

Conclusions

The present FGs will improve the representation of copepods in global marine ecosystem models. This study improves the understanding of the patterns and drivers of copepods trait biogeography and will serve as a basis for studying links between zooplankton biodiversity and ecosystem functioning in a context of climate change.
Database-independent analysis of probable post-translational modifications of soil proteins across ecosystems
Authors: Starke Robert , Schäpe Stephanie Serena , van den Bossche Tim , Martinovic Tijana , Parente Fernandes Maysa Lima , Delgado-Baquerizo Manuel , Bastida Felipe , Jehmlich Nico
Published: February 2022

The identification rate of measured peptide spectra to proteins barely scratches 1% in best-case scenarios. Hundreds of thousands of valuable spectra are lost as no viable match in the database is found. Here, we apply the delta m/z plot that was previously implemented in MSnbase as tool for quality control to 63 soil samples from three ecosystems with different vegetation (39 forests, 11 grasslands, and 13 shrublands) with the aim to extract probable post-translational modifications (PTM) without the need of a reference database. The validity of the approach was verified with amino acids proposed for their respective 1 Da mass interval and compared to their relative abundance in proteins. We found that the average probable PTM and most known PTMs proposed for the mass intervals are similar across ecosystems. Otherwise, 11 mass intervals changed significantly in relative abundance in the three ecosystems but only for one an annotation could be proposed. Our approach not only highlights the opportunity of the database-independent analysis in soil metaproteomics but paves the way for targeted analysis of the yet unknown PTMs.
EagleImp-Web: A Fast and Secure Genotype Phasing and Imputation Web Service using Field-Programmable Gate Arrays
Authors: Wienbrandt Lars , Prieß Christoph , Kässens Jan Christian , Franke Andre , Uhing Franziska , Ellinghaus David
Published: February 2022

Imputation web servers have been developed that allow phasing and imputation of genome-wide data without the need for own computing resources. However, their terms of use, data sharing with third parties, the security architecture of the web service, and the algorithms and parameter settings used are only partially disclosed. We developed EagleImp-Web, a fast, secure and convenient web service for phasing and imputation of genome-wide data. The web service uses technical improvements in phasing and imputation algorithms and a field-programmable gate array (FPGA) accelerator design to reduce computation time without loss of phasing and imputation quality. Other key features include no exposure of user information and input/output data to third parties, high data security and fast secure download, user authentication through 2-factor authentication, full control in managing user accounts, and full transparency of algorithms and their settings. EagleImp-Web provides simple and convenient functionalities for monitoring running jobs and selecting parameter settings and output information. Due to the speed advantage over a purely CPU-based implementation, EagleImp-Web offers the user the ability to choose a more resource-intensive parameter setting in exchange for computation time to further improve phasing and imputation quality. EagleImp-Web is freely availabe at https://hybridcomputing.ikmb.uni-kiel.de.
Organotopic Organization of the Cervical Vagus Nerve
Authors: Thompson Nicole , Ravagli Enrico , Mastitskaya Svetlana , Iacoviello Francesco , Stathopoulou Thaleia-Rengina , Perkins Justin , Shearing Paul R. , Aristovich Kirill , Holder David
Published: February 2022

Despite detailed characterization of fascicular organization of somatic nerves, the functional anatomy of fascicles evident in human and large mammal cervical vagus nerve is unknown. The vagus nerve is a prime target for intervention in the field of electroceuticals due to its extensive distribution to the heart, larynx, lungs, and abdominal viscera. However, current practice of the approved vagus nerve stimulation (VNS) technique is to stimulate the entire nerve. This produces indiscriminate stimulation of non-targeted effectors and undesired side effects. Selective neuromodulation is now a possibility with a spatially-selective vagal nerve cuff. However, this requires the knowledge of the fascicular organization at the level of cuff placement to inform selectivity of only the desired target organ or function. We imaged function over milliseconds with fast neural electrical impedance tomography and selective stimulation, and found consistent spatially separated regions within the nerve correlating with the three fascicular groups of interest, suggesting organotopy. This was independently verified with structural imaging by tracing anatomical connections from the end organ with microCT and the development of an anatomical map of the vagus nerve. This confirmed organotopic organization. Here we show, for the first time, localized fascicles in the porcine cervical vagus nerve which map to cardiac, pulmonary and recurrent laryngeal function (N=4). These findings pave the way for improved outcomes in VNS as unwanted side effects could be reduced by targeted selective stimulation of identified organ-specific fascicles and the extension of this technique clinically beyond the currently approved disorders to treat heart failure, chronic inflammatory disorders and more.
Phospho-proteomic analysis of CAR-T cell signaling following activation by antigen-presenting cancer cells
Authors: MacMullan Melanie A. , Dunn Zachary S. , Qu Yun , Wang Pin , Graham Nicholas A.
Published: February 2022

