bluuebunny/crossref_metadata_2025 huggingface dataset:
CryoEM Reconstruction of Yeast ADP-Actin Filament at 2.5 Å resolution. A comparison with mammalian and avian F-actin
Authors: Stevenson Sarah R. , Tzokov Svetomir B. , Lahiri Indrajit , Ayscough Kathryn R. , Bullough Per A.
Published: September 2024

The core component of the actin cytoskeleton is the globular protein G-actin, which reversibly polymerises into filaments (F-actin). Budding yeast possesses a single actin which shares 87-89% sequence identity with vertebrate actin isoforms. Previous structural studies indicate very close overlap of main-chain backbones. Intriguingly however, substitution of yeastACT1with vertebrate β-cytoplasmic actin severely disrupts cell function and substitution with a skeletal muscle isoform is lethal. Here we report a 2.5 Å structure of budding yeast F-actin. Previously unresolved side-chain information now highlights four main differences in the comparison of yeast and vertebrate ADP F-actins: a more open nucleotide binding pocket; a more solvent exposed C-terminus; a rearrangement of intersubunit binding interactions in the vicinity of the D-loop and changes in the hydrogen bonding network in the vicinity of histidine 73 (yeast actin) and methyl-histidine 73 (vertebrate actin).
Development of a DNA-encoded library screening method DEL Zipper to empower the study of RNA-targeted chemical matter
Authors: Ma Zhongyao , Zou Bin , Zhao Jiannan , Zhang Rui , Zhu Qiaoqiao , Wang Xiaofeng , Xu Linan , Gao Xiang , Hu Xinyue , Feng Wei , Luo Wen , Wang Min , He Yunyun , Yu Zhifeng , Cui Weiren , Zhang Qi , Kuai Letian , Su Wenji
Published: September 2024

To date, RNA-targeted chemical matter is under explored due to a lack of robust screening assays. In this study, we present a novel RNA-targeted small molecule screening approach using a specialized DNA-encoded library (DEL). Our findings reveal that the specialized DEL library, called DEL Zipper, can significantly reduce single-stranded DNA-RNA region interaction signals during various kinds of RNA selection. By performing the selection against both G-quadruplex, we have identified novel hits that interact with RNA targets and the results are validated through binding. This study demonstrates that the DEL Zipper method is a robust screening assay that has potential for discovering small molecule ligands for diverse RNA targets.
Differential restriction of chikungunya virus in primary human cardiac endothelial cells occurs at multiple steps in the viral life cycle
Authors: Spector Sophie N. , Noval Maria G. , Stapleford Kenneth A.
Published: September 2024

Arthropod-borne viruses (arboviruses) constitute a significant ongoing public health threat, as the mechanisms of pathogenesis remain incompletely understood. Cardiovascular symptomatology is emerging as an important manifestation of arboviral infection. We have recently studied the cardiac tropism and mechanisms implicated in cardiac damage in mice for the alphavirus chikungunya virus (CHIKV), and we therefore sought to evaluate the cardiac tropism of other emerging alphaviruses and arboviruses. Using human primary cardiac cells, we found that arboviruses from diverse viral families were able to replicate within these cells. Interestingly, we noted that while the closely related alphavirus Mayaro virus (MAYV) could replicate to high titers in primary human cardiac microvascular endothelial cells, pulmonary, and brain endothelial cells, the Indian Ocean Lineage of CHIKV (CHIKV-IOL) was completely restricted in all endothelial cells tested. Upon further investigation, we discovered that this restriction occurs at both entry and egress stages. Additionally, we observed that compared to CHIKV, MAYV may antagonize or evade the innate immune response more efficiently in human cardiac endothelial cells to increase infection. Overall, this study explores the tropism of arboviruses in human primary cardiac cells and characterizes the strain-specific restriction of CHIKV-IOL in human endothelial cells. Further work is needed to understand how the differential restriction of alphaviruses in human endothelial cells impacts pathogenesis in a living model, as well as the specific host factors responsible.

Author Summary

Mosquito-borne viruses, such as those within the alphavirus genus, are an ongoing concern to human health globally. While we have recently begun to explore the mechanisms of CHIKV-induced pathology in the heart, little is known about other arboviruses and alphaviruses related to CHIKV. Here, we identified that multiple cardiac cell types are susceptible to infection by several arboviruses important to public health. Specifically, we noted differences in how two related viruses, CHIKV and MAYV, infect endothelial cells from multiple origins. This work highlights the potential for other emerging arboviruses, in addition to CHIKV, to directly infect cardiac tissue. Moreover, it emphasizes the ability for MAYV, an emerging virus that is less studied, to infect various endothelial cell types to high titers, suggesting the need for further research on MAYV pathogenesis. Finally, this research identifies differences in infection between individual strains of CHIKV, suggesting a finely tuned mechanism of restriction in endothelial cells that can be further explored, as well as the need to study use different viral strains for future alphavirus research.
Isolating selective from non-selective forces using site frequency ratios
Authors: Hey Jody , Pavinato Vitor
Published: September 2024

