bluuebunny/crossref_metadata_2025 huggingface dataset:
Decision support system to evaluate VENTilation in the Acute Respiratory Distress Syndrome (DeVENT study) – Trial Protocol
Authors: Patel Brijesh , Mumby Sharon , Johnson Nicholas , Falaschetti Emanuela , Hansen Jorgen , Adcock Ian , McAuley Danny , Takata Masao , Karbing Dan S. , Jabaudon Matthieu , Schellengowski Peter , Rees Stephen E.
Published: September 2021

Background

The Acute Respiratory Distress Syndrome (ARDS) occurs in response to a variety of insults, and mechanical ventilation is life-saving in this setting, but ventilator induced lung injury can also contribute to the morbidity and mortality in the condition. The Beacon Caresystem is a model-based bedside decision support system using mathematical models tuned to the individual patient’s physiology to advise on appropriate ventilator settings. Personalised approaches using individual patient description may be particularly advantageous in complex patients, including those who are difficult to mechanically ventilate and wean, in particular ARDS.

Methods

We will conduct a multi-centre international randomised, controlled, allocation concealed, open, pragmatic clinical trial to compare mechanical ventilation in ARDS patients following application of the Beacon Caresystem to that of standard routine care to investigate whether use of the system results in a reduction in driving pressure across all severities and phases of ARDS.

Discussion

Despite 20 years of clinical trial data showing significant improvements in ARDS mortality through mitigation of ventilator induced lung injury, there remains a gap in its personalised application at the bedside. Importantly, the protective effects of higher positive end-expiratory pressure (PEEP) were noted only when there were associated decreases in driving pressure. Hence, the pressures set on the ventilator should be determined by the diseased lungs’ pressure-volume relationship which is often unknown or difficult to determine. Knowledge of extent of recruitable lung could improve the ventilator driving pressure. Hence, personalised management demands the application of mechanical ventilation according to the physiological state of the diseased lung at that time. Hence, there is significant rationale for the development of point-of-care clinical decision support systems which help personalise ventilatory strategy according to the current physiology. Furthermore, the potential for the application of the Beacon Caresystem to facilitate local and remote management of large numbers of ventilated patients (as seen during this COVID-19 pandemic), could change the outcome of mechanically ventilated patients during the course of this and future pandemics.

Trial registration

ClinicalTrials.gov identifier (NCT number): NCT04115709

Administrative information

Note: the numbers in curly brackets in this protocol refer to SPIRIT checklist item numbers. The order of the items has been modified to group similar items (see http://www.equator-network.org/reporting-guidelines/spirit-2013-statement-defining-standard-protocol-items-for-clinical-trials/).
High definition analyses of single cohort, whole genome sequencing data provides a direct route to defining sub-phenotypes and personalising medicine
Authors: Joyce KE , Onabanjo E , Brownlow S , Nur F , Olupona KO , Fakayode K , Sroya M , Thomas G , Ferguson T , Redhead J , Millar CM , Cooper N , Layton DM , Boardman-Pretty F , Caulfield MJ ,   Genomics England Research Consortium, Shovlin CL
Published: September 2021

Possession of a clinical or molecular disease label alters the context in which life-course events operate, but rarely explains the phenotypic variability observed by clinicians. Whole genome sequencing of unselected endothelial vasculopathy patients demonstrated more than a third had rare, likely deleterious variants in clinically-relevant genes unrelated to their vasculopathy (1 in 10 within platelet genes; 1 in 8 within coagulation genes; and 1 in 4 within erythrocyte hemolytic genes). High erythrocyte membrane variant rates paralleled genomic damage and prevalence indices in the general population. In blinded analyses, patients with greater hemorrhagic severity that had been attributed solely to their vasculopathy had more deleterious variants in platelet (Spearman ρ=0.25, p=0.008) and coagulation (Spearman ρ=0.21, p=0.024) genes. We conclude that rare diseases can provide insights for medicine beyond their primary pathophysiology, and propose a framework based on rare variants to inform interpretative approaches to accelerate clinical impact from whole genome sequencing.
Past dynamics of HIV transmission among men who have sex with men in Montréal, Canada: a mathematical modelling study
Authors: Milwid Rachael M , Xia Yiqing , Doyle Carla , Cox Joseph , Lambert Gilles , Thomas Réjean , Mishra Sharmistha , Grace Daniel , Lachowsky Nathan J , Hart Trevor A , Boily Marie-Claude , Maheu-Giroux Mathieu
Published: September 2021

Background

Gay, bisexual, and other men who have sex with men (gbMSM) experience disproportionate risks of HIV acquisition/transmission. In 2017, Montréal became the first Canadian Fast-Track city, setting the 2030 goal of zero new HIV infections. To inform local elimination efforts, we estimate the evolving role of prevention/risk behaviours and HIV transmission dynamics among gbMSM in Montréal between 1975-2019.

Methods

Data from local bio-behavioural surveys were analyzed to develop, parameterize, and calibrate an agent-based model of sexual HIV transmission. Partnership dynamics, the HIV natural history, and treatment and prevention strategies were considered. The model simulations were analyzed to estimate the fraction of HIV acquisitions/transmissions attributable to specific age-groups and unmet prevention needs.

