bluuebunny/crossref_metadata_2025 huggingface dataset:
Connexin43 in mesenchymal lineage cells regulates body adiposity and energy metabolism in mice
Authors: Lee Seung-Yon , Fontana Francesca , Sugatani Toshifumi , Castillo Ignacio Portales , Leanza Giulia , Coler-Reilly Ariella , Civitelli Roberto
Published: January 2024

Connexin43 (Cx43) is the most abundant gap junction protein present in the mesenchymal lineage. In mature adipocytes, Cx43 mediates white adipose tissue (WAT) “beiging” in response to cold exposure and maintains the mitochondrial integrity of brown adipose tissue (BAT). We found that genetic deletion ofGja1(Cx43 gene) in cells that give rise to chondro-osteogenic and adipogenic precursors driven by theDermo1/Twist2promoter leads to lower body adiposity and partial protection against the weight gain and metabolic syndrome induced by a high fat diet (HFD) in both sexes. These protective effects from obesogenic diet are related to increased locomotion, fuel utilization, energy expenditure, non-shivering thermogenesis, and better glucose tolerance in conditionallyGja1ablated mice. Accordingly,Gja1mutant mice exhibit reduced adipocyte hypertrophy, partially preserved insulin sensitivity, increased BAT lipolysis and decreased whitening under HFD. This metabolic phenotype is not reproduced with more restrictedGja1ablation in differentiated adipocytes, suggesting that Cx43 has a hitherto unknown function in adipocyte progenitors or other targeted cells, resulting in restrained energy expenditures and fat accumulation. These results disclose an hitherto unknown action of Cx43 in adiposity, and offer a promising new pharmacologic target for improving metabolic balance in diabetes and obesity.

Graphical Abstract
Rapid Emergence and Evolution of SARS-CoV-2 Variants in Advanced HIV Infection
Authors: Ko Sung Hee , Radecki Pierce , Belinky Frida , Bhiman Jinal N. , Meiring Susan , Kleynhans Jackie , Amoako Daniel , Guerra Canedo Vanessa , Lucas Margaret , Kekana Dikeledi , Martinson Neil , Lebina Limakatso , Everatt Josie , Tempia Stefano , Bylund Tatsiana , Rawi Reda , Kwong Peter D. , Wolter Nicole , von Gottberg Anne , Cohen Cheryl , Boritz Eli A.
Published: January 2024

Previous studies have linked the evolution of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) genetic variants to persistent infections in people with immunocompromising conditions1–4, but the evolutionary processes underlying these observations are incompletely understood. Here we used high-throughput, single-genome amplification and sequencing (HT-SGS) to obtain up to ∼103SARS-CoV-2 spike gene sequences in each of 184 respiratory samples from 22 people with HIV (PWH) and 25 people without HIV (PWOH). Twelve of 22 PWH had advanced HIV infection, defined by peripheral blood CD4 T cell counts (i.e., CD4 counts) <200 cells/μL. In PWOH and PWH with CD4 counts ≥200 cells/μL, most single-genome spike sequences in each person matched one haplotype that predominated throughout the infection. By contrast, people with advanced HIV showed elevated intra-host spike diversity with a median of 46 haplotypes per person (IQR 14-114). Higher intra-host spike diversity immediately after COVID-19 symptom onset predicted longer SARS-CoV-2 RNA shedding among PWH, and intra-host spike diversity at this timepoint was significantly higher in people with advanced HIV than in PWOH. Composition of spike sequence populations in people with advanced HIV fluctuated rapidly over time, with founder sequences often replaced by groups of new haplotypes. These population-level changes were associated with a high total burden of intra-host mutations and positive selection at functionally important residues. In several cases, delayed emergence of detectable serum binding to spike was associated with positive selection for presumptive antibody-escape mutations. Taken together, our findings show remarkable intra-host genetic diversity of SARS-CoV-2 in advanced HIV infection and suggest that adaptive intra-host SARS-CoV-2 evolution in this setting may contribute to the emergence of new variants of concern (VOCs).
Spatial characterization of interface dermatitis in cutaneous lupus reveals novel chemokine ligand-receptor pairs that drive disease
Authors: Shakiba Saeed , Haddadi Nazgol-Sadat , Afshari Khashayar , Lubov Janet E. , Raef Haya S. , Li Robert , Yildiz-Altay Ümmügülsüm , Daga Mridushi , Refat Maggi Ahmed , Kim Evangeline , Galindo de Laflin Johanna , Akabane Andressa , Sherman Shany , MacDonald Elizabeth , Strassner James P. , Zhang Liang , Leon Michael , Baer Christina E. , Dresser Karen , Liang Yan , Whitley James B , Skopelja-Gardner Sladjana , Harris John E , Deng April , Vesely Matthew D. , Rashighi Mehdi , Richmond Jillian
Published: January 2024