A new method is introduced for estimating the distribution of mutation fitness effects using site frequency spectra. Unlike previous methods, which make assumptions about non-selective factors, or that try to incorporate such factors into the underlying model, this new method mostly avoids non-selective effects by working with the ratios of counts of selected sites to neutral sites. An expression for the likelihood of a set of selected/neutral ratios is found by treating the ratio of two Poisson random variables as the ratio of two gaussian random variables. This approach also avoids the need to estimate the relative mutation rates of selected and neutral sites. Simulations over a wide range of demographic models, with linked selection effects show that the new SF-Ratios method performs well for statistical tests of selection, and it performs well for estimating the distribution of selection effects. Applications to two populations of Drosophila melanogaster reveal clear but very weak selection on synonymous sites. For nonsynonymous sites, selection was estimated to be far weaker than previous estimates for Drosophila populations.
Toxoplasma gondiitranscription factor AP2XII-8: a key regulator of G1 phase progression and parasite division
Authors: Shi Yuehong , Yang Qiang , He Yue , Song Xingju , Hu Dandan
Published: September 2024

The apicomplexan parasiteToxoplasma gondiican infect humans and virtually all warm-blooded animals worldwide, posing a significant threat to public health and being of veterinary importance. Acute infections are characterized by the fast replication of tachyzoites inside host cells. During this fast amplification process, gene expression is highly regulated by a series of regulatory networks. The G1 phase, which is usually conserved across species, is responsible for preparing the materials necessary for the next replicating cell cycle; however, few regulators have been identified at this stage. Here, we functionally characterized the C/G1 phase-expressed ApiAP2 transcription factor, TgAP2XII-8, inT. gondiitachyzoites. Conditional knockdown of TgAP2XII-8 leads to significant growth defects and asexual division disorders. Additionally, parasite cell cycle progression was disrupted following TgAP2XII-8 depletion, characterized by G1 phase arrest. RNA-seq and CUT&Tag experiments revealed that TgAP2XII-8 acts as an activator of ribosomal proteins expressed in the G1 phase. Moreover, TgAP2XII-8 binds to a specific DNA motif ([T/C]GCATGCA), which is abundant and conserved in the intergenic region of several other apicomplexans, possibly suggesting a broad and conserved role for this ApiAP2 in the Phylum of Apicomplexa. Our study reveals that TgAP2XII-8 acts as a critical C/G1 phase regulator, orchestrating the cell cycle inT. gondiitachyzoites. This study contributes to a broader understanding of the complexity of the parasite’s cell cycle and its key regulators.
Acacia gerrardii, a desert plant as a sustainable source of natural products with antimicrobial properties
Authors: Elkahoui Salem , Ghoniem Ahmed Eisa Mahmoud , Snoussi Mejdi , Barhoumi Zohaier , Badraoui Riadh
Published: September 2024

The aim of this study was to determine the chemical composition of methanolic extracts obtained from Acacia gerrardii (Benth.) leaves collected from the Hail Region of Saudi Arabia, to identify new bioactive compounds, and to their antioxidant activities, antibacterial and antifungal potentials, and their mineral composition. The methanolic extract showed the highest inhibition zone against methicillin-resistant Staphylococcus aureus (MRSA), Klebsiella pneumoniae and Escherichia coli. This activity is dose dependent. MIC and MBC values obtained against these bacteria in the presence of the extract were 3.125 and 50 mg/mL, respectively. The antifungal activity showed low effect against all the tested fungi (MIC: 6.25 mg/mL, MBC: 25 mg/mL). GC-MS analysis revealed the presence of 15 potent compounds. 4-O-Methylmannose was determined as the main compound of the methanolic extract. Omega 3 and 9 fatty acids, as well as diterpene and sterols were also characterized in the extract. Forty-nine (49) compounds were identified by HR-LC/MS, the main identified classes are amino acids, alkaloids, flavonol, flavones, flavonoids, dipeptide, indoles, stilbenes, pyrans, terpenes, glycosides, and sphingolipids. Moreover, the computational study revealed that the compounds identified in A. gerrardii extract binds to TyrRS from S. aureus (1JIJ), and the secreted aspartic proteinase 1 from C. albicans (2QZW) receptors with acceptable affinities and interactions. The rich phytochemical composition might be the cause of the outlined pharmacokinetic properties and molecular interactions associated antimicrobial effects, as reported by both computational and in vitro studies.
Lactobacillus crispatus S-layer proteins modulate innate immune response and inflammation in the lower female reproductive tract.
Authors: Decout Alexiane , Krasias Ioannis , Roberts Lauren , Gimeno Molina Belen , Charenton Chloe , Brown Romero Daniel , Tee Qiong Yu , Marchesi Julian R , Ng Sherrianne , Sykes Lynne , Bennett Phillip R , MacIntyre David Alan
Published: September 2024

Lactobacillus species dominance of the vaginal microbiome is a hallmark of vaginal health. Pathogen displacement of vaginal lactobacilli drives innate immune activation and mucosal barrier disruption which increases the risks of STI acquisition and, in pregnancy, of preterm birth. Using cell reporter systems, we describe differential TLR mediated activation of the proinflammatory transcription factor NF-κB by vaginal pathogens and commensals. Vaginal Lactobacillus strains associated with optimal health were shown to selectively interact with anti-inflammatory innate immune receptors whereas species associated with suboptimal health including L. iners and Gardnerella vaginalis interacted with both pro- and anti-inflammatory receptors. Anti-inflammatory action of L. crispatus was regulated by surface layer protein (SLPs)-mediated shielding of TLR ligands and selective interaction with anti-inflammatory receptor, DC-SIGN. In pregnant women, cervicovaginal SLPs were predominately associated with Lactobacillus-enriched microbiota. These data offer new mechanistic insights into how vaginal microbiota modulate host immune response and influences risk of preterm birth.
gsQTL: Associating genetic risk variants with gene sets by exploiting their shared variability
Authors: Bouland Gerard A. , Tesi Niccolò , Mahfouz Ahmed , Reinders Marcel J.T.
Published: September 2024