Results

The model-estimated HIV incidence peaked in 1985 (2.2%; 90%CrI: 1.3-2.8%) and decreased to 0.1% (90%CrI: 0.04-0.3%) in 2019. Between 1990-2017, the majority of HIV acquisitions/transmissions occurred among men aged 25-44 years, and men aged 35-44 thereafter. The unmet prevention needs of men with >10 annual anal sex partners contributed 92-94% of transmissions and 63-73% of acquisitions annually. The primary stage of HIV played an increasing role over time, contributing to 12%-27% of annual transmissions over 1990-2019. In 2019, approximately 75% of transmission events occurred from men who had discontinued, or never initiated ART.

Conclusions

The evolving HIV landscape has contributed to the recent low HIV incidence among MSM in Montréal. The shifting dynamics identified in this study highlight the need for continued population-level surveillance to identify unmet prevention needs and core groups on which to prioritize elimination efforts.
Potential Pharmacodynamic Mechanism of the Main ingredients in Licorice for Chronic Obstructive Pulmonary Disease
Authors: Chen Cai , An Jianpeng , Shen Guodong , Shen Yang
Published: September 2021

Purpose

This study aimed to investigate the effect of essential ingredients of licorice on the chronic obstructive pulmonary disease (COPD).

Method

The ingredients information were obtained fromPubChem(https://pubchem.ncbi.nlm.nih.gov/), related genes about COPD was collected fromgeneCards(http://www.genecards.org/).Networkpharmacology was utilized in this study.

Result

The intersection data set contains 20 molecular targets between COPD and liquorice. Protein-protein interaction network showed that there are a total of 58 nodes and 137 edges involved. The link number of AKT1 in PPI network was 39, which is the highest level of interaction. MAPK1 is an important target of Licorice on COPD.

Conclusion

MAPK signaling pathway could be the important key target of main ingredients of licorice on COPD.
Compensatory reconfiguration of functional networks between white matter and grey matter in Alzheimer’s disease
Authors: Zhang Xingxing , Guan Qing , Dong Debo , Chen Fuyong , Yi Jing , Luo Yuejia , Zhang Haobo ,   Alzheimer’s Disease Neuroimaging Initiative
Published: September 2021

The temporal synchronization of BOLD signals within white matter (WM) and between WM and grey matter (GM) exhibited intrinsic architecture and cognitive relevance. However, few studies examined the network property within- and between-tissue in Alzheimer’s disease (AD). The hub regions with high weighted degree (WD) were prone to the neuropathological damage of AD. To systematically investigate the changes of hubs within- and between-tissue functional networks in AD patients, we used the resting-state fMRI data of 30 AD patients and 37 normal older adults (NC) from the ADNI open database, and obtained four types of voxel-based WD metrics and four types of distant-dependent WD metrics (ddWD) based on a series of Euclidean distance ranges with a 20mm increment. We found that AD patients showed decreased within-tissue ddWD in the thalamic nucleus and increased between-tissue ddWD in the occipito-temporal cortex, posterior thalamic radiation, and sagittal stratum, compared to NC. We also found that AD patients showed the increased between-tissue FCs between the posterior thalamic radiation and occipito-temporal cortex, and between the sagittal stratum and the salience and executive networks. The dichotomy of decreased and increased ddWD metrics and their locations were consistent with previous studies on the neurodegnerative and compensatory mechanisms of AD, indicating that despite the disruptions, the brain still strived to compensate for the neural inefficiency by reorganizing functional circuits. Our findings also suggested the short-to-medium ranged ddWD metrics between WM and GM as useful biomarker to detect the compensatory changes of functional networks in AD.
Explainable Machine Learning Analysis Reveals Gender Differences in the Phenotypic and Neurobiological Markers of Cannabis Use Disorder
Authors: Niklason Gregory , Rawls Eric , Ma Sisi , Kummerfeld Erich , Maxwell Andrea M. , Brucar Leyla R. , Drossel Gunner , Zilverstand Anna
Published: September 2021

Background

Cannabis Use Disorder (CUD) has been linked to environmental, personality, mental health, neurocognitive and neurobiological risk factors. While many studies have revealed gender differences in CUD, the relative importance of these complex factors by gender has not been described.

Methods

We conducted a data-driven examination of gender differences in CUD in a community sample of young adults (Human Connectome Project [HCP]; n = 1204, 54% female). We employed state-of-the-art machine learning methods [gradient tree boosting, XGBoost] in combination with novel factor ranking tools [SHapley’s Additive exPlanations (SHAP)] as an ‘explainable machine learning approach’ in the multimodal data collected by the HCP (phenotypic and brain data).

Results

We were able to successfully classify both cannabis dependence and cannabis use levels. Previously identified environmental, personality, mental health, neurocognitive, and brain factors highly contributed to the classification. Predominantly-male risk factors included personality (high openness), mental health (high externalizing, high childhood conduct disorder, high fear somaticism), neurocognitive (impulsive delay discounting, slow working memory performance) and brain (low hippocampal volume) factors. Conversely, predominantly-female risk factors included environmental (low education level, low instrumental support) factors.