Chemokines play critical roles in the recruitment and activation of immune cells in both homeostatic and pathologic conditions. Here, we examined chemokine ligand-receptor pairs to better understand the immunopathogenesis of cutaneous lupus erythematosus (CLE), a complex autoimmune connective tissue disorder. We used suction blister biopsies to measure cellular infiltrates with spectral flow cytometry in the interface dermatitis reaction, as well as 184 protein analytes in interstitial skin fluid using Olink targeted proteomics. Flow and Olink data concordantly demonstrated significant increases in T cells and antigen presenting cells (APCs). We also performed spatial transcriptomics and spatial proteomics of punch biopsies using digital spatial profiling (DSP) technology on CLE skin and healthy margin controls to examine discreet locations within the tissue. Spatial and Olink data confirmed elevation of interferon (IFN) and IFN-inducible CXCR3 chemokine ligands. Comparing involved versus uninvolved keratinocytes in CLE samples revealed upregulation of essential inflammatory response genes in areas near interface dermatitis, includingAIM2. Our Olink data confirmed upregulation of Caspase 8, IL-18 which is the final product of AIM2 activation, and induced chemokines including CCL8 and CXCL6 in CLE lesional samples. Chemotaxis assays using PBMCs from healthy and CLE donors revealed that T cells are equally poised to respond to CXCR3 ligands, whereas CD14+CD16+ APC populations are more sensitive to CXCL6 via CXCR1 and CD14+ are more sensitive to CCL8 via CCR2. Taken together, our data map a pathway from keratinocyte injury to lymphocyte recruitment in CLE via AIM2-Casp8-IL-18-CXCL6/CXCR1 and CCL8/CCR2, and IFNG/IFNL1-CXCL9/CXCL11-CXCR3.

One Sentence Summary

We mapped chemokine orchestrators of interface dermatitis in lupus using spatial approaches on archival tissue and confirmed with fresh tissues.
Evidence forNR2F2/COUP-TFII involvement in human testis development
Authors: Wankanit Somboon , Zidoune Housna , Bignon-Topalovic Joëlle , Schlick Laurène , Houzelstein Denis , Fusée Leila , Boukri Asma , Nouri Nassim , McElreavey Ken , Bashamboo Anu , Elzaiat Maëva
Published: January 2024

NR2F2encodes COUP-TFII, an orphan nuclear receptor involved in mammalian gonadogenesis. COUP-TFII is expressed in the interstitial/stromal compartment of both fetal testes and ovaries, where it is required for developing steroidogenic lineages. Pathogenic variants in humanNR2F2are linked to testis formation in 46,XX individuals (46,XXdisorders ofsexdevelopment, 46,XX DSD). Such findings propose a regulatory role of COUP-TFII in the developing ovary, whereas its function in testis remains unknown. We evaluate the effect of ade novoheterozygous, predicted damaging, missense variant inNR2F2(p.Arg246His) in a 46,XY under-masculinized boy.In-vitroassays show that the mutant protein significantly loses the inhibitory effect on NR5A1-mediated activation of both theLHBandINSL3promoters. The data support the pathogenicity of the p.Arg246His variant in 46,XY DSD and a role forNR2F2in human testis formation. In addition toNR5A1andWT1,NR2F2variants are thus associated with both 46,XX and 46,XY DSD. This expands the list of genes that function in both male and female sex development, which is originally thought to be regulated by two entirely different sets of genes.
Nitrogen Management on Improving Resilience of Flood-Tolerant (Sub1) Rice Varieties in Flood-Prone Rainfed Lowlands
Authors: Singh A. K. , Pandey A. K. , Singh Ankit , Tiwari Deeksha , Singh Bijendra
Published: January 2024

Flash flood causing submergence adversely affects rice production in vast areas of rainfed lowlands of South and Southeast Asia. Introgression at the Sub1 locus by molecularly assisted backcrossing incorporated tolerance into varieties traditional/modern high-yielding varieties and has released some varieties for commercial planting by Indian farmers. In the present study, we investigated the application of fractional N to further improve the survival and productivity potential of two Sub1 introgression varieties, Sambha Mahsuri-Sub1, and BR11-Sub1.Thirty days (d) old seedlings were transplanted, and 28-d after transplanting i.e. 58 days old seedlings were completely submerged for 18-d. Full doses of phosphorus and potassium were applied as a basal, while nitrogen was applied in split doses. Plant survival was recorded at 0, 5 and 20 days after de-submergence (AS) to study the recovery dynamics of cultivar Sub1. Results evidently established that varieties with higher biomass, tiller number, total chlorophyll and soluble sugars concentrations before flooding have higher survival rates after water drainage. They also had faster growth and better recovery, which was reflected in yield characteristics and grain yield. The 20-day and 40- day AS N application replenished flood-disturbed soil N pools, resulting in higher N uptake and N-use efficiency. These results may contribute to better nitrogen fertilization programs for rice crops and improve stress tolerance in newly developed tolerant rice varieties. It is evident from perusal of results that treatment T2i.e.1/4 N applied as a basal and rest of residual N applied in three split doses respectively after de-submergence on days 5, 20 and 40 resulted in higher survival thanT1 and T3 respectively minimising mortality rate. This method of nitrogen fertilization (T2) also significantly affected plant-1stem biomass, plant-1tiller number, total chlorophyll, and soluble sugar concentrations before and after flooding. Based on these findings we proposed that application of lower dose of N (30kg/ha) as basal and rest amount of N in three split doses along with P and K (40kg/ha) in the field might be exploited to improve submergence tolerance and to obtain higher yield under flood prone ecosystem due to higher survival after de submergence corresponding to less post oxidative damage through proper N management during before and post submergence period.
Comparing multifunctional viral and eukaryotic proteins for generating scission necks in membranes
Authors: Alimohamadi Haleh , Luo Elizabeth Wei-Chia , Gupta Shivam , de Anda Jaime , Yang Rena , Mandal Taraknath , Wong Gerard C. L.
Published: January 2024