To investigate the functional significance of genetic risk loci identified through genome-wide association studies (GWASs), genetic loci are linked to genes based on their capacity to account for variation in gene expression, resulting in expression quantitative trait loci (eQTL). Following this, gene set analyses are commonly used to gain insights into functionality. However, the efficacy of this approach is hampered by small effect sizes and the burden of multiple testing. We propose an alternative approach: instead of examining the cumulative associations of individual genes within a gene set, we consider the collective variation of the entire gene set. We introduce the concept of gene set QTL (gsQTL), and show it to be more adept at identifying links between genetic risk variants and specific gene sets. Notably, gsQTL experiences less susceptibility to inflation or deflation of significant enrichments compared with conventional methods. Furthermore, we demonstrate the broader applicability of shared variability within gene sets. This is evident in scenarios such as the coordinated regulation of genes by a transcription factor or coordinated differential expression.
Preventive Effects of Pomegranate Seed Oil on Transient Middle Cerebral Artery Occlusion via the Keap1/Nrf2/NQO1 Pathway in the rats Cortex
Authors: Li Dongyang , Zhang Chaoying , Luo Qi , Li Man , Tian Meiqi , Jiao Hanyi , Xi Xiangwen , Weng Qifang
Published: September 2024

Ischemic stroke remains a pressing challenge that needs to be solved. Energy metabolic failure is a critical factor contributing to mitochondrial dysfunction and oxidative stress in the pathogenesis of brain ischemia, leading to the generation of excessive reactive oxygen species. Pomegranate seed oil (PSO) exhibits antioxidant properties; however, its protective effects against cerebral ischemia/reperfusion injury and the underlying mechanisms remain unclear. In this study, a transient middle cerebral artery occlusion (tMCAO) rat model was employed to simulate cerebral ischemia/reperfusion injury. We investigated the mechanisms by which different concentrations of PSO modulate oxidative damage caused by cerebral ischemia/reperfusion injury through the Keap1/Nrf2/NQO1 pathway in cortex. SD male rats were randomly divided into four groups: Control, tMCAO+NaCl, tMCAO+LO (low concentration of PSO), tMCAO+MO (medium concentration of PSO), and tMCAO+HO (high concentration of PSO). Our findings suggest that low concentration of PSO exerts neuroprotective effects by activating Nrf2 and NQO1, thereby reducing oxidative stress. Furthermore, LO significantly improved neurological scores and reduced neuronal edema. Additionally, the results demonstrated an increase in superoxide dismutase (SOD) levels and a decrease in malondialdehyde (MDA) levels. In contrast, MO and HO exhibited suboptimal effects. To sum up, these results indicate that PSO activates neuroprotective pathways against oxidative stress following cerebral ischemia/reperfusion injury via the Keap1/Nrf2/NQO1 pathway, providing novel insights into potential preventive therapies for cerebral ischemia/reperfusion.
Production of bioactive structural motifs from wheat arabinoxylan via colonic fermentation and enzymatic catalysis: evidence of interaction with toll-like receptors fromin vitro, in silicoand functional analysis
Authors: de A Guerreiro Caroline , Andrade Leandro AD , Fraga Layanne N , Marques Tatiana M , Prado Samira BR , Brummer Robert J , Nascimento João Roberto O , Castro-Alves Victor
Published: September 2024

It is well known that dietary fibers (DF) from plant-source foods can induce beneficial health effects through their physicochemical properties and utilization by the gut microbiota during fermentation, which is mainly explored with a focus on changes in the gut microbiota profile and the production of microbial-derived metabolites. Here, we characterized structural motifs (i.e., oligomers) produced during DF breakdown upon colonic fermentation and explored their interaction with toll-like receptors (TLRs) present on the surface of human intestinal and immune system cells. Firstly, a source of wheat arabinoxylan (WAX) was subjected toin vitrosimulation of human colonic fermentation, followed by characterization and quantification of WAX structural motifs to explore their dynamics throughout fermentation. The identified structural motifs were further produced through enzymatic catalysis of WAX using carbohydrate-active enzymes and fractionated into six well-defined fractions of arabinoxylans and linear xylans. These fractions of structural motifs were then tested for interaction with TLR2 and TLR4 using a reporter cell assay. Results revealed structure-dependent effects, primarily with inhibition of TLR2 and activation of TLR4 depending on the degree of polymerization and branching of WAX structural motifs. The role of the fine structure of WAX structural motifs was confirmed by molecular docking, which revealed that minor structural changes substantially influence the interaction between structural motifs and TLRs. The results fromin vitroandin silicostudies also support the hypothesis that the direct effects of oligomers and polysaccharides on cell receptors are likely the result of complex interactions involving multiple cell surface receptors. Finally, in addition to highlighting that direct effects of structural motifs might play an important role in the overall effects of DF, this work suggests that enzymatic-tailoring design of DF can be a potential tool for producing functional ingredients with specific effects on human health.
A high-speed OLED monitor for precise stimulation in vision, eye-tracking, and EEG research
Authors: Dimigen Olaf , Stein Arne
Published: September 2024