Conclusions

Our data-driven analysis of gender differences in the multimodal risk factors underlying cannabis dependence and use levels demonstrate that environmental factors contribute more strongly to CUD in women, whereas individual factors such as personality, mental health and neurocognitive factors have a larger importance in men. This warrants further investigations, and suggests the importance of understanding how these differences relate to the development of effective treatment approaches.
Predation risk differentially affects aphid morphotypes: impacts on prey behavior, fecundity and transgenerational dispersal morphology
Authors: Hermann Sara L. , Bird Sydney A. , Ellis Danielle R. , Landis Douglas A.
Published: September 2021

To avoid predation, prey initiate anti-predator defenses such as altered behavior, physiology and/or morphology. Prey trait changes in response to perceived predation risk can influence several aspects of prey biology that collectively contribute to individual success and thus population growth. However, studies often focus on single trait changes in a discrete life stage or morphotype. We assessed how predation risk by Harmonia axyridis affects several important traits in the aphid, Myzus persicae: host plant preference, fecundity and investment in dispersal. Importantly, we examined whether these traits changed in a similar way between winged (alate) and wingless (apterous) adult aphid morphotypes, which differ in morphology, but also in life-history characteristics important for reproduction and dispersal. Host plant preference was influenced by the presence of H.axyridis odors in choice tests; wingless aphids were deterred by the odor of plants with H.axyridis whereas winged aphids preferred plants with H.axyridis present. Wingless aphids reared in the presence of ladybeetle cues produced fewer offspring in the short-term, but significantly more when reared with exposure to predator cues for multiple generations. However, winged aphid fecundity was unaffected by H.axyridis cues. Lastly, transgenerational plasticity was demonstrated in response to predation risk via increased formation of winged aphid morphotypes in the offspring of predator cue-exposed wingless mothers. Importantly, we found that responses to risk differ across aphid polyphenism and that plasticity in aphid morphology occurs in response to predation risk. Together our results highlight the importance of considering how predation risk affects multiple life stages and morphotypes.
Systematic shifts in the variation among host individuals must be considered in climate-disease theory
Authors: Mihaljevic Joseph R , Páez David J.
Published: September 2021

To make more informed predictions of host-pathogen interactions under climate change, studies have incorporated the thermal performance of host, vector, and pathogen traits into disease models. However, this body of work has ignored the fact that disease spread and long-term patterns of host population dynamics are largely determined by the variation in susceptibility among individuals in the host population. Furthermore, and especially for ectothermic host species, variation in susceptibility is likely to be plastic, influenced by variables such as environmental temperature. For example, as host individuals respond idiosyncratically to temperature, this could affect the population-level variation in susceptibility, such that there may be predictable functional relationships between variation in susceptibility and temperature. Quantifying the relationship between temperature and among-host trait variation will therefore be critical for predicting how climate change and disease will interact to influence host-pathogen population dynamics. Here, we use a model to demonstrate how short-term effects of temperature on the variation of host susceptibility can drive epidemic characteristics, fluctuations in host population sizes, and probabilities of host extinction. Our results emphasize that more research is needed in disease ecology and climate biology to understand the mechanisms that shape trait variation, not just trait averages.
Coordinated drift of receptive fields during noisy representation learning
Authors: Qin Shanshan , Farashahi Shiva , Lipshutz David , Sengupta Anirvan M. , Chklovskii Dmitri B. , Pehlevan Cengiz
Published: September 2021

Long-term memories and learned behavior are conventionally associated with stable neuronal representations. However, recent experiments showed that neural population codes in many brain areas continuously change even when animals have fully learned and stably perform their tasks. This representational “drift” naturally leads to questions about its causes, dynamics, and functions. Here, we explore the hypothesis that neural representations optimize a representational objective with a degenerate solution space, and noisy synaptic updates drive the network to explore this (near-)optimal space causing representational drift. We illustrate this idea in simple, biologically plausible Hebbian/anti-Hebbian network models of representation learning, which optimize similarity matching objectives, and, when neural outputs are constrained to be nonnegative, learn localized receptive fields (RFs) that tile the stimulus manifold. We find that the drifting RFs of individual neurons can be characterized by a coordinated random walk, with the effective diffusion constants depending on various parameters such as learning rate, noise amplitude, and input statistics. Despite such drift, the representational similarity of population codes is stable over time. Our model recapitulates recent experimental observations in hippocampus and posterior parietal cortex, and makes testable predictions that can be probed in future experiments.
Weakly activated core inflammation pathways were identified as a central signaling mechanism contributing to the chronic neurodegeneration in Alzheimer’s disease
Authors: Li Fuhai , Eteleeb Abdallah , Buchser William , Wang Guoqiao , Xiong Chengjie , Payne Philip R. , McDade Eric , Karch Celeste M. , Harari Oscar , Cruchaga Carlos
Published: September 2021

Neuro-inflammation signaling has been identified as an important hallmark of Alzheimer’s disease (AD) in addition to amyloid β plaques (Aβ) and neurofibrillary tangles (NFTs). However, our knowledge of neuro-inflammation is very limited; and the core signaling pathways associated with neuro-inflammation are missing. From a novel perspective, i.e., investigating weakly activated molecular signals (rather than the strongly activated molecular signals), in this study, we uncovered the core neuro-inflammation signaling pathways in AD. Our novel hypothesis is that weakly activated neuro-inflammation signaling pathways can cause neuro-degeneration in a chronic process; whereas, strongly activated neuro-inflammation often cause acute disease progression like in COVID-19. Using the two large-scale genomics datasets, i.e., Mayo Clinic (77 control and 81 AD samples) and RosMap (97 control and 260 AD samples), our analysis identified 7 categories of signaling pathways implicated on AD and related to virus infection: immune response, x-core signaling, apoptosis, lipid dysfunctional, biosynthesis and metabolism, and mineral absorption signaling pathways. More interestingly, most of genes in the virus infection, immune response and x-core signaling pathways, are associated with inflammation molecular functions. Specifically, the x-core signaling pathways were defined as a group of 9 signaling proteins: MAPK, Rap1, NF-kappa B, HIF-1, PI3K-Akt, Wnt, TGF-beta, Hippo and TNF, which indicated the core neuro-inflammation signaling pathways responding to the low-level and weakly activated inflammation and hypoxia, and leading to the chronic neuro-degeneration. The core neuro-inflammation signaling pathways can be used as novel therapeutic targets for effective AD treatment and prevention.
Visible light optical coherence tomography of peripapillary retinal nerve fiber layer reflectivity in glaucoma
Authors: Song Weiye , Zhang Sui , Kim Yumi Mun , Saldlak Natalie , Fiorello Marissa G. , Desai Manishi , Yi Ji
Published: September 2021

