bluuebunny/crossref_metadata_2025 huggingface dataset:
N-cadherin facilitates trigeminal sensory neuron outgrowth and target tissue innervation
Authors: Halmi Caroline A. , Leonard Carrie E. , McIntosh Alec T. , Taneyhill Lisa A.
Published: May 2024

During embryonic development, diverse cell types coordinate to form functionally complex tissues. Exemplifying this process, the trigeminal ganglion emerges from the condensation of two distinct precursor cell populations, cranial placodes and neural crest, with neuronal differentiation of the former preceding the latter. While its dual cellular origin has been understood for decades, the molecules orchestrating trigeminal ganglion formation remain relatively obscure. Initial assembly of the trigeminal ganglion is mediated by cell adhesion molecules, including neural cadherin (N-cadherin), which is first expressed by placodal neurons and is required for their proper condensation with other neurons and neural crest cells. Axon outgrowth first occurs from placodal neurons, but as gangliogenesis proceeds, neural crest cells also differentiate into N-cadherin-expressing neurons, and together both extend axons toward target tissues. However, a role for N-cadherin in regulating axon outgrowth and innervation of target tissues by trigeminal neurons has not been explored. To this end, we depleted N-cadherin from chick trigeminal placode cells and observed decreases in trigeminal ganglion size, nerve growth, and target innervationin vivo, phenotypes that could only partially be attributed to increased apoptosis early in gangliogenesis. Accordingly, neurite number and branching of neural crest-derived neurons was decreasedin vitroin response to N-cadherin knockdown in placode cells, providing a novel non-cell autonomous explanation for these morphological changes. Inhibiting N-cadherin-mediated adhesion with a function-blocking antibody prevented axon extension in most, but not all, placode-derived trigeminal neuronsin vitro, indicating potential unique requirements for N-cadherin in various neuronal subtypes. Collectively, these findings reveal persistent cell autonomous and non-cell autonomous functions for N-cadherin, thus highlighting the critical role of N-cadherin in mediating reciprocal interactions between neural crest and placode neuronal derivatives during trigeminal ganglion development.
Sphingosine-1-phosphate signaling inhibition suppresses Th1-like Treg generation by reversing mitochondrial uncoupling
Authors: Coulombeau Rachel , Selck Claudia , Giang Nicolas , Al-Mohammad Abdulrahman , Ng Natalie , Maher Allison K , Argüello Rafael , Scalfari Antonio , Varley James , Nicholas Richard , Dominguez-Villar Margarita
Published: May 2024

Inflammatory environments induce the generation of dysfunctional IFNγ+T-bet+FOXP3+Th1-like Tregs, which show defective function and are found in autoimmune conditions including multiple sclerosis (MS). The pathways that control the generation of Th1-like Tregs are not well understood. Sphingosine-1-phosphate (S1P) signaling molecules are upregulated in Th1-like Tregs, andin vivoS1P inhibition with Fingolimod (FTY720) inhibits the expression of genes responsible for Treg plasticity in MS patients. However, the underlying mechanisms are unknown. Here we show that S1P signaling inhibition by FTY720 inhibits the generation of Th1-like Tregs and rescues their suppressive function. These effects are mediated by a decrease in mTORC1 signaling and reversal of the mitochondrial uncoupling that Tregs undergo during their reprogramming into Th1-like Tregsin vitro. Finally, these results are validated inin vivogenerated Th1-like Tregs, as the Tregs from MS patients treated with FTY720 display decreased Th1-like Treg frequency, increased suppressive function, and mitochondrial metabolism rebalance. These results highlight the involvement of mitochondrial uncoupling in Treg reprogramming and identify S1P signaling inhibition as a target to suppress the generation of dysfunctional Th1-like Tregs.
The insulin / IGF axis is critically important controlling gene transcription in the podocyte
Authors: Hurcombe Jennifer A. , Dayalan Lusyan , Barrington Fern , Burdet Frédéric , Ni Lan , Coward Joseph T. , Brinkkoetter Paul T. , Holzenberger Martin , Jeffries Aaron , Oltean Sebastian , Welsh Gavin I. , Coward Richard J.M.
Published: May 2024

Signalling to the podocyte via the structurally related insulin receptor (IR) or insulin-like growth factor 1 receptor (IGF1) is important for podocyte function. This study sought to elucidate the compound role of the insulin/IGF1 axis in podocytes using transgenic mice and cell culture models deficient in both receptors.

Podocyte specific IR/IGF1R knockdown mice developed a severe kidney phenotype with albuminuria, glomerulosclerosis and renal failure with death occurring in some mice between 4 and 24 weeks. Simultaneous knockdown of both receptors in cultured podocytes resulted in >50% cell death by 7 days.

Proteomic analysis revealed a striking downregulation of spliceosome-related proteins in IR/IGF1R knockdown podocytes with long-read RNA sequence data indicating an increased fraction of transcripts with intron retention/premature termination codons in these cells.