The recent introduction of organic light-emitting diode (OLED) monitors with refresh rates of 240 Hz or more opens new possibilities for their use as precise stimulation devices in vision research, experimental psychology, and electrophysiology. These affordable high-speed monitors, targeted at video gamers, promise several advantages over the cathode ray tube (CRT) and liquid crystal display (LCD) monitors commonly used in these fields. Unlike LCDs, OLED displays have self-emitting pixels that can show true black, resulting in superior contrast ratios, a broad color gamut, and good viewing angles. More importantly, the latest gaming OLEDs promise excellent timing properties with minimal input lags and rapid transition times. However, OLED technology also has potential drawbacks, notably Auto-Brightness Limiting (ABL) behavior, where the local luminance of a stimulus can change with the number of currently illuminated pixels. This study characterized a 240 Hz OLED monitor, the ASUS PG27AQDM, in terms of its timing properties, spatial uniformity, viewing angles, warm-up times, and ABL behavior. We also compared its responses to those of CRTs and LCDs. Results confirm the monitor’s excellent temporal properties with CRT-like transition times (around 0.3 ms), wide viewing angles, and decent spatial uniformity. Additionally, we found that ABL could be prevented with appropriate settings. We illustrate the monitor’s benefits in two time-critical paradigms: Rapid “invisible” flicker stimulation and the gaze-contingent presentation of stimuli during eye movements. Ourfindings suggest that the newest gaming OLEDs are precise and cost-effective stimulation devices for visual experiments that have several key advantages over CRTs and LCDs.
Identification of a fungal antibacterial endopeptidase that modulates immune responses
Authors: Machata Silke , Bertsche Ute , Hoffmann Franziska , Fattal Zaher M. , Kage Franziska , Flak Michal , Iliou Alexander N. J. , Hillmann Falk , von Eggeling Ferdinand , Slevogt Hortense , Brakhage Axel A. , Jacobsen Ilse D.
Published: September 2024

Aspergillus fumigatusis a saprophytic fungus dwelling in soil and on decaying plant material, but also an opportunistic pathogen in immunocompromised patients. In its environmental niche,A. fumigatusfaces competition from other microorganisms including bacteria. Here, we describe the discovery of the first secreted antibacterial protein inA. fumigatus. We identified a secreted fungal endopeptidase, designated CwhA, that cleaves peptidoglycan of Gram-positive bacteria at specific residues within the peptidoglycan stem peptide. Cleavage leads to bacterial lysis and the release of peptidoglycan cleavage products. Expression ofcwhAis induced by the presence of bacteria. Furthermore, CwhA is highly abundant in murine lungs during invasive pulmonary aspergillosis and peptidoglycan cleavage products generated by CwhA stimulate cytokine production of human immune cells. Although CwhA does not affect human cells directly, this novel player in fungal-bacterial interactions could affectA. fumigatusinfections by inhibiting Gram-positive bacteria in its vicinity, and modulating the immune system.
Muscle spindles provide flexible sensory feedback for movement sequences
Authors: Olson William P. , Chokshi Varun B. , Kim Jeong Jun , Cowan Noah J. , O’Connor Daniel H.
Published: September 2024

Sensory feedback is essential for motor performance and must adapt to task demands. Muscle spindle afferents (MSAs) are a major primary source of feedback about movement, and their responses are readily modulated online by gain-controller fusimotor neurons and other mechanisms. They are therefore a powerful site for implementing flexible sensorimotor control. We recorded from MSAs innervating the jaw musculature during performance of a directed lick sequence task. Jaw MSAs encoded complex jaw–tongue kinematics. However, kinematic encoding alone accounted for less than half of MSA spiking variability. MSA coding of kinematics changed based on sequence progression (beginning, middle, or end of the sequence, or reward consumption), suggesting that MSAs are flexibly tuned across the task. Dynamic control of incoming feedback signals from MSAs may be a strategy for adaptable sensorimotor control during performance of complex behaviors.
Community-Based Entomological Surveillance and Control of Vector-Borne Diseases: A Scoping Review
Authors: Eastman P. , Awolola T.S. , Yoshimizu M. , Govellla N. , Chaki P. , Zohdy S.
Published: September 2024

Community-based surveillance and control methods (CBMs) present opportunities to decentralize surveillance and control efforts while simultaneously enhancing community education, leadership, and participation in the fight against vector-borne diseases (VBDs). A scoping review was conducted to describe how CBMs are being utilized currently to combat malaria, dengue fever, Chagas disease, tick-borne diseases (TBDs) and other mosquito-borne diseases (MBD) exclusive of dengue and malaria, and to overall highlight key approaches, lessons learned, potential challenges, and recommendations. A total of 304 potential publications were identified among which 82 met the inclusion criteria. This scoping review highlighted the following benefits to CBMs: cost savings, increased sustainability, increased community knowledge, human behavior changes, increased surveillance coverage, ease in deployment, and the creation of larger, more diverse entomological datasets. Potential challenges highlighted include: participant retention and motivation, participant recruitment and incentives, continued governmental support, data quality, and collaboration with local municipal authorities. CBMs are commonly and successfully used in vector surveillance and control systems, but the chosen vector management method varies by vector-borne disease and region of the world. Additional research is needed to support the implementation of CBMs including cost-effectiveness studies and those studies with negative outcomes. Taken together, this scoping review highlights key aspects, potential challenges, and benefits of CBMs, and outlines potential future directions for incorporating CBMs into VBD control and elimination programming, and potential for community based integrated vector management (IVM) approaches.
Mild activation of the mitochondrial unfolded protein response increases lifespan without increasing resistance to stress
Authors: Di Pede Alexa , Ko Bokang , AlOkda Abdelrahman , Tamez González Aura A. , Van Raamsdonk Jeremy M.
Published: September 2024