Purpose

To evaluate the clinical utility of visible light optical coherence tomography (VIS-OCT) and to test whether VIS-OCT reflectivity and spectroscopy of peripapillary retinal nerve fiber layer (pRNFL) are correlated with severity of glaucoma, compared with standard-of-care OCT thickness measurements.

Design

Cross-sectional study.

Method

Fifty-four eyes from three groups of subjects (normal, glaucoma suspect, and glaucoma) were scanned by a custom-designed dual-channel device that simultaneously acquired visible (VIS-OCT) and near-infrared OCT (NIR-OCT) images. A 5×5 mm2 scan was taken of the peripapillary nerve fiber layer (pRNFL).

The pRNFL reflectivity was calculated for both channels and the spectroscopic marker was quantified by pVN, defined by the ratio of VIS-OCT to NIR-OCT pRNFL reflectivity. The results were compared with ophthalmic exams and clinical Zeiss OCT measurements. Mixed linear model was used to evaluate the association of imaging markers with glaucoma severity.

Results

VIS-OCT pRNFL reflectivity significantly, se uentially declined from normal to suspect to glaucoma (3.42 ± 0.35, 2.58 ± 0.37, 2.16 ± 0.39, mean ± SD), as did NIR-OCT pRNFL reflectivity (2.46 ± 0.24, 2.27 ± 0.24, 1.96 ± 0.25). The pVN also had the same decreasing trend among three groups (1.39 ± 0.08, 1.14 ± 0.08, 1.08 ± 0.10). Normal and suspect eyes were significantly different in VIS-OCT pRNFL reflectivity (p=0.002), pVN (p<0.001) but not in NIR-OCT pRNFL reflectivity (p=0.14), circumpapillary RNFL (cpRNFL) thickness (p=0.17), or macular ganglion cell layer and inner plexiform layer (GCL+IPL) thickness (p=0.07) in the mixed linear model.

Conclusion

VIS-OCT pRNFL reflectivity and pVN was more sensitive in separating suspect eyes from normal ones than clinical OCT thickness measurements. VIS-OCT pRNFL reflectivity and pVN could be a useful metric in early detection of glaucoma upon further longitudinal validation.
Single-Cell transcriptional changes in hypothalamic CRH-expressing neurons after early-life adversity inform enduring alterations in responses to stress
Authors: Short Annabel K , Wilcox Christina , Chen Yuncai , Pham Aidan L , Birnie Matthew T , Bolton Jessica L , Mortazavi Ali , Baram Tallie Z.
Published: September 2021

Mental and cognitive health, as well as vulnerability to neuropsychiatric disorders, involve the interplay of genes with the environment, particularly during sensitive developmental periods. Early-life stress / adversity (ELA) promotes vulnerabilities to stress-related affective disorders, yet it is unknown how a transient ELA dictates life-long neuroendocrine and behavioral reactions to stress. The population of hypothalamic corticotropin-releasing hormone (CRH)-expressing neurons that regulate stress-responses is a promising candidate to mediate the enduring influences of ELA on stress-related behavioral and hormonal responses via enduring transcriptional and epigenetic mechanisms. Capitalizing on a well-characterized model of ELA, we examined here the ELA-induced changes in gene expression profiles of stress-sensitive CRH-neurons in the hypothalamic paraventricular nucleus (PVN) of male mice. Given the known heterogeneity of these neuronal populations, we employed single-cell RNA sequencing (RNA-seq) approaches. The use of single-cell transcriptomics identified distinct CRH-expressing neuronal populations characterized by both their gene expression repertoire and their neurotransmitter profiles. Expression changes provoked by ELA clustered around genes involved in neuronal differentiation, synapse formation, altered energy metabolism and the cellular responses to stress and injury. Notably, the ELA-induced transcriptional changes took place primarily in subpopulations of glutamatergic CRH cells. Finally, ELA-induced transcriptional reprogramming of hypothalamic CRH-expressing neurons heralded significant, enduring disruptions of both hormonal and behavioral responses to stress throughout life.
Caspase-mediated nuclear pore complex trimming in cell differentiation and endoplasmic reticulum stress
Authors: Cho Ukrae H. , Hetzer Martin W.
Published: September 2021