This work underlines the critical importance of podocyte insulin/IGF signalling revealing a novel role for this extrinsic hormonal signalling axis in regulating gene transcription.
Cigarette smoke impairs pulmonary vascular function through nAChR activation
Authors: Munar-Rubert O , Andreu-Martínez R , Rodríguez-Pérez J , López N , Barreira B , Fernández-Malavé E , Peces-Barba G , Muñoz-Calleja C , Cogolludo A , Calzada MJ
Published: May 2024

Tobacco smoke is the main risk factor for the development of chronic obstructive pulmonary disease (COPD), a major health concern worldwide. Despite current therapies alleviate symptoms; there remain some limitations in the efficacy of treatments to curb COPD and its cardiovascular morbidities, particularly pulmonary hypertension. Our previous studies demonstrate that cigarette smoke (CS) has direct effects on pulmonary vascular tone homeostasis and contribute to pulmonary arterial dysfunction. This is in part due to altered activity of the voltage-dependent K+channel, and to an exacerbated oxidative stress promoting a switch in the sGCs redox state. However, further characterization of the molecular basis of CS-mediated PA dysfunction is needed for more effective targeted treatment and prevention. Our current studies explored these molecular pathways and specifically addressed their contribution to the cellular contractile apparatus within pulmonary arteries. Our results proved deleterious effects on the contractile machinery of pulmonary artery smooth muscle cells. Increased oxidative stress and calcium dysregulation resulting from the activation of acetylcholine receptors (nAChR) in the pulmonary artery led to the manifestation of these effects. This groundbreaking discovery unveiled, for the first time, the expression of these receptors in human pulmonary arteries. Furthermore, we proved that inhibitors directed at these receptors demonstrate efficacy in alleviating various harmful effects of smoking and safeguarding pulmonary artery function from damage. These discoveries hold significant clinical implications, as they suggest that treatment with nAChR-targeted inhibitors could constitute a viable therapeutic option for COPD-related pulmonary hypertension in patients who do not respond to conventional medication.
Cell type-specific network analysis in Diversity Outbred mice identifies genes potentially responsible for human bone mineral density GWAS associations
Authors: Dillard Luke J , Calabrese Gina M , Mesner Larry D , Farber Charles R
Published: May 2024

Genome-wide association studies (GWASs) have identified many sources of genetic variation associated with bone mineral density (BMD), a clinical predictor of fracture risk and osteoporosis. Aside from the identification of causal genes, other difficult challenges to informing GWAS include characterizing the roles of predicted causal genes in disease and providing additional functional context, such as the cell type predictions or biological pathways in which causal genes operate. Leveraging single-cell transcriptomics (scRNA-seq) can assist in informing BMD GWAS by linking disease-associated variants to genes and providing a cell type context for which these causal genes drive disease. Here, we use large-scale scRNA-seq data from bone marrow–derived stromal cells cultured under osteogenic conditions (BMSC-OBs) from Diversity Outbred (DO) mice to generate cell type-specific networks and contextualize BMD GWAS-implicated genes. Using trajectories inferred from the scRNA-seq data, we identify networks enriched with genes that exhibit the most dynamic changes in expression across trajectories. We discover 21 network driver genes, which are likely to be causal for human BMD GWAS associations that colocalize with expression/splicing quantitative trait loci (eQTL/sQTL). These driver genes, includingFgfrl1andTpx2,along with their associated networks, are predicted to be novel regulators of BMD via their roles in the differentiation of mesenchymal lineage cells. In this work, we showcase the use of single-cell transcriptomics from mouse bone-relevant cells to inform human BMD GWAS and prioritize genetic targets with potential causal roles in the development of osteoporosis.
Sequential deactivation across the thalamus-hippocampus-mPFC pathway during loss of consciousness
Authors: Chen Xiaoai , Cramer Samuel R. , Chan Dennis C.Y. , Han Xu , Zhang Nanyin
Published: May 2024

How consciousness is lost in states such as sleep or anesthesia remains a mystery. To gain insight into this phenomenon, we conducted concurrent recordings of electrophysiology signals in the anterior cingulate cortex and whole-brain functional magnetic resonance imaging (fMRI) in rats exposed to graded propofol, undergoing the transition from consciousness to unconsciousness. Our results reveal that upon the loss of consciousness (LOC), as indicated by the loss of righting reflex, there is a sharp increase in low-frequency power of the electrophysiological signal. Additionally, simultaneously measured fMRI signals exhibit a cascade of deactivation across a pathway including the hippocampus, thalamus, and medial prefrontal cortex (mPFC) surrounding the moment of LOC, followed by a broader increase in brain activity across the cortex during sustained unconsciousness. Furthermore, sliding window analysis demonstrates a temporary increase in synchrony of fMRI signals across the hippocampus-thalamus-mPFC pathway preceding LOC. These data suggest that LOC might be triggered by sequential activities in the hippocampus, thalamus and mPFC, while wide-spread activity increases in other cortical regions commonly observed during anesthesia-induced unconsciousness might be a consequence, rather than a cause of LOC. Taken together, our study identifies a cascade of neural events unfolding as the brain transitions into unconsciousness, offering critical insight into the systems-level neural mechanisms underpinning LOC.
Determination of metal ion transport rate of human ZIP4 using stable zinc isotopes
Authors: Jiang Yuhan , MacRenaris Keith , O’Halloran Thomas V. , Hu Jian
Published: May 2024