The mitochondrial unfolded protein response (mitoUPR) is a stress response pathway that responds to mitochondrial insults by altering gene expression to recover mitochondrial homeostasis. The mitoUPR is mediated by the stress-activated transcription factor ATFS-1. Constitutive activation of ATFS-1 increases resistance to exogenous stressors but paradoxically decreases lifespan. In this work, we determined the optimal levels of expression of activated ATFS-1 with respect to lifespan and resistance to stress by treating constitutively-activeatfs-1(et17)worms with different concentrations of RNA interference (RNAi) bacteria targetingatfs-1.We observed the maximum lifespan ofatfs-1(et17)worms at full-strengthatfs-1RNAi, which was significantly longer than wild-type lifespan. Under the conditions of maximum lifespan,atfs-1(et17)worms did not show enhanced resistance to stress, suggesting a trade-off between stress resistance and longevity. The maximum resistance to stress inatfs-1(et17)worms occurred on empty vector (0%atfs-1knockdown). Under these conditions,atfs-1(et17)worms are short-lived. This indicates that constitutive activation of ATFS-1 can increase lifespan or enhance resistance to stress but not both, at the same time. Finally, we determined the timing requirements for ATFS-1 to affect lifespan. We found that knocking downatfs-1expression only during development is sufficient to extendatfs-1(et17)lifespan, while adult-only knockdown has no effect. Overall, these results demonstrate that constitutively active ATFS-1 can extend lifespan when expressed at low levels and that this lifespan extension is not dependent on the ability of ATFS-1 to enhance resistance to stress.
Ripply3overdosage induces mid-face shortening throughTbx1downregulation in Down syndrome models
Authors: Ahumada Saavedra José Tomás , Chevalier Claire , Bloch Zupan Agnes , Herault Yann
Published: September 2024

The most frequent and unique features of Down syndrome (DS) are learning disability and ucraniofacial (CF) dysmorphism. The DS-specific CF features are an overall reduction in head dimensions (microcephaly), relatively wide neurocranium (brachycephaly), reduced mediolaterally orbital region, reduced bizygomatic breadth, small maxilla, small mandible, and increased individual variability. Until now, the cellular and molecular mechanisms underlying the specific craniofacial phenotype have remained poorly understood. Investigating a new panel of DS mouse models with different segmental duplications on mouse chromosome 16 in the region homologous to human chromosome 21, we identified new regions and the role of two candidate gene for DS-specific CF phenotypes. First, we confirmed the role ofDyrk1ain the neurocranium brachycephaly. Then, we identified the role of the transcription factorRipply3overdosage in midface shortening through the downregulation ofTbx1, another transcription factor involved in the CF midface phenotype encountered in DiGeorge syndrome. This last effect occurs during branchial arches development through a reduction in cell proliferation. Our findings define a new dosage-sensitive gene responsible for the DS craniofacial features and propose new models for rescuing all aspects of DS CF phenotypes. This data may also provide insights into specific brain and cardiovascular phenotypes observed in DiGeorge and DS models, opening avenues for potential targeted treatment to soften craniofacial dysmorphism in Down syndrome.
Practical media formulations for rapid growth ofLactobacillus inersand other vaginal bacteria
Authors: Serrador Daniella , Campbell Jhenielle R. , Cheung Dorothy , Shefraw Gelila , Kaul Rupert , Navarre William W.
Published: September 2024

Vaginal microbiome composition is closely tied to host health. A microbiome dominated by specific anaerobes (e.g.,Gardnerella vaginalis) is termed bacterial vaginosis (BV) and is associated with negative health outcomes, while colonization byLactobacillusspecies is thought to protect against BV. However, the role of the speciesLactobacillus inersin vaginal health is controversial, with evidence that some strains may not protect against BV while others do. To better characterizeL. inersstrains, their interactions with vaginal bacteria and human cells need to be investigatedin vitro, but this has been impeded by the lack of liquid media for rapidL. inersgrowth. We have developed three liquid media formulations forL. inersgrowth: Serrador’s Lactobacilli-adapted Iscove’s Medium (SLIM) which leads to robustL. inersgrowth, a vaginally adapted version of SLIM (SLIM-V) and a chemically defined medium (SLIM-CD). SLIM and SLIM-V lead to dramatically improvedL. inersgrowth compared to previously published formulations and support growth of other vaginal bacteria, includingL. crispatus, L. jensenii, L. gasseriandG. vaginalis. SLIM-CD leads to slower growth but could prove useful for characterizingL. inersnutrient requirements or metabolite production. A modified version of SLIM-V supports growth of human cervical epithelial cells and provides a base for future co-culture work. Here, we present the formulations of SLIM, SLIM-V and SLIM-CD, and compare the growth of bacterial strains and human cells in the media.
A brain-body feedback loop driving HPA-axis dysfunction in breast cancer
Authors: Gomez Adrian , Wu Yue , Zhang Chao , Boyd Leah , Wee Tse-Luen , Gewolb Joseph , Amor Corina , Cheadle Lucas , Borniger Jeremy C.
Published: September 2024