Chimeric antigen receptors (CARs) are synthetic biomolecules comprised of an extracellular antigen recognition domain and intracellular signaling domains. When expressed in immune cells, CARs direct their host cells to kill diseased cells expressing the antigen recognized by the CAR. Although signaling pathways downstream of CAR activation control the cytotoxic function of CAR-expressing cells, phospho-proteomic studies of CAR signaling have been limited. Most approaches have used antibodies or soluble ligands, rather than cell-displayed antigens, to activate CAR signaling. Here, we demonstrate an efficient and cost-effective label-free phospho-proteomic approach to analyze CAR signaling in immune cells stimulated with antigen-presenting cancer cells. Following co-culture of CAR-T cells with cancer cells, we first preserve phospho-signaling by cross-linking proteins with formalin. Then, we use magnet-activated cell sorting (MACS) to isolate CAR-T cells from the co-culture. Validation experiments demonstrated that formalin fixation did not alter the phospho-proteome and that MACS achieved >90% CAR-T cell purity. Next, we compared the phospho-proteome in CAR-T cells stimulated with either CD19-expressing or non-CD19-expressing SKOV3 ovarian cancer cells. This analysis revealed that CAR signaling activated known pathways including the mitogen- activated protein kinases (MAPKs) ERK1/2. Bioinformatic approaches further showed that CAR activation induced other signaling pathways including the MAPK p38α, protein kinase A, and checkpoint kinase 1 (CHK1). Taken together, this work presents an easy and inexpensive method to better understand CAR immunotherapy by label-free phospho-proteomic analysis of CAR signaling in immune cells stimulated by antigen- presenting cancer cells.
Load Non-Linearly Modulates Movement Reflex Gain in an Insect Leg via a Distributed Network of Identified Nonspiking Interneurons
Authors: Gebehart Corinna , Hooper Scott L. , Büschges Ansgar
Published: February 2022

Producing context-specific motor acts requires sensorimotor neural networks to integrate multiple sensory modalities. Some of this integration occurs via presynaptic interactions between proprioceptive afferent neurons themselves,1,2 other by afferents of different modalities targeting appropriate motor neurons (MNs).3–5 How the interneuronal network typically interposed between sensory afferents and MNs contributes to this integration, particularly at single-neuron resolution, is much less understood. In stick insects, this network contains nonspiking interneurons (NSIs) converging onto the posture-controlling slow extensor tibiae motor neuron (SETi). We analyzed how load altered movement signal processing by tracing the interaction of movement (femoral chordotonal organ, fCO) and load (tibial campaniform sensilla, tiCS) signals from the afferents through the NSI network to the motor output. On the afferent level, load reduced movement signal gain by presynaptic inhibition; tiCS stimulation elicited primary afferent depolarization and reduced fCO afferent action potential amplitude. In the NSI network, graded responses to movement and load inputs summed nonlinearly and increased the gain of NSIs opposing movement-induced reflexes. The gain of SETi and muscle movement reflex responses consequently decreased. Gain modulation was movement parameter-specific and required presynaptic inhibition; pharmacologically blocking presynaptic inhibition abolished load-dependent tuning of SETi responses. These data describe sensorimotor gain control at the sensory, premotor, and motor levels. Presynaptic inhibition-mediated nonlinear integration allowed the NSI network to respond to movement sensory input in a context (load)-dependent manner. These findings show how gain changes can allow premotor networks to integrate multiple sensory modalities and thus generate context-appropriate motor activity.
ABIOTIC STRESS GENE 1 mediates aroma volatiles accumulation by activating MdLOX1a in apple
Authors: Zhang Jing , Zhang Susu , Wang Yongxu , Zhang Shuhui , Liu Wenjun , Wang Nan , Fang Hongcheng , Zhang Zongying , Chen Xuesen
Published: February 2022

Fruit aroma is an important organoleptic quality, which influences consumer preference and market competitiveness. Aroma compound synthesis pathways in plants have been widely identified of which the lipoxygenase pathway is crucial for fatty acid catabolism to form esters in apple. However, the regulatory mechanism of this pathway remains elusive. In this study, linear regression analysis and transgene verification revealed that the lipoxygenase MdLOX1a participates in ester biosynthesis. Yeast one-hybrid library screening indicated that a novel abiotic stress gene, MdASG1 (ABIOTIC STRESS GENE 1), was a positive regulator of the MdLOX1a promoter and ester production based on yeast one-hybrid and dual-luciferase assays, and correlation analysis among eight apple cultivars. Overexpression of MdASG1 in apple and tomato stimulated the lipoxygenase pathway and increased the fatty acid-derived volatile content, whereas the latter was decreased by MdASG1 silencing. Furthermore, MdASG1 overexpression enhanced the salt-stress tolerance of tomato and apple ‘Orin’ calli accompanied by a higher content of fatty acid-derived volatiles compared with that of non-stressed transgenic tomato fruit. Collectively, these findings indicate that MdASG1 activates MdLOX1a expression and participates in the lipoxygenase pathway, subsequently increasing the accumulation of aroma compounds especially under moderate salt stress treatment. The results also provide insight into the regulation of aroma production, and the potential strategy of prudent development and utilization of saline-alkali land to produce high-quality fruit, thereby reducing pressure on arable land and ensuring national food security.