The essential microelement zinc is absorbed in the small intestine mainly by the zinc transporter ZIP4, a representative member of the Zrt/Irt-like protein (ZIP) family. ZIP4 is reportedly upregulated in many cancers, making it a promising oncology drug target. To date, there have been no reports on the turnover number of ZIP4, which is a crucial missing piece of information needed to better understand the transport mechanism. In this work, we used a non-radioactive zinc isotope,70Zn, and inductively coupled plasma mass spectrometry (ICP-MS) to study human ZIP4 (hZIP4) expressed in HEK293 cells. Our data showed that70Zn can replace the radioactive65Zn as a tracer in kinetic evaluation of hZIP4 activity. This approach, combined with the quantification of the cell surface expression of hZIP4 using biotinylation or surface-bound antibody, allowed us to estimate the apparent turnover number of hZIP4 to be in the range of 0.08-0.2 s-1. The turnover numbers of the truncated hZIP4 variants are significantly smaller than that of the full-length hZIP4, confirming a crucial role for the extracellular domain in zinc transport. Using64Zn and70Zn, we measured zinc efflux during the cell-based transport assay and found that it has little effect on the zinc import analysis under these conditions. Finally, we demonstrated that use of laser ablation (LA) ICP-TOF-MS on samples applied to a solid substrate significantly increased the throughput of the transport assay. We envision that the approach reported here can be applied to the studies of metal transporters beyond the ZIP family.
Polar flagellin glycan metaheterogeneity ofAeromonas hydrophilastrain ATCC 7966T
Authors: Fulton Kelly M. , Mendoza-Barberà Elena , Tomás Juan M. , Twine Susan M. , Smith Jeffrey C. , Merino Susana
Published: May 2024

Motile pathogens often rely upon flagellar motility as an essential virulence factor and in many species the structural flagellin protein is glycosylated. This post-translational modification has been shown to be necessary for proper folding of the flagellin structural proteins and proper function of the flagellar filament in a number of bacterial species.Aeromonas hydrophilais a ubiquitous aquatic pathogen with a constitutively expressed polar flagellum. Using a suite of mass spectrometry techniques, the flagellin FlaA and FlaB structural proteins ofA. hydrophilastrain ATCC 7966Twere shown to be glycosylated with significant metaheterogeneity: heterologous glycans were observed with variable site occupancy. The penta- and hexa-saccharide glycan chains contained a previously unreported pseudaminic acid derivative with a mass of 422 Da as the linking sugar, followed in sequence by two hexoses, anN-acetylglucosamine derivative, a deoxyN-acetylglucosamine derivative, and sometimes an additionalN-acetylglucosamine.
Decoding Secondary Motor Cortex Neuronal Activity during Cocaine Self-Administration: Insights from Longitudinalin vivoCalcium Imaging
Authors: Chen Yingying , Fu Haoying , Korada Amith , Lange Michal A. , Rayanki Chandrashekar , Lu Tao , Lai Dongbing , Fang Shiaofen , Guo Changyong , Ma Yao-Ying
Published: May 2024

Background

We recently reported that the risk of cocaine relapse is linked to hyperexcitability in the secondary motor cortex (M2) after prolonged withdrawal following intravenous self-administration (IVSA). However, the neuronal mechanisms underlying drug-taking behaviors and the response of M2 neurons to contingent drug delivery remain poorly understood.

Methods

Mice received cocaine as reinforcement (RNF) following active lever presses (ALP) but not inactive lever presses (ILP). Usingin vivocalcium imaging during cocaine IVSA, we tracked M2 neuronal activity with single-cell resolution. We then analyzed Ca2+transients in M2 at the earlyvs. late stages during the 1-hr daily sessions on IVSA Day 1 and Day 5.

Results

M2 neurons adapted to both operant behaviors and drug exposure history. Specifically, saline mice showed a reduction in both saline taking behaviors and Ca2+transient frequency with the 1-hr session. In contrast, cocaine mice maintained high ALP and RNF counts, with increased Ca2+transient frequency and amplitude on Day 1, persisting through Day 5. Compared to saline controls, cocaine mice exhibited a lower % of positively responsive neurons (Pos) and higher % of negatively responsive neurons (Neg) before ALP and after RNF, a difference not seen before ILP. Furthermore, as drug-taking behaviors progressed during the daily session, cocaine mice showed greater neuronal engagement with a larger population, particularly linked to ALP and RNF, with reduced overlap in neurons associated with ILP.

Conclusion

The M2 undergoes dynamic neuronal adaptations during early drug-taking behaviors, indicating its role as a potential substrate mediating the persistence of drug-seeking behaviors in cocaine relapse.
High-density CRISPRi screens reveal adaptive transcriptional gradients in cyanobacteria
Authors: Hren Andrew , Lollini Nicole , Carper Dana L. , Abraham Paul , Cameron Jeffrey C. , Fox Jerome M. , Eckert Carrie
Published: May 2024

Cyanobacteria are the oldest form of photosynthetic life on Earth and contribute to primary production in nearly every habitat, from permafrost to hot springs. Despite longstanding interest in the biochemical basis of environmental adaptation in these microbes, it remains poorly understood and challenging to re-wire. This study uses a high-density, genome-wide CRISPR interference (CRISPRi) screen to examine the influence of gene-specific transcriptional variation on the growth ofSynechococcussp. PCC 7002 under environmental extrema. Surprisingly, many partial knockdowns enhanced fitness under cold monochromatic conditions. Notably, transcriptional repression of a gene for a core subunit of the NDH-1 complex, which is important for photosynthesis and carbon uptake, improved growth rates under both red and blue light but at distinct, color-specific optima. In general, most genes with fitness-improving knockdowns were distinct to each light color, evidencing unique stress responses and alleviation mechanisms. Multi-target transcriptional repression produced nonadditive effects. Findings reveal diverse mechanisms of environmental adaptation in cyanobacteria and provide a new approach for using gradients in sgRNA activity to pinpoint biochemically influential transcriptional changes in cells.