Deterministic formation of membrane scission necks by protein machinery with multiplexed functions is critical in biology. A microbial example is the M2 viroporin, a proton pump from the influenza A virus which is multiplexed with membrane remodeling activity to induce budding and scission in the host membrane during viral maturation. In comparison, the dynamin family constitutes a class of eukaryotic proteins implicated in mitochondrial fission, as well as various budding and endocytosis pathways. In the case of Dnm1, the mitochondrial fission protein in yeast, the membrane remodeling activity is multiplexed with mechanoenzyme activity to create fission necks. It is not clear why these functions are combined in these scission processes, which occur in drastically different compositions and solution conditions. In general, direct experimental access to changing neck sizes induced by individual proteins or peptide fragments is challenging due to the nanoscale dimensions and influence of thermal fluctuations. Here, we use a mechanical model to estimate the size of scission necks by leveraging Small-Angle X-ray Scattering (SAXS) structural data of protein-lipid systems under different conditions. The influence of interfacial tension, lipid composition, and membrane budding morphology on the size of the induced scission necks is systematically investigated using our data and molecular dynamic simulations. We find that the M2 budding protein from the influenza A virus has robust pH-dependent membrane activity that induces nanoscopic necks within the range of spontaneous hemi-fission for a broad range of lipid compositions. In contrast, the sizes of scission necks generated by mitochondrial fission proteins strongly depend on lipid composition, which suggests a role for mechanical constriction.
From Understudied to Understood: Multi-Omics Analysis with MiniENCODE Exemplified by Zebrafish
Authors: Yang Hang , Shan Zhaojun , Shang Hanqiao , Jiang Puxuan , Li Yingshu , Tu Qiang
Published: January 2024

The ENCODE project provides a comprehensive map of genomic activity in humans and mice. However, its extensive toolbox can be too costly and technically challenging for smaller research communities. This presents a hurdle for understudied model organisms where resource constraints and complex omics data integration further compound the challenge. To bridge this gap, our study introduces a miniENCODE project, using the zebrafish as an exemplar. We collected, generated and integrated datasets from three experimental assays across developmental stages and adult tissues. This analysis is facilitated by our newly developed miniODP database, a computational tool designed for the integration and visualization of multiomics data. Utilizing these methods, we have cataloged over 52,000 candidate enhancers, identified various stage-specific, tissue-specific, and constantly active enhancers, and experimentally validated a subset of heart-specific enhancers. We have delineated key transcription factors and their corresponding regulatory networks across developmental stages and adult tissues. This study not only delivers valuable regulatory insights for the zebrafish research community but also illustrates how an integrative approach can aid in understanding complex regulatory mechanisms even with limited resources. This strategy could empower scientific communities working with various understudied model organisms to expedite their genomic research without overstretching their resources.
Understanding feeding competition under laboratory conditions: Rohu (Labeo rohita)versus Amazon sailfin catfish (Pterygoplichthysspp.)
Authors: Mallick Suman , Sundaray Jitendra Kumar , Ghosal Ratna
Published: January 2024

Competitive interactions between species is widely prevalent within the animal world. In this manuscript, we attempted to understand feeding competitions between the Amazon sailfin catfish, an invasive species introduced globally, and rohu, a keystone species native to several countries within southeast Asia. We used two different size classes of each species, large-size having total length (TL, from snout tip to caudal fin) of 15-20 cm and fingerling having TL <6 cm, and feeding duration was used as a proxy to understand competition. Our results demonstrated that feeding durations of large-size rohu were either similar or significantly (P<0.05) higher in presence of catfish when compared to trials in presence of conspecifics, indicating that large-size rohu is not a weak competitor. However, feeding durations of fingerling rohu was significantly (P<0.05) reduced in presence of both large-size and fingerling catfish, when compared to trials in presence of conspecifics. Moreover, fingerling rohu also displayed freeze (alarm) behavior in presence of the catfish. Interestingly, presence of rohu had no significant (P>0.05) impact on feeding durations of catfish. Overall, the study demonstrated that invasive catfish may behaviorally outcompete fingerling rohu, thus, threatening the sustenance of a species that is native to several freshwaters around the globe.
Interactors and effects of overexpressing YlxR/RpnM, a conserved RNA binding protein in cyanobacteria
Authors: Hemm Luisa , Miucci Anna , Riediger Matthias , Tholen Stefan , Kraus Alexander , Georg Jens , Schilling Oliver , Hess Wolfgang R.
Published: January 2024

Throughout the tree of life RNA-binding proteins play important roles, but they are poorly characterized in cyanobacteria. Structural prediction suggests an RNA-binding interface for the protein YlxR/Ssr1238 in the cyanobacteriumSynechocystis6803. Two pairs of cysteine residues are arranged as possibly coordinating an Fe-S cluster and appear widely conserved in the homologous proteins of other cyanobacteria. Overexpression of Ssr1238 for 24 h led to higher levels of RNase P RNA, tRNAs, and stress-related mRNAs. Co-immunoprecipitation of proteins followed by MS analysis and sequencing of UV crosslinked, co-immunoprecipitated RNA samples identified potential interaction partners of Ssr1238. The most enriched transcript was RNase P RNA, and RnpA, the protein component of RNase P, was among the most highly enriched proteins. A second highly enriched transcript derived from genessl3177, which encodes a central enzyme in cell wall remodeling during cell division. The data also showed a strong connection to the RNA maturation and modification system indicated by co-precipitation of RNA modifying enzymes, riboendonuclease E and enolase. Surprisingly, cyanophycin synthetase and urease were highly enriched as well. In conclusion, Ssr1238 specifically binds to two different transcripts and participates in the coordination of RNA maturation, translation, cell division, and aspects of nitrogen metabolism. Our results are consistent with recent findings that theB. subtilisYlxR protein functions as an RNase P modulator (RnpM), but suggest additional functionalities and extend its proposed role to the phylum cyanobacteria.
Demonstration of chemotherapeutic mediated lymphatic changes in meningeal lymphatics in vitro, ex vivo, and in vivo
Authors: Roberts L. Monet , Hammel Jennifer H , Azar Francesca , Feng Tzu-Yu (Alkaid) , Cunningham Jessica J. , Rutkowski Melanie , Munson Jennifer
Published: January 2024