One-sentence Summary

MdASG1 directly activates MdLOX1a expression to promote aroma volatiles accumulation especially under moderate salt stress.
Differential recognition of computationally optimized H3 hemagglutinin influenza vaccine candidates by human antibodies
Authors: Abbadi Nada , Nagashima Kaito , Pena-Briseno Alma , Ross Ted M. , Mousa Jarrod J.
Published: February 2022

Among circulating influenza viruses in humans, H3N2 viruses typically evolve faster than other subtypes and have caused severe illness and deaths in millions of people since emerging in 1968. Computationally optimized broadly reactive antigen (COBRA) technology is one strategy to broaden vaccine-elicited antibody responses among influenza subtypes. In this study, we determined the structural integrity of an H3N2 COBRA HA, TJ5, and, as nearly all humans have pre-existing immunity to H3N2 influenza viruses, we probed the antigenic profile of several H3N2 COBRA HAs by assessing recognition of these immunogens by human B cells and monoclonal antibodies (mAbs). Of three recently described COBRA H3 HA antigens (TJ5, NG2, and J4), we determined that TJ5 and J4 HA proteins recognize pre-existing B cells (from the 2017-2018 vaccine season) more effectively than NG2 HA and a wild type Hong Kong/4801/2014 protein. H3 HA-specific human mAbs recognize wild type and COBRA HA proteins, and have functional activity against a broad panel of H3N2 viruses. mAb TJ5-5 recognizes TJ5 and J4 HA proteins, but has poor recognition of NG2 HA, similar to the global B cell analysis. To probe these recognition differences and to verify the structural integrity of the TJ5 HA protein, we determined a 3.4 Å structure via cryo-electron microscopy of TJ5-5 complexed with the TJ5 HA, which revealed residues important to the differential binding. Overall, these studies determined that COBRA H3 HA proteins have correct antigenic and structural features, and are recognized by B cells and mAbs isolated from seasonally vaccinated humans.

Importance

Vaccine development for circulating influenza viruses, particularly for the H3N2 subtype, remains challenging due to consistent antigenic drift. Computationally optimized broadly reactive antigen (COBRA) technology has proven effective for broadening influenza hemagglutinin (HA) elicited antibody responses compared to wild type immunogens. Here we determined the structural features and antigenic profiles of H3 COBRA HA proteins. Two H3 COBRA HA proteins, TJ5 and J4, are better recognized by pre-existing B cells and monoclonal antibodies from the 2017-2018 vaccine season compared to COBRA NG2 and a wild type A/Hong Kong/2014 HA protein. We determined a cryo-EM structure of one mAb that poorly recognizes NG2, mAb TJ5-5, in complex with the TJ5 COBRA HA protein and identified residues critical to mAb recognition. As NG2 is more effective than TJ5 for a recent Hong Kong/2019 virus, these data provide insights into the diminished effectiveness of influenza vaccines across vaccine seasons.
A robust modular automated neuroimaging pipeline for model inputs to TheVirtualBrain
Authors: Frazier-Logue Noah , Wang Justin , Wang Zheng , Sodums Devin , Khosla Anisha , Samson Alexandria , McIntosh Anthony R. , Shen Kelly
Published: February 2022

TheVirtualBrain, an open-source platform for large-scale network modelling, can be personalized to an individual using a wide range of neuroimaging modalities. With the growing number and scale of neuroimaging data sharing initiatives of both healthy and clinical populations comes an opportunity to create large and heterogeneous sets of dynamic network models to better understand individual differences in network dynamics and their impact on brain health. Here we present TheVirtualBrain-UK Biobank pipeline, a robust, automated and open-source brain image processing solution to address the expanding scope of TheVirtualBrain project. Our pipeline generates connectome-based modelling inputs compatible for use with TheVirtualBrain. We leverage the existing multimodal MRI processing pipeline from the UK Biobank made for use with a variety of brain imaging modalities. We add various features and changes to the original UK Biobank implementation specifically for informing large-scale network models, including user-defined parcellations for the construction of matching whole-brain functional and structural connectomes. Changes also include detailed reports for quality control of all modalities, a streamlined installation process, modular software packaging, updated software versions, and support for various publicly available datasets. The pipeline has been tested on both healthy and clinical populations and is robust to the morphological changes observed in aging and dementia. In this paper, we describe these and other pipeline additions and modifications in detail, as well as how this pipeline fits into the TheVirtualBrain ecosystem.
Microbial Community Dynamics Provide Evidence for Hypoxia During a Coral Reef Mortality Event
Authors: Doyle Shawn M. , Self Miabel J. , Hayes Joseph , Shamberger Kathryn E.F. , Correa Adrienne M.S. , Davies Sarah W. , Santiago-Vázquez Lory Z. , Sylvan Jason B.
Published: February 2022