Breast cancer patients often exhibit disrupted circadian rhythms in circulating glucocorticoids (GCs), such as cortisol. This disruption correlates with reduced quality of life and higher cancer mortality1–3. The exact cause of this phenomenon — whether due to treatments, stress, age, co-morbidities, lifestyle factors, or the cancer itself remains unclear. Here, we demonstrate that primary breast cancer alone blunts host GC rhythms by disinhibiting neurons in the hypothalamus, and that circadian phase-specific neuromodulation of these neurons can attenuate tumor growth by enhancing anti-tumor immunity. We find that mice with mammary tumors exhibit blunted GC rhythms before tumors are palpable, alongside increased activity in paraventricular hypothalamic neurons expressing corticotropin-releasing hormone (i.e., PVNCRHneurons). Tumor-bearing mice have fewer inhibitory synapses contacting PVNCRHneurons and reduced miniature inhibitory post-synaptic current (mIPSC) frequency, leading to net excitation. Tumor-bearing mice experience impaired negative feedback on GC production, but adrenal and pituitary gland functions are largely unaffected, indicating that alterations in PVNCRHneuronal activity are likely a primary cause of hypothalamic-pituitary-adrenal (HPA) axis dysfunction in breast cancer. Using chemogenetics (hM3Dq) to stimulate PVNCRHneurons at different circadian phases, we show that stimulation just before the light-to-dark transition restores normal GC rhythms and reduces tumor progression. These mice have significantly more effector T cells (CD8+) within the tumor than non-stimulated controls, and the anti-tumor effect of PVNCRHneuronal stimulation is absent in mice lacking CD8+ T cells. Our findings demonstrate that breast cancer distally regulates neurons in the hypothalamus that control output of the HPA axis and provide evidence that therapeutic targeting of these neurons could mitigate tumor progression.
CD11B+CD36+cells are bone anabolic macrophages that limit age-associated bone loss
Authors: Koroth Jinsha , Karkache Ismael Y. , Vu Elizabeth K. , Mansky Kim C. , Bradley Elizabeth W.
Published: September 2024

Disruptions in the bone remodeling cycle that occur with increasing age lead to degeneration of the skeleton and increased risk of fragility fractures. Our understanding of how bone remodeling within cortical bone is controlled and altered with age in males and females is limited. Here, we generated bone marrow chimeric mice to understand the impacts of age and sex on bone remodeling. We demonstrate that transplantation of aged male or female bone marrow into young, lethally irradiated male hosts unexpectedly enhances cortical bone mass without impacting cancellous bone. Our single cell RNA-sequencing data show that mice reconstituted with aged bone marrow exhibited subsets of cells marked by CD11B/CD36 expression that demonstrate enhanced production of anabolic cytokines as compared to young counterparts, and that these myeloid subsets exist under conditions of normal physiology in aged mice. Importantly, CD11B+CD36+cells do not differentiate into osteoclasts in vitro, and CD36 does not mark TRAP+ cells in vivo. Instead, CD36+cells localize to resorption sites, including within cortical bone defects, suggesting their involvement in cortical bone remodeling and healing. CD11B+CD36+cells also express elevated levels of bone anabolic WNT ligands, especially Wnt6. In functional assays, we demonstrate that soluble factors produced by CD11B+CD36+cells enhance osteoblast progenitor commitment, mineralization, and activation of WNT signaling in vitro. Moreover, CD11B/CD36 exquisitely mark a subset of anabolic myeloid cells within human bone marrow. In conclusion, our studies identified a novel population of aged macrophages that limit cortical bone loss.
tRNA lysidinylation is essential for the minimal translation system found in the apicoplast ofPlasmodium falciparum
Authors: Elahi Rubayet , Prigge Sean T.
Published: September 2024

For decades, researchers have sought to define minimal genomes to elucidate the fundamental principles of life and advance biotechnology. tRNAs, essential components of this machinery, decode mRNA codons into amino acids. The apicoplast of malaria parasites encodes 25 tRNA isotypes in its organellar genome - the lowest number found in known translation systems. Efficient translation in such minimal systems depends heavily on post-transcriptional tRNA modifications, especially at the wobble anticodon position. Lysidine modification at the wobble position (C34) of tRNACAUdistinguishes between methionine (AUG) and isoleucine (AUA) codons, altering the amino acid delivered by this tRNA and ensuring accurate protein synthesis. Lysidine is formed by the enzyme tRNA isoleucine lysidine synthetase (TilS) and is nearly ubiquitous in bacteria and essential for cellular viability. We identified a TilS ortholog (PfTilS) located in the apicoplast ofPlasmodium falciparumparasites. By complementingPfTilS with a bacterial ortholog, we demonstrated that the lysidinylation activity ofPfTilS is critical for parasite survival and apicoplast maintenance, likely due to its impact on apicoplast protein translation. Our findings represent the first characterization of TilS in an endosymbiotic organelle, advancing eukaryotic organelle research and our understanding of minimal translational machinery. Due to the absence of lysidine modifications in humans, this research also exposes a potential vulnerability in malaria parasites that could be targeted by antimalarial strategies.

Significance

In recent decades, synthetic biologists have sought the minimal cellular components required for life, focusing on simpler systems for easier modeling. The apicoplast organelle of malaria parasites, with only 25 tRNA isotypes, contains the smallest known complete tRNA set, even smaller than in synthetic organisms. This makes it an ideal model for studying minimal translational machinery, where tRNAs depend on post-transcriptional modifications for efficient protein translation. A key modification, lysidine, is crucial for decoding isoleucine and methionine. This study describes a tRNA-isoleucine lysidine synthetase (TilS) enzyme, essential for apicoplast protein translation. These findings have implications for understanding eukaryotic organelles and minimal translation machinery. Additionally, the absence of lysidine in humans suggests a potential target for antimalarial strategies.
Operant alcohol self-administration targets GluA2-containing AMPA receptor expression and synaptic activity in the nucleus accumbens in a manner that drives the positive reinforcing properties of the drug
Authors: Faccidomo Sara , Saunders Briana L , May Ashley M. , Eastman Vallari R. , Kim Michelle , Taylor Seth M. , Hoffman Jessica L. , McElligott Zoé A , Hodge Clyde W
Published: September 2024

