bluuebunny/crossref_metadata_2025 huggingface dataset:
Urea amidolyase as the enzyme for urea utilization in algae: functional display inChlamydomonas reinhardtiiand evolution in algae
Authors: Liang Honghao , Lin Senjie , Chen Yuanhao , Wang Jingtian , Aslam Muhammad , Chen Jing , Du Hong , Li Tangcheng
Published: May 2024

Urea is a crucial nitrogen nutrient source for algae with the potential to stimulate harmful algal blooms, but the molecular machinery underpinning urea uptake and assimilation by algae is not fully understood. Urease (URE) is commonly regarded as the responsible enzyme, but the urea amidolyase (UAD) system, albeit known to exist, has hardly been studied. Here, the phylogenetic distribution, expression patterns, and functional roles of UAD system are examined, which comprises subunitsDUR1,DUR2, andDUR3. We find a widespread occurrence of UAD, spanning four major phytoplankton lineages, and potentially independent evolution of URE and lineage-specific loss. Besides, a stronger regulation of UAD by environmental nitrogen concentrations compared to URE is uncovered in both global ocean and local dinoflagellate-dominant bloom events. CRISPR-based mutation inChlamydomonas reinhardtiishows that subunitDUR2is essential for urea utilization.DUR2inactivation led to completely growth restriction and upregulation ofDUR1andDUR3A, suggesting its functional interaction with them. In contrast,DUR3Binactivation only partially halted urea uptake and cell growth but significantly reduced gene expression across the entire UAD system. These findings not only reveal the crucial role ofDUR2in urea utilization inC. reinhardtiiand potentially in many other algae, but also suggestDUR2to be a more suitable indicator of urea utilization than urease, and underscore the importance to consider both URE and UAD enzyme systems when urea utilization by algae is assessed.
A receptor kinase complex refines cambium activity inArabidopsis
Authors: He Qing , Han Jingyi , Wei Wenbin , Pohl Ehmke , Wightman Raymond , Etchells J Peter
Published: May 2024

In plant development, key receptor kinases are often active in disparate cell types, with each requiring vastly different signalling outputs. The ERECTA (ER) receptor kinase and its homologs ERL1 and ERL2 exemplify this pleiotropy. InArabidopsis, they influence stomatal patterning, shoot meristem function, ovule morphogenesis, xylem fiber differentiation, and cell division in the vascular cambium1–6. Such diverse expression and functionality raises the question of how ER signalling can specify such distinct cell behaviours. One mechanism is via cell-type specific interactions with co-receptors, ligands, or other proteins that modulate signalling. However, little is known about ER interactors in the vascular cambium, a bifacial stem cell niche that generates phloem and xylem (Figure 1A). Combinatorial mutations betweenER, ERL1andERL2and receptor kinases of a second family,PXY, PXL1, andPXL2, show severe cambial defects5,7, but the mechanism underpinning these phenotypes is not known. Here we discovered that PXY proteins form protein complexes with ER and ERL2. PXY signalling can be manipulated by altering levels of its cognate ligand, TDIF. In genetic analysis, plant lines in which TDIF levels were altered had dramatic phenotypic changes that required the presence of ER or ERL2. Our results demonstrate that PXY signalling mediated cambium regulation depends on ER signalling and explains ER function in the cambium. Because the cambium produces xylem, which constitutes the wood in vascular plants, our findings position PXY-ER complexes at the centre of the accumulation of this versatile biomaterial and essential carbon sink.
The dithiocarbamate pesticides maneb and mancozeb disturb the metabolism of lipids and xenobiotics in an in vitro model of metabolic dysfunction-associated steatotic liver disease
Authors: Petitjean Kilian , Dicara Giovanna , Bristeau Sébastien , Coppens-Exandier Hugo , Amalric Laurence , Baran Nicole , Savary Camille C. , Corlu Anne , Loyer Pascal , Fromenty Bernard
Published: May 2024