Systemic chemotherapeutics target cancer cells but are also known to impact other cells away from the tumor. Questions remain whether systemic chemotherapy crosses the blood-brain barrier and causes inflammation in the periphery that impacts the central nervous system (CNS) downstream. The meningeal lymphatics are a critical component that drain cerebrospinal fluid from the CNS to the cervical lymph nodes for immunosurveillence. To develop new tools for understanding chemotherapy-mediated effects on the meningeal lymphatics, we present two novel models that examine cellular and tissue level changes. Our in vitro tissue engineered model of a meningeal lymphatic vessel lumen, using a simple tissue culture insert system with both lymphatic endothelial and meningeal cells, examines cell disruption. Our ex vivo model culturing mouse meningeal layers probes structural changes and remodeling, correlating to an explant tissue level. To gain a holistic understanding, we compare our in vitro and ex vivo models to in vivo studies for validation and a three-tier methodology for examining the chemotherapeutic response of the meningeal lymphatics. We have demonstrated that the meningeal lymphatics can be disrupted by systemic chemotherapy but show differential responses to platinum and taxane chemotherapies, emphasizing the need for further study of off-target impacts in the CNS.
Behaviour-based movement cut-off points in 3-year old children comparing wrist- with hip-worn actigraphs MW8 and GT3X
Authors: Jansson Daniel , Westlander Rikard , Sandlund Jonas , West Christina E. , Domellöf Magnus , Wulff Katharina
Published: January 2024

Introduction

Behaviour-based physical intensities have not undergone rigorous calibration in long-term recordings of 3-year-old children’s sleep/activity patterns. This study aimed at (i) calibrating activity counts of motor behaviour measured simultaneously with MotionWatch 8 (MW8) and ActiGraph (GT3X) in 3-year-old children, (ii) documenting movement intensities in 30s-epochs at wrist/hip positions, and (iii) evaluating the accuracy of cut-off agreements between different behavioural activities.

Methods

Thirty 3-year-old children of the NorthPop cohort performed six directed behavioural activities individually, each for 8-10 minutes while wearing two pairs of devices at hip and wrist position. Directly observed naturally-occurring behaviours included: watching cartoons, recumbent story listening, sit and handcraft, floor play with toys, engaging in a walk and a sprinting game. Receiver-Operating-Curve classification was applied to determine activity count thresholds and to assign context-guided, physical activity composite classes.

Results

Activity counts of MW8 and GT3X pairs of wrist-worn (r = 0.94) and hip-worn (r = 0.79) devices correlated significantly (p < 0.001). Activity counts at hip position were significantly lower compared to those at the wrist position (p < 0.001), irrespective of device type. Sprinting, floorball/walk and floorplay assigned as ‘physicallymobile’classes achieved outstanding accuracy (AUC >0.9) and two sedentary and a motionless activities assigned into ‘physicallystationary’classes achieved excellent accuracy (AUC >0.8).

Conclusion

This study provides useful cut-offs for physical activity levels of preschool children using two different devices. Contextual information of behaviour is advantageous over intensity classifications only, because interventions reallocate time among behaviours, which allows to establish dose-response relationships between behavioural changes and health outcomes. Our comparative calibration is one step forward to inform behaviour-based public health guidelines for 3-year-old children.
Predicting Serotonin Detection with DNA-Carbon Nanotube Sensors Across Multiple Spectral Wavelengths
Authors: Kelich Payam , Adams Jaquesta , Jeong Sanghwa , Navarro Nicole , Landry Markita P. , Vuković Lela
Published: January 2024

Owing to the value of DNA-wrapped single-walled carbon nanotube (SWNT) based sensors for chemically-specific imaging in biology, we explore machine learning (ML) predictions DNA-SWNT serotonin sensor responsivity as a function of DNA sequence based on the whole SWNT fluorescence spectra. Our analysis reveals the crucial role of DNA sequence in the binding modes of DNA-SWNTs to serotonin, with a smaller influence of SWNT chirality. Regression ML models trained on existing datasets predict the change in the fluorescence emission in response to serotonin, ΔF/F, at over a hundred wavelengths for new DNA-SWNT conjugates, successfully identifying some high- and low-response DNA sequences. Despite successful predictions, we also show that the finite size of the training dataset leads to limitations on prediction accuracy. Nevertheless, incorporating entire spectra into ML models enhances prediction robustness and facilitates the discovery of novel DNA-SWNT sensors. Our approaches show promise for identifying new chemical systems with specific sensing response characteristics, marking a valuable advancement in DNA-based system discovery.
Synthetase and Hydrolase Specificity Collectively Excludes 2’-Deoxyguanosine from Bacterial Alarmone
Authors: Zhou Rich W. , Gordon Isis J. , Hei Yuanyuan , Wang Boyuan
Published: January 2024