In July 2016, a severe coral reef invertebrate mortality event occurred approximately 200km southeast of Galveston, Texas at the East Flower Garden Bank wherein upwards of 80% of corals in a 0.06 km2 area died. Based on surveys of dead corals and other invertebrates shortly after this mortality event, responders hypothesized that localized hypoxia was the most likely direct cause. However, no dissolved oxygen data were available to test this hypothesis because oxygen is not continuously monitored within the Flower Garden Banks sanctuary. Here we quantify microbial plankton community diversity based on four cruises over two years at the Flower Garden Banks, including a cruise just 5-8 days after the mortality event was first observed. In contrast with observations collected during baseline conditions, microbial plankton communities in the thermocline were differentially enriched with taxa known to be active and abundant in oxygen minimum zones or that have known adaptations to oxygen limitation shortly after the mortality event (e.g. SAR324, Thioglobaceae, Nitrosopelagicus, and Thermoplasmata MGII). Unexpectedly, these enrichments were not localized to the East Bank, but were instead prevalent across the entire study area, suggesting there was a widespread depletion of dissolved oxygen concentrations in the thermocline around the time of the mortality event. Hydrographic analysis revealed the southern East Bank coral reef (where the localized mortality event occurred) was uniquely within the thermocline at this time. Our results demonstrate how temporal monitoring of microbial communities can be a useful tool to address questions related to past environmental events.

IMPORTANCE

In the northwestern Gulf of Mexico in July 2016, upwards of 80% of corals in a small area of the East Flower Garden Bank coral reef suddenly died without warning. Oxygen depletion is believed to have been the cause. However, there was considerable uncertainty as no oxygen data is available from the time of the event. Microbes are sensitive to changes in oxygen and can be used as bioindicators of oxygen loss. In this study, we analyze microbial communities in water samples collected over several years at the Flower Garden Banks, including shortly after the mortality event. Our findings indicate that compared to normal conditions, oxygen depletion was widespread in the deep-water layer during the mortality event. Hydrographic analysis of water masses further revealed some of this low oxygen water likely upwelled onto the coral reef.
Construction of a constitutively active type III secretion system for heterologous protein secretion
Authors: Liang Julie Ming , Burdette Lisa Ann , Wong Han Teng , Tullman-Ercek Danielle
Published: February 2022

Proteins comprise a multibillion-dollar industry in enzymes and therapeutics, but bacterial protein production can be costly and inefficient. Proteins of interest (POIs) must be extracted from lysed cells, purified, and resolubilized. The Salmonella pathogenicity island 1 type III secretion system (T3SS) is a protein secretion complex in Salmonella that has been engineered to secrete heterologous proteins and addresses the problems associated with bacterial protein production. However, the current best practices method of T3SS pathway activation for secretion is not ideal for industrial scaleup. Previously, the T3SS was activated by plasmid-based overexpression of the T3SS transcriptional regulator, hilA, which requires the addition of a small molecule inducer to the culture media and adds significant cost to the production media. Plasmid-based expression is also subject to instability in large-scale fermentation. Here, we show that we can constitutively activate the T3SS by modulating the upstream transcriptional regulator, hilD, either through knocking out hilE, a repressor of HilD, or by adding transcriptional fusions to hilD. Finally, we combine the two most promising genomic modifications to build a constitutively active T3SS capable of secreting a range of heterologous proteins at titers comparable to those reported with synthetic induction of hilA. These improvements further our goal of making an industrially competitive protein production strain that reduces the challenges associated with plasmid induction and maintenance.

Importance

Proteins are used in our everyday lives as therapeutics (insulin), industrial enzymes (laundry detergent), and bio-based materials (spider silk). Current industrial protein production in bacteria is costly because it requires purification of the target protein from the other proteins inside the cell. We solve this problem by engineering the Salmonella pathogenicity island 1 type III secretion system (T3SS) to export the target protein into the cell growth media. This makes the protein purification process more efficient and cheaper. However, this system currently requires an expensive inducer reagent to activate it which significantly increases the cost of the production media. We show here the creation of a constitutively active T3SS, meaning the T3SS pathway is always on. In doing so, we successfully created a Salmonella strain that eliminates the need for the inducer reagent and exports proteins at levels comparable to the inducer-activated system, ultimately reducing the cost of T3SS protein production.
Rapid genome surveillance of SARS-CoV-2 and study of risk factors using shipping container laboratories and portable DNA sequencing technology
Authors: Bilooei Sara Farahi , Jovicevic Dejana , Iranzadeh Arash , Thomas Anthony , Muscat Ivan , Mpofu Cynthia , Steiner Helene , Meany Thomas
Published: February 2022