Abstract

During apoptosis, caspases degrade 8 out of ∼30 nucleoporins to irreversibly demolish the nuclear pore complex. However, for poorly understood reasons, caspases are also activated during cell differentiation. Here, we show that sublethal activation of caspases during myogenesis results in the transient proteolysis of four peripheral Nups and one transmembrane Nup. “Trimmed” NPCs become nuclear export-defective, and we identified in an unbiased manner several classes of cytoplasmic, plasma-membrane, and mitochondrial proteins that rapidly accumulate in the nucleus. NPC trimming by non-apoptotic caspases was also observed in neurogenesis and endoplasmic reticulum stress. Our results suggest that caspases can reversibly modulate nuclear transport activity, which allows them to function as agents of cell differentiation and adaptation at sublethal levels.
Optogenetic activators of apoptosis, necroptosis and pyroptosis for probing cell death dynamics and bystander cell responses
Authors: Shkarina Kateryna , Hasel de Carvalho Eva , Santos José Carlos , Leptin Maria , Broz Petr
Published: September 2021

Targeted and specific induction of cell death in individual or groups of cells holds the potential for new insights into the response of tissues or organisms to different forms of death. Here we report the development of optogenetically-controlled cell death effectors (optoCDEs), a novel class of optogenetic tools that enables light-mediated induction of three types of programmed cell death (PCD) – apoptosis, pyroptosis and necroptosis – using Arabidopsis thaliana photosensitive protein Cryptochrome2. OptoCDEs enable rapid and highly specific induction of PCD in human, mouse and zebrafish cells and are suitable for a wide range of applications, such as sub-lethal cell death induction or precise elimination of single cells or cell populations in vitro and in vivo. As the proof-of-concept, we utilize optoCDEs to assess the differences in the neighboring cell response to apoptotic or necrotic PCD, revealing a new role for shingosine-1-phosphate signaling in regulating the efferocytosis of apoptotic cell by epithelia.
Empirical estimation of marine phytoplankton assemblages in coastal and offshore areas using anin situmulti-wavelength excitation fluorometer
Authors: Kodama Taketoshi , Taniuchi Yukiko , Kasai Hiromi , Yamaguchi Tamaha , Nakae Misato , Okumura Yutaka
Published: September 2021

Phytoplankton assemblages are important for understanding the quality of primary production in marine ecosystems. Here, we describe development of a methodology for monitoring marine phytoplankton assemblages using anin situmulti-wavelength excitation fluorometer (MEX) and its application for seasonal observations in coastal and offshore areas around Japan. The MEX recorded the fluorescence excited with nine light-emitting diodes, temperature, and sensor depth. We prepared reference datasets comprising temperature, MEX fluorescence, and plant-pigment-based chemotaxonomy phytoplankton assemblages. Target MEX fluorescence was decomposed by reference MEX fluorescence using a linear inverse model for calculating coefficients after the reference data were limited by temperature, followed by reconstruction of plant-pigment-based chemotaxonomy of the target MEX fluorescence using the coefficients and the chemotaxonomy assemblages of the reference data. Sensitivity analysis indicated poor estimation of the proportion and/or chlorophylla-based abundance of chlorophytes, haptophytes, prasinophytes, and prochlorophytes; however, limiting the estimations to five chemotaxonomic groups [diatoms, dinoflagellates, cryptophytes, cyanobacteria (cyanophytes and prochlorophytes), and other eukaryotes (chlorophytes, haptophytes, and prasinophytes)] resulted in positive correlations of both the proportion and abundances, suggesting that the five taxonomic abundances were well-estimated using the MEX. Additionally, MEX observations denoted spatial and seasonal variations of phytoplankton assemblages, with high contributions from other eukaryotes in every area and season, cyanobacteria highly during the summer in surface Kuroshio and Japan Sea waters, and diatoms in the Oyashio and Oyashio–Kuroshio transition areas and the Okhotsk Sea. Furthermore, ratios of water-column-integrated chlorophyll-based abundances to those on the surface at the chemotaxonomy group level were differed among the areas and groups. These findings suggested that phytoplankton-assemblage monitoring in the vertical direction is essential for evaluation of their current biomass, and that the MEX promotes the acquisition of these observations.
Translational gene expression control in Chlamydia trachomatis
Authors: Grieshaber Nicole A , Chiarelli Travis J , Appa Cody R , Neiswanger Grace , Peretti Kristina , Grieshaber Scott S
Published: September 2021

The human pathogen Chlamydia trachomatis proceeds through a multi phenotypic developmental cycle with each cell form specialized for different roles in pathogenesis. Understanding the mechanisms regulating this complex cycle has historically been hampered by limited genetic tools. In an effort to address this issue, we developed a translational control system to regulate gene expression in Chlamydia using a synthetic riboswitch. Here we demonstrate that translational control via a riboswitch can be used in combination with a wide range of promoters in C. trachomatis. The synthetic riboswitch E, inducible with theophylline, was used to replace the ribosome binding site of the synthetic promoter T5-lac, the native chlamydial promoter of the pgp4 plasmid gene and an anhydrotetracycline responsive promoter. In all cases the riboswitch inhibited translation, and high levels of protein expression was induced with theophylline. Combining the Tet transcriptional inducible promoter with the translational control of the riboswitch resulted in strong repression and allowed for the cloning and expression of the potent chlamydial regulatory protein, HctB. The ability to control the timing and strength of gene expression independently from promoter specificity is a new and important tool for studying chlamydial regulatory and virulence genes.
Structure and RNA template requirements of Arabidopsis RNA-DEPENDENT RNA POLYMERASE 2
Authors: Fukudome Akihito , Singh Jasleen , Mishra Vibhor , Reddem Eswar , Martinez-Marquez Francisco , Wenzel Sabine , Yan Rui , Shiozaki Momoko , Yu Zhiheng , Wang Joseph Che-Yen , Takagi Yuichiro , Pikaard Craig S.
Published: September 2021