In response to starvation, virtually all bacteria pyrophosphorylate the 3’-hydroxy group of GTP or GDP to produce two messenger nucleotides collectively denoted as (p)ppGpp. Also known as alarmones, (p)ppGpp reprograms bacterial physiology to arrest growth and promote survival. Intriguingly, although cellular concentration of dGTP is two orders of magnitude lower than that of GTP, alarmone synthetases are highly selective against using 2’-deoxyguanosine (2dG) nucleotides as substrates. We thus hypothesize that production of 2dG alarmone, (p)pp(dG)pp, is highly deleterious, which drives a strong negative selection to exclude 2dG nucleotides from alarmone signaling. In this work, we show that theB. subtilisSasB synthetase prefers GDP over dGDP with 65,000-fold higherkcat/Km, a specificity stricter than RNA polymerase selecting against 2’-deoxynucleotides. Using comparative chemical proteomics, we found that although most known alarmone-binding proteins inEscherichia colicannot distinguish ppGpp from pp(dG)pp, hydrolysis of pp(dG)pp by the essential hydrolase, SpoT, is 1,000-fold slower. This inability to degrade 2’-deoxy-3’-pyrophosphorylated substrate is a common feature of the alarmone hydrolase family. We further show that SpoT is a binuclear metallopyrophoshohydrolase and that hydrolysis of ppGpp and pp(dG)pp shares the same metal dependence. Our results support a model in which 2’-OH directly coordinates the Mn2+at SpoT active center to stabilize the hydrolysis-productive conformation of ppGpp. Taken together, our study reveals a vital role of 2’-OH in alarmone degradation, provides new insight on the catalytic mechanism of alarmone hydrolases, and leads to the conclusion that 2dG nucleotides must be strictly excluded from alarmone synthesis because bacteria lack the key machinery to down-regulate such products.
Acquisition phase-specific contribution of climbing fiber transmission to cerebellum-dependent motor memory
Authors: Seo Jewoo , Kim Seung Ha , Lee Jaegeon , Kim Min Seok , Lee Yong-Seok , Kim Sang Jeong
Published: January 2024

Climbing fiber (CF) transmission from the inferior olivary triggers complex spikes (Cs) in Purkinje cells (PCs) driven by a burst of calcium spikes. In the context of motor learning, especially the compensatory optic response, CF transmission serves as instructive signals selectively conveyed to PCs. While the significance of CF input in motor memory formation is widely acknowledged, a comprehensive understanding of its distinct contribution across different temporal windows, spanning from the initial learning phase to the retrieval period, remains incomplete. Therefore, we aimed to investigate the necessity of CF-induced instructive signals in motor learning by assessing their roles in memory acquisition, consolidation, and retrieval. We employed optogenetics to selectively inhibit CF transmission during targeted time windows. Consequently, the absence of CF-induced instructive signals during motor learning impairs memory acquisition. However, when these signals were suppressed during the consolidation and retrieval period, there was neither a loss of long-term memory nor prevention of memory retrieval. Our results highlight that CF transmission plays a specialized and critical role primarily in memory acquisition, rather than subsequent processes.
Capsaicin acts as a novel NRF2 agonist to suppress ethanol induced gastric mucosa oxidative damage by directly disrupting the KEAP1-NRF2 interaction
Authors: Gao Xiaoning , Guo WuYan , Liu Peiyuan , Yuwen Mingyue , Liu Zixiang , Tan Ruyang , Liu Kairui , Yang Zhiru , Ba Junli , Bai Xue , Shama Shiti , Tang Cong , Miao Kai , Pei Haozhi , Liu Liren , Zhu Cheng , Wang Tao , Zhang Bo , Kang Jun
Published: January 2024

Excessive drinking poses serious health risks and is closely associated with oxidative damage. The KEAP1-NRF2-ARE axis serves as the primary antioxidant system. However, the existing small molecule inhibitors are all covalently bound to KEAP1, meaning that once bound, they are not easily dissociated, while continuous inhibition of KEAP1 exhibits severe side effects. In this study, BLI, CETSA, Pull-down, Co-IP and HDX-MS assay analysis were conducted to detect the KEAP1 binding behavior of natural product, capsaicin (CAP), bothin vitroand in GES-1 cells. The ethanol-induced acute gastric mucosal damage rat model was also established to determine the therapeutic effect of CAP. We demonstrated that CAP ameliorated mitochondrial damage, facilitated the nuclear translocation of NRF2, thereby promoting the expression of downstream antioxidant response elements, HO-1, Trx, GSS and NQO1 in GES-1 cells. Subsequently, CAP could directly bind to KEAP1 and inhibit the interaction between KEAP1 and NRF2. While in the KEAP1-knockout 293T cells, CAP failed to activate NRF2 expression. It was also found that CAP non-covalently bound to Kelch domain and allosterically regulated three regions of KEAP1:L342-L355, D394-G423 and N482-N495. To enhance drug solubility and delivery efficiency, we designed IR-Dye800 modified albumin coated CAP nanoparticle. The nanoparticles significantly alleviated the gastric mucosal inflammation and activated the NRF2 downstream genesin vivo. Finally, we further verified our hypothesis in Nfe2l2-knockout mice. Our work provided new insights that CAP is a safe and novel NRF2 agonist by allosterically regulating KEAP1, which may contribute to the development of lead drugs for oxidative stress-related illness, e.g. aging, cancer, neurodegenerative and cardiovascular diseases.