SIGNIFICANCE STATEMENT

Cyanobacteria are the most abundant photosynthetic organisms on Earth, where they endure a striking variety of environmental fluctuations. This study examines the biochemical basis of environmental adaptation inSynechococcussp. PCC 7002, an important model strain, by modulating the expression of every gene in its genome. Results show that partial, but not complete, reduction in the expression of a subset of influential genes can improve growth under cold monochromatic conditions. Optimal expression levels differ between red and blue light and shift with multi-gene adjustments. Findings show how minor transcriptional adjustments can yield major improvements in growth under environmental extrema and provide a powerful systems-level approach for studying—and fine-tuning—the adaptive capacity of microbes.
Target gene responses differ when transcription factor levels are acutely decreased by nuclear export versus degradation
Authors: McGehee James , Stathopoulos Angelike
Published: May 2024

Defining the time of action for morphogens requires tools capable of temporally controlled perturbations. To study how the transcription factor Dorsal affects patterning of theDrosophilaembryonic dorsal-ventral axis, we used two light-inducible tags that result in either nuclear export or degradation of Dorsal when exposed to blue light. Nuclear export of Dorsal results in loss of expression for the high threshold, ventrally-expressed target genesnail(sna) but retention of the low threshold, laterally-expressed target geneshort-gastrulation(sog). In contrast, degradation of Dorsal results in retention ofsna,loss ofsog, and lower nuclear levels than when Dorsal is exported from the nucleus. To elucidate how nuclear export results in loss ofsnabut degradation does not, we investigated Dorsal kinetics using photobleaching and found it reenters the nucleus even under conditions of blue-light when export is favored. The associated kinetics of being imported and exported continuously are likely responsible for loss ofsnabut, alternatively, can supportsog. Collectively, our results show that this dynamic patterning process is influenced by both Dorsal concentration and nuclear retention.

SUMMARY STATEMENT

This study shows how optogenetic tools can be used to determine how a transcription factor’s levels and nuclear retention impact a dynamic patterning process.
Reprogramming SREBP1-dependent lipogenesis and inflammation in high-risk breast with licochalcone A: a novel path to cancer prevention
Authors: Hajirahimkhan Atieh , Bartom Elizabeth T. , Chung Carolina H , Guo Xingyu , Berkley Kyli , Lee Oukseub , Chen Ruohui , Cho Wonhwa , Chandrasekaran Sriram , Clare Susan E. , Khan Seema A.
Published: May 2024

Background

Anti-estrogens have had limited impact on breast cancer (BC) prevention. Novel agents with better tolerability, and efficacy beyond estrogen receptor (ER) positive BC are needed. We studied licochalcone A (LicA) for ER-agnostic BC prevention.

Methods

We evaluated antiproliferative effects of LicA in seven breast cell lines and its suppression of ER+ and ER− xenograft tumors in mice. High-risk human breast tissue was treated with LicAex vivo, followed by RNA sequencing and metabolism flux modeling. Confirmatory testing was performed in an independent specimen set and ER+/− BC cell lines using NanoString metabolic panel, proteomics, western blots, and spatiotemporally resolved cholesterol quantification in single cells.

Results

LicA suppressed proliferationin vitroand xenograft tumor growthin vivo. It downregulated pivotal steps in PI3K-AKT-SREBP1-dependent lipogenesis, suppressed PI3K and AKT phosphorylation, SREBP1 protein expression, and cholesterol levels in the plasma membrane inner leaflet, to the levels in normal breast cells. LicA also suppressed prostaglandin E2 synthesis and PRPS1-catalyzedde novonucleotide biosynthesis, stalling proliferation; further evident by reduced MKI67 and BCL2 proteins.

Conclusions

LicA targets SREBP1, a central regulator of lipogenesis and immune response, reducing pro-tumorigenic aberrations in lipid homeostasis and inflammation. It is a promising non-endocrine candidate for BC prevention.
Pseudomonas syringaesocially-induced swimming motility requires the molybdenum cofactor
Authors: Yang Zichu , Swingle Bryan
Published: May 2024

Social interactions among bacteria can induce behaviors that affect their fitness and influence how complex communities assemble. Here we report a new socially-induced motility behavior that we refer to as baited expansion inPseudomonas syringaepv. tomato DC3000 (PstDC3000), a plant pathogenic bacterium. We foundPstDC3000 displayed strongly-induced swimming motility towards nearby colonies ofDickeya dianthicolaorEscherichia coli. We developed a controlled system to visualize and characterize the development of baited expansion. Our results provide evidence that baited expansion behavior occurs in response to a chemical gradient established and maintained by the bait colony. We also found this behavior correlated with distinct transcriptional profiles and identified molybdenum cofactor as a crucial factor in facilitating the baited expansion behavior.
Neuraminidase 1 regulates the cellular state of microglia by modulating the sialylation of Trem2
Authors: Fremuth Leigh Ellen , Hu Huimin , van de Vlekkert Diantha , Annunziata Ida , Weesner Jason Andrew , Gomero Elida , d’Azzo Alessandra
Published: May 2024