RNA-dependent RNA polymerases play essential roles in RNA-mediated gene silencing in eukaryotes. In Arabidopsis, RNA-DEPENDENT RNA POLYMERASE 2 (RDR2) physically interacts with DNA-dependent NUCLEAR RNA POLYMERASE IV (Pol IV) and their activities are tightly coupled, with Pol IV transcriptional arrest or termination, involving the nontemplate DNA strand, somehow enabling RDR2 to engage Pol IV transcripts and generate double-stranded RNAs. The dsRNAs are then released from the Pol IV-RDR2 complex and diced into siRNAs that guide RNA-directed DNA methylation and silencing. Here we report the structure of full-length RDR2, at an overall resolution of 3.1 Å, determined by cryo-electron microscopy. The N-terminal region contains an RNA-recognition motif (RRM) adjacent to a positively charged channel that leads to a catalytic center with striking structural homology to the catalytic centers of multisubunit DNA-dependent RNA polymerases. We show that RDR2 initiates 1-2 nucleotides (nt) internal to the 3’ ends of its templates and can transcribe the RNA of an RNA-DNA hybrid provided that 9 or more nucleotides at the RNA’s 3’ end is unpaired. Using a nucleic acid configuration that mimics the arrangement of RNA and DNA strands upon Pol IV transcriptional arrest, we show that displacement of the RNA 3’ end occurs as the DNA template and non-template strands reanneal, enabling RDR2 transcription. These results suggest a model in which Pol IV arrest and backtracking displaces the RNA 3’ end as the DNA strands reanneal, allowing RDR2 to engage the RNA and transcribe the second strand.

Significance

RDR2 is critical for siRNA-directed DNA methylation in Arabidopsis, functioning in physical association with DNA-dependent Pol IV to synthesize the second strands of double-stranded siRNA precursors. Basepairing between the DNA template strand transcribed by Pol IV and the nontemplate DNA strand is known to induce Pol IV arrest and Pol IV-RDR2 transcriptional coupling, but how this occurs is unknown. We report the structure of RDR2 and experimental evidence for how RDR2 engages its RNA templates and initiates transcription. RDR2 engages the ends of RNAs displaced from RNA-DNA hybrids, suggesting a model in which Pol IV arrest and backtracking, accompanied by DNA strand reannealing, extrudes the 3’ end of the Pol IV transcript, allowing RNA engagement and second-strand synthesis.
Mechanical loading of cranial joints minimizes the craniofacial phenotype in Crouzon syndrome
Authors: Moazen Mehran , Hejazi Mahbubeh , Savery Dawn , Jones Dominic , Marghoub Arsalan , Alazmani Ali , Pauws Erwin
Published: September 2021

Children with syndromic forms of craniosynostosis undergo a plethora of surgical interventions to resolve the clinical features caused by the premature fusion of cranial sutures. While surgical correction is reliable, the need for repeated rounds of invasive treatment puts a heavy burden on the child and their family.

This study explores a non-surgical alternative using mechanical loading of the cranial joints to prevent or delay craniofacial phenotypes associated with Crouzon syndrome.

We treated Crouzon syndrome mice before the onset of craniosynostosis by cyclical mechanical loading of cranial joints using a custom designed set-up.

Cranial loading applied to the frontal bone partially restores normal skull morphology, significantly reducing the typical brachycephalic appearance. This is underpinned by the delayed closure of the coronal suture and of the intersphenoidal synchondrosis.

This study provides a novel treatment alternative for syndromic craniosynostosis which has the potential to be an important step towards replacing, reducing or refining the surgical treatment of all craniosynostosis patients.
An integrated pipeline and multi-model graphical user interface for accurate nano-dosimetry
Authors: Botte Ermes , Vagaggini Pietro , Zanoni Ilaria , Gardini Davide , Costa Anna Luisa , Ahluwalia Arti
Published: September 2021

Accurate dosing of nanoparticles is crucial for risk assessment and for their safe use in medical and other applications. Although it is well-known that nanoparticles sediment, diffuse and aggregate as they move through a fluid, and that therefore the effective dose perceived by cells may not necessarily be that initially administered, dose quantification remains a challenge. This is because to date, methods for accurate dose estimation are difficult to implement, involving precise characterization of the nanomaterial and the exposure system as well as complex mathematical operations. Here we present a pipeline for accurate nano-dosimetry of engineered nanoparticles on cell monolayers, based on an easy-to-use graphical software - DosiGUI - which integrates two well-established particokinetic and particodynamic models. DosiGUI is an open source tool which was developed to facilitate nano-dosimetrics. The pipeline includes methods for determining the stickiness index which describes the affinity between nanoparticles and cells. Our results show that accurate estimations of the effective dose cannot prescind from rigorous characterization of the stickiness index, which depends on both nanoparticle characteristics and cell type.
Targeting current species ranges and carbon stocks fails to conserve biodiversity in a changing climate: opportunities to support climate adaptation under 30×30
Authors: Dreiss Lindsay M. , Lacey L. Mae , Weber Theodore C. , Delach Aimee , Niederman Talia E. , Malcom Jacob W.
Published: September 2021