In this paper we report on genome sequencing of 154 SARS-CoV-2 samples between June and July 2021 (Summer outbreak) in the Bailiwick of Jersey, a UK channel island. We have analysed extensive data collected on 598,155 RT-qPCR tests that identified 8,950 positive cases as part of public health surveillance from September 2020 to August 2021. Our study implemented an amplicon-based sequencing approach using the Oxford Nanopore Technology (ONT) portable device. This revealed the emergence of twelve AY sublineages and were clustered into the Delta sub-clades 21I and 21J. This was integrated alongside an existing RT-qPCR diagnostic laboratory to provide a sample-to-sequence turnaround time of approximately 30 hours with significant scope for optimisation. Owing to the geographic remoteness of the island from large scale sequencing infrastructure, this presents an opportunity to provide policy makers with near real-time sequencing findings. Our analysis suggests that age and sex remained a substantial risk factor for mortality. We observe viral loads are higher in advanced ages and unvaccinated individuals. The median age of SARS-CoV-2 positive individuals was higher during winter than the summer outbreak, and the contact tracing program showed that younger individuals stayed positive for longer.
Predictors of longitudinal cognitive ageing from age 70 to 82 including APOE e4 status, early-life and lifestyle factors
Authors: Corley Janie , Conte Federica , Harris Sarah E. , Taylor Adele M. , Redmond Paul , Russ Tom C. , Deary Ian J. , Cox Simon R.
Published: February 2022

Discovering why some people’s cognitive abilities decline more than others is a key challenge for cognitive ageing research. The most effective strategy may be to address multiple risk factors from across the life-course simultaneously in relation to robust longitudinal cognitive data. We conducted a 12-year follow-up of 1091 (at age 70) men and women from the longitudinal Lothian Birth Cohort 1936 study. Comprehensive repeated cognitive measures of visuospatial ability, processing speed, memory, verbal ability, and a general cognitive factor, were collected over five assessments (age 70, 73, 76, 79, and 82 years) and analysed using multivariate latent growth curve modelling. Fifteen life-course variables were used to predict variation in cognitive ability levels at age 70 and cognitive slopes from age 70 to 82. Only APOE e4 carrier status was found to be reliably informative of general- and domain-specific cognitive decline, despite there being many life-course correlates of cognitive level at age 70. APOE e4 carriers had significantly steeper slopes across all three fluid cognitive domains compared with non-carriers, especially for memory (β = −0.234, P< 0.001) and general cognitive function (β = −0.246, P<0.001), denoting a widening gap in cognitive functioning with increasing age. Our findings suggest that when many other candidate predictors of cognitive ageing slope are entered en masse, their unique contributions account for relatively small proportions of variance, beyond variation in APOE e4 status. We conclude that APOE e4 status is important for identifying those at greater risk for accelerated cognitive ageing, even among ostensibly healthy individuals.
Deficiency of the RNA-binding protein Cth2 extends yeast replicative lifespan by alleviating its repressive effects on mitochondrial function
Authors: Patnaik Praveen K. , Beaupere Carine , Barlit Hanna , Romero Antonia María , Tsuchiya Mitsuhiro , Muir Michael , Martínez-Pastor María Teresa , Puig Sergi , Kaeberlein Matt , Labunskyy Vyacheslav M.
Published: February 2022

Iron dyshomeostasis contributes to aging, but little information is available about the molecular mechanisms. Here, we provide evidence that, in Saccharomyces cerevisiae, aging is associated with altered expression of genes involved in iron homeostasis. We further demonstrate that defects in the conserved mRNA-binding protein Cth2, which controls stability and translation of mRNAs encoding iron-containing proteins, increase lifespan by alleviating its repressive effects on mitochondrial function. Mutation of the conserved cysteine residue in Cth2 that inhibits its RNA-binding activity is sufficient to confer longevity, whereas Cth2 gain-of-function shortens replicative lifespan. Consistent with its function in RNA degradation, we demonstrate that Cth2 deficiency relieves Cth2-mediated post-transcriptional repression of nuclear-encoded components of the electron transport chain. Our findings uncover a major role of the RNA-binding protein Cth2 in the regulation of lifespan and suggest that modulation of iron starvation signaling can serve as a target for potential aging interventions.
Association Mapping Across a Multitude of Traits Collected in Diverse Environments Identifies Pleiotropic Loci in Maize
Authors: Mural Ravi V. , Sun Guangchao , Grzybowski Marcin , Tross Michael C. , Jin Hongyu , Smith Christine , Newton Linsey , Andorf Carson M. , Woodhouse Margaret R. , Thompson Addie M. , Sigmon Brandi , Schnable James C.
Published: February 2022