Neuraminidase 1 (Neu1) cleaves terminal sialic acids from sialoglycoproteins in endolysosomes and at the plasma membrane. As such, Neu1 regulates immune cells, primarily those of the monocytic lineage. Here we examined how Neu1 influences microglia by modulating the sialylation of full-length Trem2 (Trem2-FL), a multifunctional receptor that regulates microglial survival, phagocytosis, and cytokine production. When Neu1 was deficient/downregulated, Trem2-FL remained sialylated, accumulated intracellularly, and was excessively cleaved into a C-terminal fragment (Trem2-CTF) and an extracellular soluble domain (sTrem2), enhancing their signaling capacities. Sialylated Trem2-FL (Sia-Trem2-FL) did not hinder Trem2-FL–DAP12–Syk complex assembly but impaired signal transduction through Syk, ultimately abolishing Trem2-dependent phagocytosis. Concurrently, Trem2-CTF–DAP12 complexes dampened NFκB signaling, while sTrem2 propagated Akt-dependent cell survival and NFAT1-mediated production of TNFα and CCL3. Because Neu1 and Trem2 are implicated in neurodegenerative/neuroinflammatory diseases, including Alzheimer disease (AD) and sialidosis, modulating Neu1 activity represents a therapeutic approach to broadly regulate microglia-mediated neuroinflammation.
Rapid detection, quantification and speciation ofLeishmaniausing real-time PCR and DNA sequencing at the rRNA Internal Transcribed Spacer 2
Authors: Paun Andrea , Grigg Michael E.
Published: May 2024

The ability to discriminate infection between closely relatedLeishmaniaspecies within theVianniaspecies complex, specificallyL. braziliensis, L. guyanensisandL. panamensisis critical to inform the clinical diagnosis and determine the most efficacious treatment modality. We designed a nested primer set targeting the rRNA Internal Transcribed Spacer 2 (ITS2), located on Chromosome 27, to distinguish among all human infectiveLeishmaniaspecies. Separate nested and single primer pairs were developed for conventional and quantitative PCR approaches respectively. Species-specific single nucleotide polymorphisms and indels located within the PCR products were identified by Sanger sequencing. This single locus approach provides a sensitive and specific platform to identify the species ofLeishmaniacausing infection.
Enhancing tumor-infiltrating T cells with an exclusive fuel source
Authors: Miller Matthew L. , Thauland Timothy J. , Nagarajan Rina , Zuo Wenqi Ellen , Moreno Lastre Miguel A. , Butte Manish J.
Published: May 2024

Solid tumors harbor immunosuppressive microenvironments that inhibit tumor-infiltrating lymphocytes (TILs) through the voracious consumption of glucose. We sought to restore TIL function by providing them with an exclusive fuel source. The glucose disaccharide cellobiose, which is the building block of cellulose, contains a β-1,4-glycosidic bond that that animals (or their tumors) cannot hydrolyze, but microbes have evolved enzymes to catabolize cellobiose into useful glucose. We equipped mouse T cells and human CAR-T cells with two proteins enabling import and hydrolysis of cellobiose and demonstrated that cellobiose supplementation during glucose withdrawal restores key anti-tumor T-cell functions: viability, proliferation, cytokine production, and cytotoxic killing. Engineered T cells offered cellobiose suppress murine tumor growth and prolong survival. Offering exclusive access to a natural disaccharide is a new tool that augments cancer immunotherapies. This approach could be used to answer questions about glucose metabolism across many cell types, biological processes, and diseases.
Opioid Modulation of Differential Gene Expression and Neuronal Differentiation in the Ventricular-Subventricular Zone of Adult Male Zebra Finches
Authors: Din Uzma , Banerjee Sourav , Iyengar Soumya
Published: May 2024

1

The endogenous opioid system modulates diverse functions including pain, physiological functions and motivation. We had earlier demonstrated an increase in cell proliferation in the neurogenic niche of adult male zebra finches, following administration of the general opioid antagonist, naloxone. Since the mechanisms underlying opioid modulation of neurogenesis are poorly understood, we have investigated the molecular mechanisms underlying cell proliferation differentiation induced by the opioid system. Systemic naloxone or vehicle administration in adult male birds was followed by a whole transcriptome microarray assay of the ventricular-subventricular zone (V-SVZ). The analysis revealed 26 differentially expressed transcripts (expression fold change ≥ 1.5), encoding genes important for neuronal functions. The transcriptomics analysis using microarray identified a significant increase in the expression of precursor miRNAtgu-mir-124-201transcript following inhibition of opioid receptors by naloxone. Our results suggested that the administration of naloxone triggers an increase in the expression of the pro-differentiating microRNA, miR-124, in the brains of adult zebra finches. Using quantitative real-time PCR, we found that the expression of miR-124-3p was higher in the V-SVZ of naloxone-treated versus control birds. Furthermore, the density of neuroblasts was higher in the ventral V-SVZ adjacent to the striatal nucleus Area X, but not in the V-SVZ above the pallial nucleus, HVC, both of which are involved in singing. Since miR-124 targets signalling pathways important for neuronal proliferation and differentiation, our findings suggest that microRNA-124 may be involved in the increase in neurogenesis in adult songbirds induced by altering opioid modulation.
Investigating neurometabolite changes in response to median nerve stimulation
Authors: Houlgreave Mairi S. , Dyke Katherine , Berrington Adam , Jackson Stephen R.
Published: May 2024

Background

Rhythmic median nerve stimulation (MNS) at 10Hz has been shown to cause a substantial reduction in tic frequency in individuals with Tourette Syndrome. The mechanism of action is currently unknown, but is hypothesised to involve entrainment of oscillations within the sensorimotor cortex.

Objective

We used functional magnetic resonance spectroscopy (fMRS) to explore the dynamic effects of MNS on neurometabolite concentrations.

Methods

Here, we investigated the effects of rhythmic and arrhythmic 10Hz MNS on glutamate (Glu) and GABA concentrations in the contralateral sensorimotor cortex in 15 healthy controls, using a blocked fMRS design. We used a Mescher-Garwood-semi-localised by adiabatic selective refocusing (MEGA-sLASER) sequence at 7T.