Protecting areas for climate adaptation will be essential to ensuring greater opportunity for species conservation well into the future. However, many proposals for protected areas expansion focus on our understanding of current spatial patterns, which may be ineffective surrogates for future needs. A science-driven call to address the biodiversity and climate crises by conserving at least 30% of lands and waters by 2030, 30×30, presents new opportunities to inform the siting of new protections globally and in the U.S. Here we identify climate refugia and corridors based on a weighted combination of currently available models; compare them to current biodiversity hotspots and carbon-rich areas to understand how 30×30 protections siting may be biased by data omission; and compare identified refugia and corridors to the Protected Areas Database to assess current levels of protection. Available data indicate that 20.5% and 27.5% of identified climate adaptation areas (refugia and/or corridor) coincides with current imperiled species hotspots and carbon-rich areas, respectively. With only 12.5% of climate refugia and corridors protected, a continued focus on current spatial patterns in species and carbon richness will not inherently conserve places critical for climate adaptation. However, there is ample opportunity for establishing future-minded protections: 52% of the contiguous U.S. falls into the top quartile of values for at least one class of climate refugia. Nearly 27% is already part of the protected areas network but managed for multiple uses that may limit their ability to contribute to the goals of 30×30. Additionally, nearly two-thirds of nationally identified refugia coincide with ecoregion-specific refugia suggesting representation of nearly all ecoregions in national efforts focused on conserving climate refugia. Based on these results, we recommend that land planners and managers make more explicit policy priorities and strategic decisions for future-minded protections and climate adaptation.
Mapping the Energy Landscape of PROTAC-Mediated Protein-protein Interactions
Authors: Villegas José A. , Vaid Tasneem M. , Johnson Michael E. , Moore Terry W.
Published: September 2021

One of the principal difficulties in computational modeling of macromolecules is the vast conformational space that arises out of large numbers of atomic degrees of freedom. This problem is a familiar issue in the area of protein-protein docking, where models of protein complexes are generated from the monomeric subunits. Although restriction of molecular flexibility is a commonly used approximation that decreases the dimensionality of the problem, the seemingly endless number of possible ways two binding partners can interact generally necessitates the use of further approximations to explore the search space. Recently, growing interest in using computational tools to build predictive models of PROTAC-mediated complexes has led to the application of state-of-the-art protein-protein docking techniques to tackle this problem. Additionally, the atomic degrees of freedom introduced by flexibility of linkers used in the construction of PROTACs further expands the configurational search space, a problem that can be tackled with conformational sampling tools. However, repurposing existing tools to carry out protein-protein docking and linker conformer generation independently results in extensive sampling of structures incompatible with PROTAC-mediated complex formation. Here we show that it is possible to restrict the search to the space of protein-protein conformations that can be bridged by a PROTAC molecule with a given linker composition by using a cyclic coordinate descent algorithm to position PROTACs into complex-bound configurations. We use this methodology to construct a picture of the energy landscape of PROTAC-mediated interactions in a model test case, and show that the global minimum lies in the space of native-like conformations.
The unfolded protein response triggers the immune deficiency pathway in ticks
Authors: Sidak-Loftis Lindsay C. , Rosche Kristin L. , Pence Natasha , Ujczo Jessica K. , Hurtado Joanna , Fisk Elis A. , Goodman Alan G. , Noh Susan M. , Peters John W. , Shaw Dana K.
Published: September 2021

The insect immune deficiency (IMD) pathway is a defense mechanism that senses and responds to Gram negative bacteria. Ticks lack genes encoding upstream components that initiate the IMD pathway. Despite this deficiency, core signaling molecules are present and functionally restrict tick-borne pathogens. The molecular events preceding activation remain undefined. Here, we show that the Unfolded Protein Response (UPR) initiates the IMD network inIxodes scapularisticks. The endoplasmic reticulum (ER) stress receptor, IRE1α, is phosphorylated in response to tick-borne bacteria, but does not splice the mRNA encoding XBP1. Instead, through protein modeling and reciprocal pulldowns, we show thatIxodesIRE1α complexes with TRAF2. Disrupting IRE1α-TRAF2 signaling blocks IMD pathway activation and diminishes the production of reactive oxygen species. Throughin vitro,in vivo,andex vivotechniques we demonstrate that the UPR-IMD pathway circuitry limits the Lyme disease-causing spirocheteBorrelia burgdorferiand the rickettsial agentsAnaplasma phagocytophilumandA. marginale(anaplasmosis). Altogether, our study uncovers a novel linkage between the UPR and the IMD pathway in ticks.
Protein disulphide isomerase (PDI) is protective against several types of DNA damage, including that induced by amyotrophic lateral sclerosis-associated mutant TDP-43 in neuronal cells/ in vitro models
Authors: Shadfar Sina , Vidal Marta , Parakh Sonam , Laird Angela S. , Atkin Julie D.
Published: September 2021