Classical genetic studies have identified many cases of pleiotropy where mutations in individual genes alter many different phenotypes. Quantitative genetic studies of natural genetic variants frequently examine one or a few traits, limiting their potential to identify pleiotropic effects of natural genetic variants. Widely adopted community association panels have been employed by plant genetics communities to study the genetic basis of naturally occurring phenotypic variation in a wide range of traits. High-density genetic marker data – 18M markers – from two partially overlapping maize association panels comprising 1,014 unique genotypes grown in field trials across at least seven US states and scored for 162 distinct trait datasets enabled the identification of of 2,154 suggestive marker-trait associations and 697 confident associations in the maize genome using a resampling-based genome-wide association strategy. The precision of individual marker-trait associations was estimated to be three genes based a reference set of genes with known phenotypes. Examples were observed of both genetic loci associated with variation in diverse traits (e.g. above-ground and below-ground traits), as well as individual loci associated with the same or similar traits across diverse environments. Many significant signals are located near genes whose functions were previously entirely unknown or estimated purely via functional data on homologs. This study demonstrates the potential of mining community association panel data using new higher density genetic marker sets combined with resampling-based genome-wide association tests to develop testable hypotheses about gene functions, identify potential pleiotropic effects of natural genetic variants, and study genotype by environment interaction.
A TREX1 model reveals double-strand DNA preference and inter-protomer regulation
Authors: Hemphill Wayne O. , Hollis Thomas , Salsbury Freddie R. , Perrino Fred W.
Published: February 2022

The TREX1 3’ → 5’ exonuclease degrades DNAin vivoto prevent chronic immune activation through the cGAS-STING pathway. TREX1 degrades ss- and dsDNA containing a free 3’-hydroxyl, but the precise nature of immune-activating DNA remains an open question. The TREX1 homodimer structure is critical for exonuclease activity with amino acids from one protomer acting across the dimer interface contributing to catalysis in the opposing protomer. The unique TREX1 obligate homodimer structure suggests an intricate connection between the TREX1 protomers that has yet to be explained. We used biochemical assays, molecular dynamics simulations, and kinetic modeling to determine relative TREX1 affinities for ss- and dsDNA and to interrogate inter-protomer communication within the TREX1 homodimer. These new findings indicate that TREX1 is a semi-processive exonuclease with at least a 20-fold greater affinity for dsDNA than for ssDNA. Furthermore, we find extensively correlated dynamics between TREX1 protomers revealing newly identified substrate interactions in the TREX1 enzyme. These data indicate that TREX1 has evolved as a semi-processive exonuclease with a likelyin vivofunction to degrade dsDNA, where the TREX1 homodimer structure facilitates a mechanism for efficient binding and catabolism of dsDNA. These studies identify previously unrecognized regions of the TREX1 enzyme involved in DNA interactions, and our findings contribute to an emerging model of TREX1 exonuclease activity.
Horizontal gene transfer increases microbiome stability
Authors: Coyte K. Z. , Stevenson C. , Knight C. G. , Harrison E. , Hall J. P. J. , Brockhurst M. A.
Published: February 2022

Genes encoding resistance to stressors, such as antibiotics or environmental pollutants, are widespread across microbiomes, often encoded on mobile genetic elements. Yet despite their prevalence, the impact of resistance genes and their mobility upon the dynamics of microbial communities remains largely unknown. Here we develop eco-evolutionary theory to explore how resistance genes alter the stability of diverse microbiomes. We show that adding resistance genes to a microbiome typically increases its overall stability, particularly for mobile resistance genes with high transfer rates that efficiently spread resistance throughout the community. However, the impact of resistance genes upon the stability of individual taxa varies depending upon the mobility of the resistance gene and the network of ecological interactions within the community. Non-mobile resistance genes can benefit susceptible taxa in cooperative communities, yet damage those in competitive communities. Moreover, whilst the transfer of mobile resistance genes generally increases the stability of previously susceptible recipient taxa to perturbation, it can, counterintuitively, decrease the stability of the originally resistant donor species. We confirmed these theoretical predictions experimentally using competitive soil microcosm communities. Here the stability of a susceptible microbial community to perturbation was increased by adding mobile resistance genes encoded on conjugative plasmids but was decreased when these same genes were encoded on the chromosome. Together these findings highlight the importance of horizontal gene transfer in driving the eco-evolutionary dynamics of diverse microbiomes.
The lipid raft-associated protein stomatin is required for accumulation of dectin-1 in the phagosomal membrane and for full activity of macrophages against Aspergillus fumigatus
Authors: Goldmann Marie , Schmidt Franziska , Cseresnyés Zoltán , Orasch Thomas , Jahreis Susanne , Hartung Susann , Figge Marc Thilo , von Lilienfeld-Toal Marie , Heinekamp Thorsten , Brakhage Axel A.
Published: February 2022