Results

Our results show no difference in the difference-from-baseline measures between the two stimulation conditions. Looking at the effect of MNS over both conditions there is a trend for an initial increase in Glu/tCr (total creatine) followed by a decrease over time, whereas GABA/tCr decreased during each stimulation block.

Conclusion

These results suggest that despite entrainment of oscillations during rhythmic MNS, there are no significant differences in the tonic neuromodulatory effects of rhythmic and arrhythmic stimulation. The reduction in glutamate over the course of stimulation may reflect a decrease in glutamatergic firing due to adaptation. This may make it less likely that an involuntary movement is generated during continuous stimulation.
Assessing the potential antifungal resistance risk from dual use of a mode of action in agriculture and medical treatment of human pathogens
Authors: Paveley Neil , van den Bosch Frank , Grimmer Michael
Published: May 2024

A mechanistic basis is described for assessment of resistance risk to medical anti-fungal treatments from agricultural use of fungicides of the same mode of action. The following need to occur in landscape environments for a risk to be posed by ‘dual use’: (i) emergence, whereby a resistant strain emerges by mutation and invasion, (ii) selection, whereby a mutation conferring a fitness advantage is selected for in the presence of fungicide, and (iii) exposure of humans to resistant strains from the landscape, potentially resulting in invasive fungal infection (IFI). We identify 20 human fungal pathogens for which there is evidence that all three processes above could, in principle, occur. A model is derived for quantitative analysis to explore what determines resistance emergence and selection in human pathogens in landscape environments. Emergence and selection were particularly affected by fitness cost associated with the resistance mutation(s) and fungicide concentration.

Emergence was also determined by the amount of pathogen reproduction (related to pathogen population size). The findings were related to an example case of observational data from the Netherlands for Aspergillus fumigatus. The analysis supports previous work that compost, including bulb waste, is towards the high-risk end of the spectrum for this species. Agricultural soils, non-agricultural land and grassland were lower risk. More generally, across species, the model output suggests that if fungicide resistance is associated with even a small fitness cost, then environments with low fungicide concentrations, such as field soils and semi-natural environments (e.g. woodland), may not be conducive to resistance emergence or selection.
Parallel Evolution of Salinity Tolerance inArabidopsis thalianaAccessions from Cape Verde Islands
Authors: Rivas Félix J Martínez , Wozny Dorothee , Xue Zeyun , Gilbault Elodie , Sapir Thomas , Rouille Melissa , Ricou Antony , Medina Joaquín , Noël Laurent D. , Lauber Emmanuelle , Voxeur Aline , Loudet Olivier , Clément Gilles , Jiménez-Gómez Jose M
Published: May 2024

Soil salinization poses a significant threat to crop production impacting one fifth of all cultivated land. The Cape Verde Islands are located 600 km from the coast of Africa and are characterized by high salinity soils and inland water sources.

In this study we find thatArabidopsis thalianaplants native to these islands accumulate a metabolite that protects them from salt stress. We partially characterized this metabolite as glucuronyl-mannose. We find that the ability to produce glucuronyl-mannose evolved independently in two different islands from the same archipelago through mutations in the same gene, an alpha glycosidase protein that we named GH38cv. These cases of parallel evolution suggest positive selection towards the increase of salt tolerance with low fitness costs. Indeed, plants carrying derived alleles of GH38cv do not present growth defects or low defenses under normal conditions, but show better germination rates, longer roots and better hydric status than non-mutated plants when exposed to salt stress. These findings provide a knowledge-based method to develop salt resilient crops using natural mechanisms, which could be attractive both to conventional and organic agriculture.
SpatialDiffusion: Predicting Spatial Transcriptomics with Denoising Diffusion Probabilistic Models
Authors: Khan Sumeer Ahmad , Lagani Vincenzo , Lehmann Robert , Kiani Narsis A. , Gomez-Cabrero David , Tegner Jesper
Published: May 2024

Spatial Transcriptomics (ST) allows deep characterization of the 2D organization of expression data within tissue slices. The ST technology provides a tissue contextualization of deep single-cell profiles. Recently, numerous computational and machine learning methods have addressed challenges such as data quality, augmentation, annotation, and the development of integrative platforms for data analysis. In contrast, here we ask whetherunseen spatial transcriptomics data can be predicted and if we can interpolate novel transcriptomic slices. To this end, we adopt a denoising diffusion probabilistic-based model (DDPM) todemonstrate the learning ofgenerative ST models for several tissues. Furthermore, our generative diffusion model interpolates (predicts) unseen slices located “between” the collected finite number of ST slices. This methodology set the stage for learning predictive deep 3D models of tissues from a finite number of spatial transcriptomics slices, thus harboring the advent of AI-augmented spatial transcriptomics.
Thermodynamics Underpinning the Microbial Community-Level Nitrogen Networks
Authors: Seto Mayumi , Sasaki Risa , Ooka Hideshi , Nakamura Ryuhei
Published: May 2024

Nitrogen species often serve as crucial electron donors or acceptors in microbial catabolism, enabling the synthesis of adenosine triphosphate (ATP). Although theoretically any nitrogen redox reactions could be an energy source, it remains unclear why specific reactions are predominantly utilized. This study evaluates energetically superior reactions from 988 theoretically plausible combinations involving 11 nitrogen species, oxygen gas, hydrogen ion, and water. Our analysis of the similarity between this model-based energetically superior network and the actual microbial community-level nitrogen network, reconstructed as a combination of enzymatic reactions, showed increased link overlap rates with thermodynamic weighting on reaction rates. In particular, existing microbial reactions involving solely nitrogen species and additionally oxygen, such as anaerobic ammonia oxidation (ANAMMOX) and complete and partial nitrification, were frequently identified as energetically superior among the examined reactions. The alignment of these reactions with thermodynamically favorable outcomes underscores the critical role of thermodynamics not only in individual metabolic processes but also in shaping the broader network interactions within ecosystems, consequently affecting biodiversity and ecological functions.