Protein disulphide isomerase (PDI) is a chaperone that catalyses the formation of thiol-disulphide bonds during protein folding. Whilst up-regulation of PDI is a protective mechanism to regulate protein folding, an increasingly wide range of cellular functions have been ascribed to PDI. Originally identified in the endoplasmic reticulum (ER), PDI has now been detected in many cellular locations, including the nucleus. However, its role in this cellular compartment remains undefined. PDI is implicated in multiple diseases, including amyotrophic lateral sclerosis (ALS), a fatal and rapidly progressing neurodegenerative condition affecting motor neurons. Loss of essential proteins from the nucleus is an important feature of ALS. This includes TAR DNA-binding protein-43 (TDP-43), a DNA/RNA binding protein present in a pathological form in the cytoplasm in almost all (97%) ALS cases, that is also mutated in a proportion of familial cases. PDI is protective against disease-relevant phenotypes associated with dysregulation of protein homeostasis (proteostasis) in ALS. DNA damage is also increasingly linked to ALS, which is induced by pathological forms of TDP-43 by impairment of its normal function in the non-homologous end-joining (NHEJ) mechanism of DNA repair. However, it remains unclear whether PDI is protective against DNA damage in ALS. In this study we demonstrate that PDI was protective against several types of DNA damage, induced by either etoposide, hydrogen peroxide (H2O2), or ALS-associated mutant TDP-43M337V in neuronal cells. This was demonstrated using widely used DNA damage markers, phosphorylated H2AX and 53BP1, which is specific for NHEJ. Moreover, we also show that PDI translocates into the nucleus following DNA damage. Here PDI is recruited directly to sites of DNA damage, implying that it has a direct role in DNA repair. This study therefore identifies a novel role of PDI in the nucleus in preventing DNA damage.
Optimised production of disulfide-bonded fungal effectors in E. coli using CyDisCo and FunCyDisCo co-expression approaches
Authors: Yu Daniel S. , Outram Megan A. , Crean Emma , Smith Ashley , Sung Yi-Chang , Darma Reynaldi , Sun Xizhe , Ma Lisong , Jones David A. , Solomon Peter S. , Williams Simon J.
Published: September 2021

Effectors are a key part of the arsenal of plant pathogenic fungi and promote pathogen virulence and disease. Effectors typically lack sequence similarity to proteins with known functional domains and motifs, limiting our ability to predict their functions and understand how they are recognised by plant hosts. As a result, cross-disciplinary approaches involving structural biology and protein biochemistry are often required to decipher and better characterise effector function. These approaches are reliant on high yields of relatively pure protein, which often requires protein production using a heterologous expression system. For some effectors, establishing an efficient production system can be difficult, particularly those that require multiple disulfide bonds to achieve their naturally folded structure. Here, we describe the use of a co-expression system within the heterologous host E. coli termed CyDisCo (cytoplasmic disulfide bond formation in E. coli) to produce disulfide bonded fungal effectors. We demonstrate that CyDisCo and a naturalised co-expression approach termed FunCyDisCo (Fungi-CyDisCo) can significantly improve the production yields of numerous disulfide bonded effectors from diverse fungal pathogens. The ability to produce large quantities of functional recombinant protein has facilitated functional studies and crystallisation of several of these reported fungal effectors. We suggest this approach could be useful when investigating the function and recognition of a broad range of disulfide-bond containing effectors.
Temporal and sex-dependent gene expression patterns in a renal ischemia-reperfusion injury and recovery pig model
Authors: Nemours Stéphane , Castro Luis , Ribatallada-Soriano Didac , Semidey Maria Eugenia , Aranda Miguel , Ferrer Marina , Sanchez Alex , Morote Joan , Cantero-Recasens Gerard , Meseguer Anna
Published: September 2021

Men are more prone to acute kidney injury (AKI) and chronic kidney disease (CKD), progressing to end-stage renal disease (ESRD) than women. Severity and capacity to regenerate after AKI are important determinants of CKD progression, and of patient morbidity and mortality in the hospital setting. To determine sex differences during injury and recovery we have generated a female and male renal ischemia/reperfusion injury (IRI) pig model, which represents a major cause of AKI. Although no differences were found in blood urea nitrogen (BUN) and serum creatinine (SCr) levels between both sexes, females exhibited higher mononuclear infiltrates at basal and recovery, while males showed more tubular damage at injury. Global transcriptomic analyses of kidney biopsies from our IRI pig model revealed a sexual dimorphism in the temporal regulation of genes and pathways relevant for kidney injury and repair, which was also detected in human samples. Enrichment analysis of gene sets revealed five temporal and four sexual patterns governing renal IRI and recovery. Overall, this study constitutes an extensive characterization of the time and sex differences occurring during renal IRI and recovery at gene expression level and offers a template of translational value for further study of sexual dimorphism in kidney diseases.

AUTHOR SUMMARY

Kidneys’ correct functioning is essential for optimal body homeostasis, being their basic functions blood filtration and excretion of wastes and toxins. Inherited or acquired conditions can cause renal dysfunction requiring renal replacement therapy, which will affect patients’ life quality and survival. A major cause of kidney failure is the renal ischemia/reperfusion injury (IRI), which occurs in many clinical situations like kidney transplantation or aortic aneurysm surgery. Interestingly, men are more susceptible to IRI than women, being women more protected against kidney injury. However, the genetics regulating these sex differences in injury and renal repair remained unknown.

Here, we provide a novel porcine model to study renal injury and recovery in both males and females. Using this model, we have identified the gene sets involved in renal injury and recovery processes. Moreover, global genetic analyses allowed us to discover the temporal and sex-dependent patterns that regulate those gene sets and, finally, kidney damage and repair. A relevant finding of our study is that males develop a feminized genetic profile during recovery, which may represent a survival mechanism to diminish the androgenic pro-damage effects on kidney cells. To sum up, our results provide novel sex-dependent targets to prevent renal injury and promote kidney recovery.