Alveolar macrophages are among the first cells to come into contact with inhaled fungal conidia of the human pathogenic fungus Aspergillus fumigatus. In lung alveoli, they contribute to phagocytosis and elimination of conidia. As a counter defense, conidia contain a grey-green pigment allowing them to survive in phagosomes of macrophages for some time. Previously, we showed that this conidial pigment interferes with the formation of flotillin-dependent lipid-rafts in the phagosomal membrane thereby preventing the formation of functional phagolysosomes. In this study, the role of the lipid raft-associated protein stomatin in macrophages during antifungal defense was investigated. To determine the function of this integral membrane protein, a stomatin-deficient macrophage cell line was generated by CRISPR/Cas9 gene editing. Immunofluorescence and flow cytometry revealed that stomatin contributes to the phagocytosis of conidia and is important for recruitment of both the β-glucan receptor dectin-1 and the vATPase to the membrane of phagosomes. In the stomatin knockout cell line, fusion of phagosomes and lysosomes was reduced when infected with pigmentless pksP conidia. Thus, our data suggest that stomatin is involved in maturation of phagosomes via fostering fusion of phagosomes with lysosomes.
Ex Vivo Herpes Simplex Virus Reactivation Involves a DLK-Dependent Wave of Lytic Gene Expression that is Independent of Histone Demethylase Activity and Viral Genome Synthesis
Authors: Whitford Abigail L. , Clinton Corinne A. , Kennedy E.B. Lane , Dochnal Sara A. , Suzich Jon B. , Cliffe Anna R.
Published: February 2022

Herpes Simplex virus-1 (HSV-1) maintains a lifelong latent infection in neurons and periodically reactivates, resulting in the production of infectious virus. The exact cellular pathways that induce reactivation are not understood. In primary neuronal models of HSV latency, the cellular protein Dual Leucine Zipper kinase (DLK) has been found to initiate a wave of viral gene expression known as Phase I. Phase I occurs independently of both viral DNA replication and the activities of histone demethylase enzymes required to remove repressive heterochromatin modifications associated with the viral genome. Here we investigated whether Phase-I like gene expression occurs in ganglia reactivated from infected mice. Using the combined trigger of explant-induced axotomy and inhibition of PI3K signaling, we found that HSV lytic gene expression was induced rapidly from both sensory and sympathetic neurons.Ex vivoreactivation involved a wave of viral late gene expression that occurred independently of viral genome synthesis and histone demethylase activity, and preceded the detection of infectious virus. Importantly, we found that DLK was required for the initial induction of lytic gene expression. These data confirm the essential role of DLK in inducing HSV-1 gene expression from the heterochromatin associated genome and further demonstrate that HSV-1 gene expression during reactivation occurs via mechanisms that are distinct from lytic replication.
Reduced polymerase pausing compensates for increased chromatin accessibility in the aging liver
Authors: Bozukova Mihaela , Grbavac Dora , Nikopoulou Chrysa , Tessarz Peter
Published: February 2022

Regulation of gene expression is tightly linked to the organization of the mammalian genome. With age, chromatin alterations occur on all levels of genome organization, accompanied by changes in the gene expression profile. However, little is known about the changes on the level of transcriptional regulation with age. Here, we used a multi-omic approach and integrated ATAC-, RNA- and NET-seq to identify age-related changes in the chromatin landscape of murine liver and to investigate how these are linked to transcriptional regulation. We provide the first systematic inventory of the connection between aging, chromatin accessibility and transcriptional regulation in a whole tissue. We observe that aging in murine liver is accompanied by an increase in chromatin accessibility at promoter regions of protein-coding genes. Yet, although promoter accessibility is a requirement for transcription, the increased accessibility does not result in enhanced transcriptional output. Instead, aging is accompanied by a decrease of promoter-proximal pausing of RNA polymerase II (Pol II). We propose that these changes in transcriptional regulation are due to a reduced stability of the pausing complex and may represent a mechanism to compensate for the age-related increase in chromatin accessibility in order to prevent aberrant transcription.
Rif-Correct, a software tool for bacterial mRNA half-life estimation
Authors: Aretakis James R. , Schrader Jared M.
Published: February 2022

Background

Genome-wide measurement of bacterial mRNA lifetimes using the antibiotic rifampicin has provided new insights into the control of bacterial mRNA decay. However, for long polycistronic mRNAs, the estimation of mRNA half-life can be confounded with transcriptional runoff caused by rifampicin’s inhibition of initiating RNA polymerases, and not elongating RNA polymerases.

Results

We present the Rif-correct software package, a free open-source tool that uses transcriptome models of transcript architecture to allow for more accurate mRNA half-life estimates that account for the transcriptional runoff. Rif-correct is implemented as a customizable python script that allows for users to control all the analysis parameters to achieve improved mRNA half-life estimates.