Graphic diagram
Label-free long-term measurements of adipocyte differentiation from patient-driven fibroblasts and quantitative analyses of in situ lipid droplet generation
Authors: Jeong Eun Young , Kim Hye-Jin , Lee Sumin , Park Yongkeun , Kim Young Mo
Published: January 2024

The visualization and tracking of adipocytes and their lipid droplets (LDs) during differentiation are pivotal in developmental biology and regenerative medicine studies. Traditional staining or labeling methods, however, pose significant challenges due to their labor-intensive sample preparation, potential disruption of intrinsic cellular physiology, and limited observation timeframe. This study introduces a novel method for long-term visualization and quantification of biophysical parameters of LDs in unlabeled adipocytes, utilizing the refractive index (RI) distributions of LDs and cells. We employ low-coherence holotomography (HT) to systematically investigate and quantitatively analyze the 42-day redifferentiation process of fat cells into adipocytes. This technique yields three-dimensional, high-resolution refractive tomograms of adipocytes, enabling precise segmentation of LDs based on their elevated RI values. Subsequent automated analysis quantifies the mean concentration, volume, projected area, and dry mass of individual LDs, revealing a gradual increase corresponding with adipocyte maturation. Our findings demonstrate that HT is a potent tool for non-invasively monitoring live adipocyte differentiation and analyzing LD accumulation. This study, therefore, offers valuable insights into adipogenesis and lipid research, establishing HT and image-based analysis as a promising approach in these fields.
Impact of high fat diet and exercise on bone and bile acid metabolism in rats
Authors: Alonso Nerea , Almer Gunter , Semeraro Maria Donatella , Rodriguez-Blanco Giovanny , Fauler Günter , Anders Ines , Ritter Gerald , vom Scheidt Annika , Hammer Niels , Gruber Hans Jürgen , Herrmann Markus
Published: January 2024

Background

Bile acids help facilitate the intestinal lipids absorption, and have endocrine activity in glucose, lipid and bone metabolism. Obesity and exercise influence bile acid metabolism and have opposite effects in bone. This given study investigates if regular exercise helps mitigate the adverse effects of obesity on bone, potentially by reversing alterations in bile acid metabolism.

Methods

Four-month-old female Sprague Dawley rats either received high fat diet (HFD) or chow-based standard diet (lean controls). During the 10-month study period, half of the animals performed 30 min of running at moderate speed on five consecutive days followed by two days of rest. The other half was kept inactive (inactive controls). At study end, bone quality was assessed by micro computed tomography and biomechanical testing. Bile acids were measured in serum and stool.

Results

HFD feeding was related to reduced trabecular (−33%, p=1.14×10-7) and cortical (−21%, p=2.9×10-8) bone mass, and lowered femoral stiffness (12-41%, p=0.005). Furthermore, HFD decreased total bile acids in serum (−37%, p=1.0×10-6), but increased bile acids in stool (+2-fold, p=7.3×10-9). These quantitative effects were accompanied by changes in the relative abundance of individual bile acids. The concentration of serum bile acids correlated positively with all cortical bone parameters (r=0.593 - 0.708), whilst stool levels showed inverse correlations at cortical (r=-0.651 - −0.805) and trabecular level (r=-0.656 - −0.750). Exercise improved some trabecular and cortical bone quality parameters (+11-31%, p=0.043 to 0.001) in lean controls, but failed to revert the bone loss related to HFD. Similarly, changes in bile acid metabolism were not mitigated by exercise.

Conclusion

Prolonged HFD consumption induced quantitative and qualitative alterations in bile acid metabolism, accompanied by bone loss. Tight correlations between bile acids and structural indices of bone quality support further functional analyses on the potential role of bile acids in bone metabolism. Regular moderate exercise improved trabecular and cortical bone quality in lean controls but failed mitigating the effects related to HFD in bone and bile acid metabolism.
The Emerging Role of 3-Hydroxyanthranilic Acid onC. elegansAging Immune Function
Authors: Espejo Luis S. , DeNicola Destiny , Chang Leah M. , Hofschneider Vanessa , Haskins Anne E. , Balsa Jonah , Freitas Samuel S. , Antenor Angelo , Hamming Sage , Hull Bradford , Castro-Portuguez Raul , Dang Hope , Sutphin George L.
Published: January 2024

3-hydroxyanthranilic acid (3HAA) is considered to be a fleeting metabolic intermediate along tryptophan catabolism through the kynurenine pathway. 3HAA and the rest of the kynurenine pathway have been linked to immune response in mammals yet whether it is detrimental or advantageous is a point of contention. Recently we have shown that accumulation of this metabolite, either through supplementation or prevention of its degradation, extends healthy lifespan inC. elegansand mice, while the mechanism remained unknown. UtilizingC. elegansas a model we investigate how 3HAA andhaao-1inhibition impact the host and the potential pathogens. What we find is that 3HAA improves host immune function with aging and serves as an antimicrobial against gram-negative bacteria. Regulation of 3HAA’s antimicrobial activity is accomplished via tissue separation. 3HAA is synthesized in theC. eleganshypodermal tissue, localized to the site of pathogen interaction within the gut granules, and degraded in the neuronal cells. This tissue separation creates a new possible function for 3HAA that may give insight to a larger evolutionarily conserved function within the immune response.

Graphical Abstract
Allostery at a protein-protein interface harboring an intermolecular dynamic network
Authors: Gomez Sara Medina , Vasa Suresh Kumar , Linser Rasmus
Published: January 2024

Motional properties of individual amino acids in proteins are strongly modulated by their specific surrounding. The dynamics of tightly interacting residues can formintramolecular dynamic networks, which influence various features of protein function and serve as an access point for their modulation within signaling cascades. However, the possible formation ofintermolecular networks shared between natural or constructed interaction partners has escaped thorough experimental assessment. Here, using fast-MAS solid-state NMR spectroscopy, we contrast the absence of a cross-talk between different residues in an apo protein with a recoupling of μs timescale dynamics effective via a mediating crystal-crystal contact. The data show that dynamic allostery is not necessarily restricted to motionally coupled elements within a single protein but can traverse molecular boundaries. Interrogation of intermolecular dynamic networks by the strategies proposed here may shed light on the mechanisms underlying allosteric modulation of protein function in biological, pharmacological, and biotechnological studies.
The holocephalan ratfish endoskeleton shares trabecular and areolar mineralization patterns, but not tesserae, with elasmobranchs little skate and catshark, and appears paedomorphic
Authors: Atake Oghenevwogaga J. , Berio Fidji , Debiais-Thibaud Melanie , Eames B. Frank
Published: January 2024