Pesticides are increasingly recognized to be hepatotoxic but less is known about their toxicity in metabolic dysfunction-associated steatotic liver disease (MASLD). Recent investigations reported oxidative stress-induced apoptosis in differentiated hepatocyte-like HepaRG cells after a single treatment with a 7-pesticide mixture that included chlorpyrifos, dimethoate, diazinon, iprodione, imazalil, and the dithiocarbamates maneb and mancozeb. These effects were reproduced by maneb, mancozeb, or manganese chloride (MnCl2). Herein, differentiated HepaRG cells cultured for 2 weeks without (-FA) or with (+FA) a mixture of stearic and oleic acids were treated with this 7-pesticide mixture, maneb, mancozeb, or MnCl2along the same period. While these molecules did not induce neutral lipid accumulation in -FA-HepaRG cells, they worsened steatosis in +FA-HepaRG cells. Maneb or MnCl2impaired very low-density lipoprotein (VLDL) secretion and increased fatty acid uptake without altering mitochondrial fatty acid oxidation andde novolipogenesis. Reduced VLDL secretion was associated with decreased mRNA levels of apolipoproteins B and C3 and microsomal triglyceride transfer protein. Zinc supplementation restored VLDL secretion, reduced fatty acid uptake and prevented the exacerbation of steatosis in +FA-HepaRG cells treated with mancozeb or MnCl2. The mixture, maneb, or MnCl2also reduced the mRNA expression and activity of several cytochromes P450 in +FA- and -FA-HepaRG cells. This was associated with impaired biotransformation of diazinon while chlorpyrifos metabolism was unaffected. Hence, maneb, mancozeb and MnCl2disturb the metabolism of lipids and xenobiotics in HepaRG cells, in particular in fatty acid-exposed cells. These findings could have major pathophysiological consequences in dithiocarbamate-exposed individuals with MASLD.
Hybrid lipid-block copolymer membranes enable stable reconstitution of a wide range of nanopores and robust sampling of serum
Authors: Vreeker Edo , Grünewald Fabian , van der Heide Nieck Jordy , Marrink Siewert-Jan , Tych Katarzyna (Kasia) , Maglia Giovanni
Published: May 2024

Biological nanopores are powerful tools for detecting biomolecules at the single-molecule level, making them appealing as sensors for biological samples. However, the lipid membranes in which nanopores reside can be unstable in the presence of biological fluids. Here, membranes formed with the amphiphilic polymers PMOXA-PDMS-PMOXA and PBD-PEO are tested as potential alternatives for nanopore sensing. We demonstrate that polymer membranes can possess increased stability towards applied potentials and high concentrations of human serum, but that the stable insertion of a wide range of biological nanopores is most often compromised. Alternatively, hybrid polymer-lipid membranes comprising a 1:1 w/w mixture of PBD11PEO8and DPhPC showed high electrical and biochemical stability while creating a suitable environment for all tested nanopores. Analytes such as proteins, DNA and sugars were efficiently sampled, indicating that in hybrid membranes nanopores showed native-like properties. Molecular dynamics simulations revealed that lipids form ∼12 nm domains interspersed by a polymer matrix. Nanopores partitioned into these lipid nanodomains and sequestered lipids possibly offering the same binding strength as in a native bilayer. This work shows that single-molecule analysis with nanopores in [PBD11PEO8+ DPhPC] membranes is feasible and present stable recordings in the presence of human serum. These results pave the way towards novel nanopore-based biosensors.
In vivo Targeting MEK and TNK2/SRC pathways in PTPN11 driven leukemia
Authors: Chang Bill H. , Thiel-Klare Karina , Tyner Jeffrey W.
Published: May 2024

PTPN11 encodes for a tyrosine phosphatase implicated in the pathogenesis of hematologic malignancies such as Juvenile Myelomonocytic Leukemia (JMML), Acute Myeloid Leukemia (AML), and Acute Lymphoblastic Leukemia (ALL). Since activating mutations of PTPN11 increase proliferative signaling and cell survival through the RAS/MAPK proliferative pathway there is significant interest in using MEK inhibitors for clinical benefit. Yet, single agent clinical activity has been minimal. Previously, we showed that PTPN11 is further activated by upstream tyrosine kinases TNK2/SRC, and that PTPN11-mutant JMML and AML cells are sensitive to TNK2 inhibition using dasatinib. In order to validate these findings, we adopted a genetically engineered mouse model of PTPN11 driven leukemia using the mouse strain 129S/Sv-Ptpn11tm6Bgn/Mmucd crossed with B6.129P2-Lyz2tm1(cre)Ifo/J. The F1 progeny expressing Ptpn11D61Ywithin hematopoietic cells destined along the granulocyte-monocyte progenitor lineage developed a fatal myeloproliferative disorder characterized by neutrophilia and monocytosis, and infiltration of myeloid cells into the liver and spleen. Cohorts of Ptpn11D61Yexpressing animals treated with combination of dasatinib and trametinib for an extended period of time was well tolerated and had a significant effect in mitigating disease parameters compared to single agents. Finally, a primary patient-derived xenograft model using a myeloid leukemia with PTPN11 F71L also displayed improved disease response to combination. Collectively, these studies point to combined therapies targeting MEK and TNK2/SRC as a promising therapeutic potential for PTPN11-mutant leukemias.