Conclusions

Rif-correct is the first free open-source computational analysis pipeline for Rif-seq dataset mRNA half-life estimation. It is simple to run, fast, and easy to run with a detailed instruction manual and example datasets.
Plant autophagosomes mature into amphisomes prior to their delivery to the central vacuole
Authors: Zhao Jierui , Bui Mai Thu , Ma Juncai , Künzl Fabian , De La Concepcion Juan Carlos , Chen Yixuan , Petsangouraki Sofia , Mohseni Azadeh , Leon Marta Garcia , Gomez Marta Salas , Giannini Caterina , Gwennogan Dubois , Kobylinska Roksolana , Clavel Marion , Schellmann Swen , Jaillais Yvon , Friml Jiri , Kang Byung-Ho , Dagdas Yasin
Published: February 2022

Autophagosomes are double-membraned vesicles that traffic harmful or unwanted cellular macromolecules to the vacuole for recycling. Although autophagosome biogenesis has been extensively studied, mechanisms of autophagosome maturation, i.e., delivery and fusion with the vacuole, remain largely unknown in plants. Here, we have identified an autophagy adaptor, CFS1, that directly interacts with the autophagosome marker ATG8 and localizes on both membranes of the autophagosome. Autophagosomes form normally in Arabidopsis thaliana cfs1 mutants, but their delivery to the vacuole is disrupted. CFS1’s function is evolutionarily conserved in plants as it also localizes to the autophagosomes and plays a role in autophagic flux in the liverwort Marchantia polymorpha. CFS1 regulates autophagic flux by connecting autophagosomes with the ESCRT-I component VPS23, leading to the formation of amphisomes. Disrupting the VPS23-CFS1 interaction affects autophagic flux and renders plants sensitive to starvation stress. Altogether, our results reveal a deeply conserved mechanism of vacuolar delivery in plants that is mediated by amphisomes.
Identification and characterisation of Klebsiella pneumoniae and Pseudomonas aeruginosa clinical isolates with atypical β-lactam susceptibility profiles using Orbitrap liquid chromatography-tandem mass spectrometry
Authors: Takebayashi Yuiko , Dulyayangkul Punyawee , Satapoomin Naphat , Wan Nur Ismah Wan Ahmad Kamil , Williams O. Martin , Macgowan Alasdair P. , Heesom Kate J. , Williams Philip B. , Avison Matthew B.
Published: February 2022

There is significant interest in the possibility of predicting antibacterial drug susceptibility directly though the analysis of bacterial DNA or protein. We report the use of Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa and Acinetobacter baumannii transformants to define baseline predictive rules for the β- lactam susceptibility profiles of β-lactamase positive clinical isolates. We then deployed a robust and reproducible shotgun proteomics methodology to identify β-lactamase positivity and predict β-lactam susceptibility by reference to our baseline predictive rules both in cultured bacteria and in extracts of culture-positive blood. Proteomics and whole genome sequencing then allowed us to characterise K. pneumoniae and P. aeruginosa isolates that differed from the expected β-lactam susceptibility profile, iteratively expanding our predictive rules. Proteomics added considerable value over and above the information generated by whole genome sequencing, allowing for gene expression, not just gene presence to be considered. Specifically, in K. pneumoniae, we identified key differences between acrR and ramR regulatory mutations and compared the effects of OmpK36 Aspartate-Threonine or Glycine-Aspartate dipeptide porin insertions on susceptibility to cefepime and carbapenems. In P. aeruginosa, we identified differences in the gene expression effects of mexR versus nalC mutations and related these to differences in β-lactam MICs against isolates hyper-producing AmpC β-lactamase and or producing a metallo-β-lactamase.
Maturation of cortical endoplasmic reticulum clusters in the mouse oocyte: changes at fertilization
Authors: Wang Huizhen , Christenson Lane K. , Kinsey William H.
Published: February 2022

Oocytes from many invertebrate and vertebrate species exhibit unique endoplasmic reticulum specializations (cortical ER clusters) thought be essential for egg activation. In examination of cortical ER clusters, we observed they were tethered to previously unreported fenestrae within the cortical actin layer. Further, studies demonstrated sperm preferentially bind to plasma membrane overlying the fenestrae, establishing close proximity to underlying ER clusters. Moreover, following sperm-oocyte fusion, cortical ER clusters undergo a previously unrecognized global maturational change in volume, shape, and calreticulin content that persists through sperm incorporation, before dispersing at the pronuclear stage. These changes did not occur in oocytes from females mated with Izumo1 -/- males demonstrating that gamete fusion plays an important role in ER cluster maturation. In addition to these global changes seen at sites distant to the sperm, highly localized ER modifications were noted at the sperm binding site as cortical ER clusters surround the sperm head during incorporation, then form a diffuse cloud surrounding the decondensing sperm nucleus. This study provides the first evidence that cortical ER clusters interact with the fertilizing sperm, indirectly through a previous unknown lattice work of actin fenestrae, then directly during sperm incorporation. These observations raise the possibility that oocyte ER cluster-sperm interactions provide a competitive advantage to the oocyte, which may not occur during assisted reproductive technologies such as intracytoplasmic sperm injection.

Summary Statement

Sperm-oocyte interactions stimulate global changes in cortical endoplasmic reticulum cluster structure as well as localized responses at the sperm binding site.