Specific character traits of mineralized endoskeletal tissues need to be clearly defined and comprehensively examined among extant chondrichthyans (elasmobranchs, such as sharks and skates, and holocephalans, such as chimaeras) to understand their evolution. For example, tiles of mineralized polygonal structures called tesserae occur at cartilage surfaces in chondrichthyans, but recent studies showing trabecular mineralization at elasmobranch cartilage surfaces suggest that tesserae are not as common as previously thought. Also, while areolar mineralized tissue in elasmobranchs is generally considered a unique, shared chondrichthyan feature, some chondrichthyan species demonstrate bone-like tissues in both a specific region of tesserae termed the cap zone and continuous (not tiled) mineralized neural arches. To clarify the distribution of specific endoskeletal features among extant chondrichthyans, adult skeletal tissues in a holocephalan chimaera (spotted ratfish) and two elasmobranchs (small-spotted catshark and little skate) were characterized using synchrotron radiation and desktop micro-CT imaging, and histological and immunofluorescent assays. Endoskeletal mineralization in the ratfish, catshark, and little skate varied both quantitively in tissue mineral density (TMD), and qualitatively in the morphology and localization of mineralized structures and tissues. For example, TMD of several skeletal elements was significantly lower in ratfish, compared to catshark and little skate. Trabecular and areolar mineralization were shared among these extant chondrichthyan species, but tesserae and bone-like tissues were not. Interestingly, three separate analyses argued that the chimaera endoskeleton retains embryonic features (i.e., paedomorphic with respect to elasmobranchs). This study further proposes general terminology for character states of the extant chondrichthyan endoskeleton and infers those states in ancestral chondrichthyans with reference to fossil data.
Analysis of theBorreliaceaePangenome Reveals a Conserved Genomic Archictecture Across Phylogenetic Scales
Authors: Lemieux Jacob E.
Published: January 2024

The FamilyBorreliaceaecontains arthropod-borne spirochetes that cause two widespread human diseases, Lyme disease (LD) and relapsing fever (RF). LD is a subacute, progressive illness with variable stage and tissue manifestations. RF is an acute febrile illness with prominent bacteremia that may recur and disseminate, particularly to the nervous system. Clinical heterogeneity is a hallmark of both diseases. While human clinical manifestations are influenced by a wide variety of factors, including immune status and host genetic susceptibility, there is evidence thatBorreliaceaemicrobial factors influence the clinical manifestations of human disease caused by this Family of spirochetes. Despite these associations, the spirochete genes that influence the severity and manifestations of human disease are, for the most part, unknown. Recent work has identified lineage-specific expansions of lipoproteome-rich accessory genome elements in virulent clones ofB. burgdorferi. Using publicly available genome assemblies, I show here that allBorreliaceaelineages for which sufficient sequence data is available harbor a similar pattern of strongly structured, lineage-specific expansions in their accessory genome, particularly among lipoproteins, and that this pattern holds across phylogenetic scales including genera, species, and genotypes. The relationships among pangenome elements suggest that infrequent episodes of marked genomic change followed by clonal expansion in geographically and enzootically structured populations may account for the unique lineage structure ofBorreliaceae. This analysis informs future genotype-phenotype studies amongBorreliaceaeand lays a foundation for studies of individual gene function guided by phylogenetic patterns of conservation, diversification, gain, and/or loss.
TDP-43 pathology links innate and adaptive immunity in amyotrophic lateral sclerosis
Authors: Evangelista Baggio A. , Ragusa Joey V. , Pellegrino Kyle , Wu Yija , Quiroga-Barber Ivana Yoseli , Cahalan Shannon R. , Arooji Omeed K. , Madren Jillann A. , Schroeter Sally , Cozzarin Joe , Xie Ling , Chen Xian , White Kristen K. , Ezzell J. Ashley , Iannone Marie A. , Cohen Sarah , Traub Rebecca E. , Li Xiaoyan , Bedlack Richard , Phanstiel Douglas H. , Meeker Rick , Stanley Natalie , Cohen Todd J.
Published: January 2024

Amyotrophic lateral sclerosis is the most common fatal motor neuron disease. Approximately 90% of ALS patients exhibit pathology of the master RNA regulator, Transactive Response DNA Binding protein (TDP-43). Despite the prevalence TDP-43 pathology in ALS motor neurons, recent findings suggest immune dysfunction is a determinant of disease progression in patients. Whether TDP-43 pathology elicits disease-modifying immune responses in ALS remains underexplored. In this study, we demonstrate that TDP-43 pathology is internalized by antigen presenting cells, causes vesicle rupture, and leads to innate and adaptive immune cell activation. Using a multiplex imaging platform, we observed interactions between innate and adaptive immune cells near TDP-43 pathological lesions in ALS brain. We used a mass cytometry-based whole-blood stimulation assay to provide evidence that ALS patient peripheral immune cells exhibit responses to TDP-43 aggregates. Taken together, this study provides a novel link between TDP-43 pathology and ALS immune dysfunction, and further highlights the translational and diagnostic implications of monitoring and manipulating the ALS immune response.
A developmental brain-wide screen identifies retrosplenial cortex as a key player in the emergence of persistent memory
Authors: Jin Benita , Gongwer Michael W , Kearney Brian P , Ohanian Lilit , Holden-Wingate Lucinda , Le Bryan , Darmawan Alfonso , Nakayama Yuka , Mora Sophia A Rueda , DeNardo Laura A
Published: January 2024