Rationale:The positive reinforcing effects of alcohol (ethanol) drive its repetitive use and contribute to alcohol use disorder (AUD). Ethanol alters the expression of glutamate AMPA receptor (AMPAR) subunits in reward-related brain regions, but the extent to which this effect regulates ethanol’s reinforcing properties is unclear.Objective:This study investigates whether ethanol self-administration changes AMPAR subunit expression and synaptic activity in the nucleus accumbens core (AcbC) to regulate ethanol’s reinforcing effects in male C57BL/6J mice.Results:Sucrose-sweetened ethanol self-administration (0.81 g/kg/day) increased AMPAR GluA2 protein expression in the AcbC, without effect on GluA1, compared to sucrose-only controls. Infusion of myristoylated Pep2m in the AcbC, which blocks GluA2 binding to N-ethylmaleimide-sensitive fusion protein (NSF) and reduces GluA2-containing AMPAR activity, reduced ethanol-reinforced responding without affecting sucrose-only self-administration or motor activity. Antagonizing GluA2-lacking AMPARs, through AcbC infusion of NASPM, had no effect on ethanol self-administration. AcbC neurons receiving projections from the basolateral amygdala (BLA) showed increased sEPSC area under the curve (a measurement of charge transfer) and slower decay kinetics in ethanol self-administering mice as compared to sucrose. Optogenetic activation of these neurons revealed an ethanol-enhanced AMPA/NMDA ratio and significantly reduced paired-pulse ratio, suggesting elevated GluA2 contributions specifically within the BLA→AcbC pathway.Conclusions:Ethanol use upregulates GluA2 protein expression in the AcbC and AMPAR synaptic activity in AcbC neurons receiving BLA projections and enhances synaptic plasticity directly within the BLA→AcbC circuit. GluA2-containing AMPAR activity in the AcbC regulates the positive reinforcing effects of ethanol through an NSF-dependent mechanism, highlighting a potential therapeutic target in AUD.
A molecular switch for stress-induced activation of retrograde mitochondrial transport
Authors: Gladkova Christina , Paez-Segala Maria G , Grant William P , Myers Samuel A , Wang Yuxiao , Vale Ronald D
Published: September 2024

The cellular distribution of mitochondria in response to stress and local energy needs is governed by the relative activities of kinesin and dynein. The mechanism for switching between these two opposite polarity microtubule motors remains unknown. Here, we coupled a novel cellular synthetic cargo transport assay with AlphaFold2-guided mutagenesis to identify a regulatory helix in the mitochondrial adaptor protein (TRAK) that mediates switching between kinesin- and dynein-driven transport. Differences in the helix sequence explain why two near-identical TRAK isoforms transport mitochondria in predominantly opposite directions. Phosphorylation of the regulatory helix by stress-activated kinases causes the activation of dynein and dissociation of kinesin. Our results reveal a molecular mechanism for coordinating the directional transport of mitochondria in response to intracellular signals.
Age-related decline of PKA-RIIβ level in SNc dopaminergic neurons underlies PD pathogenesis
Authors: Zhao Yun , Wang Bingwei , Zhao Miao , Liu Jiarui , Hou Yujia , Wei Shizhang , Xiong Changxian , Li Daotong , Zheng Ruimao
Published: September 2024

The cyclic-AMP dependent protein kinase A (Protein kinase A, PKA) regulates dopaminergic function in the substantia nigra pars compacta (SNc). However, whether PKA is involved in the pathogenesis of Parkinson’s disease (PD) is unknown. Here, by collecting and analyzing the current worldwide SNc transcriptomic datasets of PD patients, we found a decline of PKA-RIIβ subunit level in the SNc of PD patients. The decreased PKA-RIIβ subunit level was positively correlated with decreased dopamine synthesis and increased oxidative stress in the SNc of PD patients. PKA-RIIβ subunit is expressed in the striatum and the SNc. PKA-RIIβ gene knockout mice (RIIβ-KO) showed a age-related parkinsonism at 12 months of age. Using Cre-LoxP system, we observed that RIIβ re-expression in the SNc dopaminergic neurons rescued parkinsonism of RΙΙβ-KO mice. RIIβ re-expression in striatal neurons did not affect parkinsonism of RΙΙβ-KO mice. The spontaneous parkinsonism could be developed in 12-month-old SNc dopaminergic neuron-specific RIIβ-deficient mice. Single-nucleus RNA sequencing revealed decreased PKA activity, reduced dopamine synthesis and raised oxidative stress in the SNc dopaminergic neurons of RIIβ-KO mice. Adeno-associated virus (AAV)-mediated gene therapy targeting PKA-RΙΙβ in the SNc dopaminergic neurons rescued parkinsonism in PD mouse model. Taken together, these findings indicate that PKA-RIIβ may be a key factor of human genetic etiologies of PD. The therapy targeting PKA-RIIβ in the SNc dopaminergic neurons may be promising for PD treatment.
Mayo Normative Studies: regression-based normative data for remote self-administration of the Stricker Learning Span, Symbols Test and Mayo Test Drive Screening Battery Composite and validation in individuals with Mild Cognitive Impairment and dementia
Authors: Stricker Nikki H. , Frank Ryan D. , Boots Elizabeth A. , Fan Winnie Z. , Christianson Teresa J. , Kremers Walter K. , Stricker John L. , Machulda Mary M. , Fields Julie A. , Lucas John A. , Hassenstab Jason , Aduen Paula A. , Day Gregory S. , Graff-Radford Neill R. , Jack Clifford R. , Graff-Radford Jonathan , Petersen Ronald C.
Published: September 2024

Objective

Few normative data for unsupervised, remotely-administered computerized cognitive measures are available. We examined variables to include in normative models for Mayo Test Drive (a multi-device remote cognitive assessment platform) measures, developed normative data, and validated the norms.