Significance Statement

This study advances our understanding of how thermodynamics governs energy metabolism at the community level within microbial ecosystems by systematically analyzing 988 potential redox reactions involving inorganic nitrogen species, oxygen gas, hydrogen ion, and water. We uncover that existing microbial reactions, such as anaerobic ammonia oxidation (ANAMMOX) and nitrification, stand out as energetically superior over other examined reactions. The robust alignment between model-predicted energetically favorable reactions and actual microbial nitrogen reactions underscores the predictive power of thermodynamic principles, even in ecological networks. Our findings extend the traditional applications of thermodynamics in biology, highlighting how thermodynamic constraints shape ecological networks and influence biodiversity and ecosystem functions in natural ecosystems.
Pro-survival roles for p21(Cip1/Waf1) in Non-Small Cell Lung Cancer
Authors: Cutty SJ , Hughes FA , Ortega-Prieto P , Desai S , Thomas P , Fets LV , Secrier M , Barr AR
Published: May 2024

Quiescence is a reversible state of proliferative arrest, distinct from senescence. While cancer is a disease of dysregulated proliferation, cancer cells can retain the ability to enter quiescence which confers advantages to tumour cells by protecting them from chemotherapy or by allowing metastasis to distant sites. Multiple mechanisms exist to induce and maintain quiescence that are not yet fully understood. Here, we show that high expression of the CDK inhibitor p21Cip1/Waf1correlates with a poor prognosis inTP53wild-type, but notTP53mutant, non-small cell lung cancer (NSCLC) patients. Using quantitative single-cell imaging of genetically-engineered NSCLC reporter cell lines, we show thatTP53wild-type NSCLC cells can enter a p21-dependent spontaneous quiescent state, downstream of replication stress. Furthermore, p21 expression confers survival advantages toTP53wild-type NSCLC cells, both under normal proliferation and in response to chemotherapy. We also show that p21 can promote tumour relapse by allowing cells to recover from both G1 and G2 arrest states after drug removal. Together, our data suggest that targeting p21 function inTP53wild-type tumours could lead to better outcomes for chemotherapy treatment in NSCLC patients.

Statement of Significance

We show thatTP53WT Non-Small Cell Lung Cancer cells can enter a p21-dependent spontaneous quiescent state and that p21 maintains the viability of NSCLC cells, is chemoprotective and can promote tumour relapse.
Paenilamicins from the honey bee pathogenPaenibacillus larvaeare context-specific translocation inhibitors of protein synthesis
Authors: Koller Timm O. , Berger Max J. , Morici Martino , Paternoga Helge , Bulatov Timur , Di Stasi Adriana , Dang Tam , Mainz Andi , Raulf Karoline , Crowe-McAuliffe Caillan , Scocchi Marco , Mardirossian Mario , Beckert Bertrand , Vázquez-Laslop Nora , Mankin Alexander , Süssmuth Roderich D. , Wilson Daniel N.
Published: May 2024

The paenilamicins are a group of hybrid non-ribosomal peptide-polyketide compounds produced by the honey bee pathogenPaenibacillus larvaethat display activity against Gram-positive pathogens, such asStaphylococcus aureus. While paenilamicins have been shown to inhibit protein synthesis, their mechanism of action has remained unclear. Here, we have determined structures of the paenilamicin PamB2 stalled ribosomes, revealing a unique binding site on the small 30S subunit located between the A- and P-site tRNAs. In addition to providing a precise description of interactions of PamB2 with the ribosome, the structures also rationalize the resistance mechanisms utilized byP. larvae. We could further demonstrate that PamB2 interferes with the translocation of mRNA and tRNAs through the ribosome during translation elongation, and that this inhibitory activity is influenced by the presence of modifications at position 37 of the A-site tRNA. Collectively, our study defines the paenilamicins as a new class of context-specific translocation inhibitors.
Lymphostatin: Structure of a large multi-functional virulence factor
Authors: Griessmann Matthias , Rasmussen Tim , Flegler Vanessa J. , Kraft Christian , Schneider Ronja , Spantzel Lukas , Stevens Mark P. , Börsch Michael , Böttcher Bettina
Published: May 2024

Enteropathogenic and EnterohaemorrhagicEscherichia coliare enteric pathogens of global importance and human infections can be life-threatening. Lymphostatin is a key virulence factor of these bacteria, being required for intestinal colonisation and a potent inhibitor of the mitogen- and antigen-activated proliferation of lymphocytes and proinflammatory responses. In some strains, it also mediates adherence to host cells and influences actin nucleation at sites of attachment. This 365 kDa protein requires glycosyltransferase and cysteine protease motifs for activity against lymphocytes, but high-resolution structural information has proven elusive and the molecular mechanisms by which it acts remain unclear. Here, we describe the structure of lymphostatin from the prototype O127:H6 enteropathogenicE. colistrain determined by electron cryo-microscopy. Our results reveal two glycosyltransferase domains, one PaTox-like protease domain, an ADP-ribosyltransferase domain, and a delivery domain. Long linkers act to hold these domains together. These linkers occlude the catalytic sites of the N-terminal glycosyltransferase and protease domains. In this dormant state, lymphostatin binds to HEK-293T cells, where it forms large clusters before being taken up and sequestered into cytosolic foci. With more functional domains than any other known large bacterial toxin, lymphostatin can be regarded as the multifunctional Swiss army knife of pathogenicEscherichia coli, enabling complex interactions with the host cells in different environments.
Cerebellar control over inter-regional excitatory/inhibitory dynamics discriminates execution from observation of an action
Authors: Lorenzi Roberta Maria , Korkmaz Gökçe , Alahmadi Adnan A.S. , Monteverdi Anita , Casiraghi Letizia , D’Angelo Egidio , Palesi Fulvia , Gandini Wheeler-Kingshott Claudia A.M.
Published: May 2024