Memories formed early in life are short-lived while those formed later persist. Recent work revealed that infant memories are stored in a latent state. But why they fail to be retrieved is poorly understood. Here we investigated brain-wide circuit mechanisms underlying infantile amnesia. We performed a screen that combined contextual fear conditioning, activity-dependent neuronal tagging at different postnatal ages, tissue clearing and light sheet microscopy. We observed striking developmental changes in regional activity patterns between infant, juvenile, and adult mice, including changes in the retrosplenial cortex (RSP) that aligned with the emergence of persistent memory. We then performed a series of targeted investigations of RSP structure and function across development. Chronic chemogenetic reactivation of tagged RSP ensembles during the week after learning enhanced memory in adults and juveniles, but not in infants. However, after 33 days, reactivating infant-tagged RSP ensembles recovered forgotten memories. Changes in the developmental functions of RSP memory ensembles were accompanied by changes in dendritic spine density and the likelihood that those ensembles could be reactivated by contextual cues. These studies show that RSP ensembles store latent infant memories, reveal the time course of RSP functional maturation, and suggest that immature RSP functional networks contribute to infantile amnesia.
Modeling Combination Chemo-Immunotherapy for Heterogeneous Tumors
Authors: Chen Shaoqing , Hu Zheng , Zhou Da
Published: January 2024

Hypermutable cancers create opportunities for the development of various immunotherapies, such as immune checkpoint blockade (ICB) therapy. However, emergent studies have revealed that many hypermutated tumors have poor prognosis due to heterogeneous tumor antigen landscapes, yet the underlying mechanisms remain poorly understood. To address this issue, we developed mathematical models to explore the impact of combining chemotherapy and ICB therapy on heterogeneous tumors. Our results uncover how chemotherapy reduces antigenic heterogeneity, creating improved immunological conditions within tumors, which, in turn, enhances the therapeutic effect when combined with ICB. Furthermore, our results show that the recovery of the immune system after chemotherapy is crucial for enhancing the response to chemo-ICB combination therapy.

Author summary

The challenge posed by intratumoral heterogeneity is gaining recognition in the field of cancer treatment. Despite the success of immune checkpoint blockade (ICB) therapies in enhancing overall survival across various cancer types, the complexity of therapeutic responses persists due to the heterogeneity of tumor antigens. In this study, we developed mathematical models to explore the evolutionary dynamics of tumors with both homogeneous and heterogeneous antigen landscapes. Our analysis reveals that tumors with heterogeneity exhibit resistance to ICB therapy, unlike their homogeneous counterparts which respond positively. Additionally, our models demonstrate that early treatment of heterogeneous tumors with chemotherapy leads to significant remission but also rapid recurrence. Notably, we identified a fascinating trade-off associated with chemotherapy—while suppressing the immune system, it creates a tumor immunological environment that becomes more conducive to ICB therapy. Finally, our modeling highlights the augmented response observed in tumors subjected to a chemo-ICB combination and shows the crucial role of immune recovery in the context of combination therapy.
Maturation and conformational switching of a de novo designed phase-separating polypeptide
Authors: Hilditch Alexander T. , Romanyuk Andrey , Hodgson Lorna R. , Mantell Judith , Neal Chris R. , Verkade Paul , Obexer Richard , Serpell Louise C. , McManus Jennifer J. , Woolfson Derek N.
Published: January 2024

Cellular compartments formed by biomolecular condensation are a widespread feature of cell biology. These organelle-like assemblies compartmentalize macromolecules dynamically within the crowded intracellular environment. However, the intermolecular interactions that produce condensed droplets may also create arrested states and potentially pathological assemblies such as fibers, aggregates, and gels, through droplet maturation. Protein liquid-liquid phase separation is a metastable process, so maturation may be an intrinsic property of phase-separating proteins, where nucleation of different phases or states arise in supersaturated condensates. Here, we describe the formation of both phase-separated droplets and proteinaceous fibers driven by a de novo designed polypeptide. We characterize the formation of supramolecular fibers in vitro and in bacterial cells. We show that client proteins can be targeted to the fibers in cells using the droplet-forming construct. Finally, we explore the interplay between phase separation and fiber formation of the de novo polypeptide, showing that the droplets mature with a post-translational switch to largely β conformations, analogous to models of pathological phase separation.
Thermodynamic Characterization of LC3/GABARAP Ligands by Isothermal Titration Calorimetry
Authors: Dopfer Johannes , Schwalm Martin P. , Knapp Stefan , Rogov Vladimir V.
Published: January 2024

Isothermal titration calorimetry (ITC) is a widely used technique for the characterization of protein-protein and protein-ligand interactions. It provides information on the stoichiometry, affinity, and the thermodynamic driving forces of interactions. This chapter exemplifies the use of ITC to investigate interactions between human autophagy modifiers (LC3/GABARAP proteins) and their interaction partners, the LIR motif containing sequences. The purpose of this report is to present a detailed protocol for the production of LC3/GABARAP-interacting LIR peptides usingE. coliexpression systems. In addition, we outline the design of ITC experiments using the LC3/GABARAP:peptide interactions as an example. Comprehensive troubleshooting notes are provided to facilitate the adaptation of these protocols to different ligand-receptor systems. The methodology outlined for studying protein-ligand interactions will help to avoid common errors and misinterpretations of experimental results.