Method

1240 Cognitively Unimpaired (CU) adults ages 32-100-years (96% white) from the Mayo Clinic Study of Aging and Mayo Alzheimer’s Disease Research Center with Clinical Dementia Rating® of 0 were included. We converted raw scores to normalized scaled scores and derived regression-based normative data adjusting for age, age2, sex and education (base model); alternative norms are also provided (age+age2+sex; age+age2). We assessed additional terms using ana prioricut-off of 1% variance improvement above the base model. We examined low test performance rates (<-1 standard deviation) in independent validation samples (n=167 CU, n=64 mild cognitive impairment (MCI), n=14 dementia). Rates were significantly different when 95% confidence intervals (CI) did not include the expected 14.7% base rate.

Results

No model terms met thea prioricut-off beyond the base model, including device type, response input source (e.g., mouse, etc.) or session interference. Norms showed expected low performance rates in CU and greater rates of low performance in MCI and dementia in independent validation samples.

Conclusion

Typical normative models appear appropriate for remote self-administered MTD measures and are sensitive to cognitive impairment. Device type and response input source did not explain enough variance for inclusion in normative models but are important for individual-level interpretation. Future work will increase inclusion of individuals from under-represented groups.
Human genetic evidence to inform clinical development of interleukin-6 signaling inhibition for abdominal aortic aneurysm
Authors: Burgess Stephen , Cronjé Héléne T. , deGoma Emil , Chyung Yung , Gill Dipender
Published: September 2024

Background

Abdominal aortic aneurysm (AAA) represents a significant cause of mortality, yet no medical therapies have proven efficacious. The aim of the current study was to leverage human genetic evidence to inform clinical development of interleukin-6 (IL6) signaling inhibition for treatment of AAA.

Methods

We focused on rs2228145, a missense variant in theIL6Rgene region whose associations are expressed per additional copy of the C allele, corresponding to the genetically-predicted effect of IL6 signaling inhibition. We consider genetic associations with AAA risk in the AAAgen consortium (39,221 cases, 1,086,107 controls) and UK Biobank (2215 cases, 365,428 controls). To validate against known effects of IL6 signaling inhibition, we present associations with rheumatoid arthritis, polymyalgia rheumatica, and severe COVID-19. To explore mechanism specificity, we present associations with thoracic aortic aneurysm, intracranial aneurysm, and coronary artery disease. We further evaluated associations with measures of the abdominal aorta in UK Biobank, and explored genetic associations in clinically-relevant subgroups of the population.

Results

We observed strong genetic associations with AAA risk in the AAAgen consortium and in UK Biobank: odds ratio (OR) 0.91 (95% confidence interval [CI]: 0.90 to 0.92, p = 4×10-30) and OR 0.90 (95% CI: 0.84, 0.96, p=0.0007), respectively. The association with AAA risk in UK Biobank was linear in the number of minor alleles: OR 0.91 (95% CI: 0.83, 1.00) in heterozygotes and OR 0.80 (95% CI: 0.71, 0.92) in minor homozygotes. The association was similar for fatal AAA, but with greater uncertainty due to the lower number of events. The association with AAA was of greater magnitude than associations with coronary artery disease and even rheumatologic disorders for which IL6 inhibitors have been approved. No strong associations were observed with thoracic aortic aneurysm, intracranial aneurysm, or abdominal aorta diameter in the general population without AAA. Associations attenuated towards the null in populations with concomitant inflammatory or connective tissue disease.

Conclusions

This drug target Mendelian randomization study supports that IL6 signaling inhibition will be efficacious for treating AAA, but not other types of aneurysmal disease. These findings serve to help inform clinical development of IL6 signaling inhibition for AAA treatment.
Dynamic network regulating phosphatidic acid homeostasis revealed using membrane editing coupled to proximity labeling
Authors: Tei Reika , Baskin Jeremy M.
Published: September 2024

Cellular lipid metabolism is subject to strong homeostatic regulation, but players involved in and mechanisms underlying these pathways remain mostly uncharacterized. Here, we develop and exploit a “Feeding–Fishing” approach coupling membrane editing using optogenetic lipid-modifying enzymes (feeding) with organelle membrane proteomics via proximity labeling (fishing) to elucidate molecular players and pathways involved in homeostasis of phosphatidic acid (PA), a multifunctional lipid central to glycerolipid metabolism. By performing proximity biotinylation using a membrane-tethered TurboID alongside membrane editing to selectively deliver phosphatidic acid to the same membrane, we identified numerous PA-metabolizing enzymes and lipid transfer proteins enriched in and depleted from PA-fed membranes. Subsequent mechanistic analysis established that PA homeostasis in the cytosolic leaflets of the plasma membrane and of lysosomes is governed by a select subset of PA metabolic pathways and, via divergent molecular mechanisms, several members of the lipid transfer protein superfamily capable of mediating interorganelle lipid transport. More broadly, the interfacing of membrane editing with organelle membrane proteomics using proximity labeling represents a powerful and generalizable strategy for revealing mechanisms governing lipid homeostasis.