The motor learning theory anticipates that cerebro-cerebellar loops perform sensorimotor prediction thereby regulating motor control. This operation has been identified during action execution (AE) and observation (AO) but the causal interaction between the cerebellum and cerebral cortex remained unclear. Here we used Dynamic Causal Modelling (DCM) to study functional MRI (fMRI) data obtained during a squeeze ball task in either the AE or AO conditions. In both cases, active regions included bilateral primary visual cortex (V1), left primary motor cortex (M1), left supplementary motor and premotor cortex (SMAPMC), left cingulate cortex (CC), left superior parietal lobule (SPL), and right cerebellum (CRBL). AE and AO networks showed the same fixed effective connectivity, with pathways between V1, CRBL, SMAPMC and CC wired in a closed loop. However, the cerebellar communication towards the cerebral cortex switched from excitatory in AE to inhibitory in AO. Moreover, in AE only, signal modulation was non-linear from SMAPMC to CRBL and within the CRBL self-connection, supporting the role of the CRBL in elaborating motor plans received from SMAPMC. Thus, the need for motor planning and the presence of a sensorimotor feedback in AE discriminate the modality of forward control operated by the CRBL on SMAPMC. While the underlying circuit mechanisms remain to be determined, these results reveal that the CRBL differentially controls the excitatory/inhibitory dynamics of inter-regional effective connectivity depending on its functional engagement, opening new prospective for the design of artificial sensorimotor controllers and for the investigation of neurological diseases.
Inhibition of Inflammatory Osteoclasts Enhances CGRP+TrkA+Signaling and Accelerates Callus Remodeling in Osteoporotic Fractures
Authors: Shu Yuexia , Tan Zhenyu , Pan Zhen , Chen Yujie , Wang Jielin , He Jieming , Wang Jia , Wang Yuan
Published: May 2024

Impaired callus remodeling significantly contributes to the delayed healing of osteoporotic fractures; however, the underlying mechanisms remain unclear. Sensory neuronal signaling plays a crucial role in bone repair. In this study, we demonstrate that in ovariectomized (OVX) mice, the loss of CGRP+TrkA+sensory neuronal signaling during callus remodeling correlates with increased Cx3cr1+iOCs expression within the bone callus. Conditional knockout of Cx3cr1+iOCs restored CGRP+TrkA+sensory neuronal, enabling normal callus remodeling progression. Mechanistically, we further demonstrate that Cx3cr1+iOCs secrete seme3A in the osteoporotic fracture repair microenvironment, inhibiting CGRP+TrkA+sensory neurons’ axonal regeneration and suppressing nerve-bone signaling exchange, thus hindering bone remodeling. Lastly, in human samples, we observed an association between the loss of CGRP+TrkA+sensory neuronal signaling and increased expression of Cx3cr1+iOCs. In conclusion, enhancing CGRP+TrkA+sensory nerve signaling by inhibiting Cx3cr1+iOCs activity presents a potential strategy for treating delayed healing in osteoporotic fractures.
Dynamic and decay kinetics of H3 variants in live cells reveal the pivotal role of HIRA/NSD2 in maintaining the distinct H3.3 specific chromatin landscape
Authors: Nehru Vishal , Ball David , Mukherjee Abhishek , Kurotaki Daisuke , Chitnis Ajay , Karpova Tatiana S. , Ozato Keiko
Published: May 2024

The incorporation of variant histone H3.3 into the genome is tightly linked with transcriptional activity, yet its precise regulatory mechanisms remain elusive. Traditional methods like Chromatin Immunoprecipitation offer static views of H3.3 distribution, lacking dynamic insights. Here, using the SNAP tag system, we employed Fluorescence Recovery After Photobleaching (FRAP) and live-cell imaging to investigate H3.3 mobility and decay kinetics in live mouse embryonic fibroblast cells. Our focus on interferon-induced transcriptional activation revealed rapid H3.3 exchange, indicative of its transcriptional regulatory role. Transcription inhibition hindered H3.3 mobility, emphasizing its involvement in transcription. Additionally, we probed into turnover dynamics(decay) of H3.1-SNAP and H3.3-SNAP variants, uncovering differential decay rates influenced by transcriptional activity and histone modifiers such as NSD2 and HIRA. Live-cell imaging showed faster decay of H3.3 compared to H3.1, further exacerbated upon NSD2/HIRA loss. Notably, HIRA and NSD2, regulators of H3.3 dynamics, proved crucial for both H3.3 mobility and decay, underscoring their pivotal role. These findings deepen our understanding of epigenetic regulation, emphasizing the dynamic nature of histone turnover in cellular function and its implications for disease pathogenesis. Taken together, this study sheds light on the dynamic behavior of H3.3 and its regulatory mechanisms, providing valuable insights into epigenetic regulation in cellular processes and disease contexts.