Key Points

Combining MEK and TNK2/SRC inhibitors has therapeutic potential in PTPN11 mutant JMML and AML
Deep mutational scanning of influenza A virus NEP reveals pleiotropic mutations in its N-terminal domain
Authors: Teo Qi Wen , Wang Yiquan , Lv Huibin , Mao Kevin J. , Tan Timothy J.C. , Huan Yang Wei , Rivera-Cardona Joel , Shao Evan K. , Choi Danbi , Dargani Zahra Tavakoli , Brooke Christopher B. , Wu Nicholas C.
Published: May 2024

The influenza A virus nuclear export protein (NEP) is a multifunctional protein that is essential for the viral life cycle and has very high sequence conservation. However, since the open reading frame of NEP largely overlaps with that of another influenza viral protein, non-structural protein 1, it is difficult to infer the functional constraints of NEP based on sequence conservation analysis. Besides, the N-terminal of NEP is structurally disordered, which further complicates the understanding of its function. Here, we systematically measured the replication fitness effects of >1,800 mutations of NEP. Our results show that the N-terminal domain has high mutational tolerance. Additional experiments demonstrate that N-terminal domain mutations pleiotropically affect viral transcription and replication dynamics, host cellular responses, and mammalian adaptation of avian influenza virus. Overall, our study not only advances the functional understanding of NEP, but also provides insights into its evolutionary constraints.
Self-assembly of microstructured protein coatings with programmable functionality for fluorescent biosensors
Authors: Jo Suna , Pearson Erin , Yoon Donghoon , Kim Jungkwun , Park Won Min
Published: May 2024

Proteins, as genetically programmable functional macromolecules, hold immense potential as biocompatible self-assembling building blocks. Despite their versatility in building coating materials, it has been often hindered from programming their functionality genetically. In this study, we demonstrate a modular self-assembly of protein coatings that are genetically programmable for a biosensor application. We designed recombinant fusion protein building blocks to form microstructured coatings on diverse substrates, such as glass or polymer, through a thermally triggered liquid-liquid phase separation and an orthogonal high-affinity coiled-coil interaction. We incorporated fluorescence proteins into coatings and controlled protein density to enable fluorescence imaging and quantification in a low-resource setting. Then, we created a coating for a calcium biosensor using a genetically engineered calcium indicator protein. This protein coating served as the foundation for our smartphone-based fluorescence biosensor, which successfully measured free calcium concentrations in the millimolar range at which extracellular calcium homeostasis is maintained. Using this fluorescence biosensor, we were able to detect abnormal physiological conditions such as mild or moderate hypercalcemia. We envision that this modular and genetically programmable functional protein coating platform could be extended to the development of highly accessible, low-cost fluorescent biosensors for a variety of targets.
Effect of SNEDDS Loading Pentagamavunon-0 on Memory Impairment and Neurogenesis in Mice with Monosodium Glutamate-Induced Alzheimer’s Disease-Like Symptoms
Authors: Anas Yance , Susidarti Ratna Asmah , Martien Ronny , Yuniarti Nunung
Published: May 2024

Pentagamavunon-0 (PGV-0), a curcumin analog, has antioxidant and HDAC2 inhibitory properties. It affects neurogenesis and cognitive function in Alzheimer’s disease (AD). Targeting neurogenesis is a strategy currently being developed for AD treatment. This study investigates the therapeutic potential of self-nanoemulsifying drug delivery systems loaded with PGV-0 (SNEDDS PGV-0) on spatial learning, memory impairment, and gene expression-related neurogenesis in mice with monosodium glutamate (MSG) induced AD-like symptoms. MSG 4 g/kg was injected (sc.) into 4-week-old Balb/C mice seven times every alternate day, followed by treatment for 60 days with CMC-Na 0.5%, PGV-0 suspension, SNEDDS PGV-0, and donepezil-HCl. The open-field test, novel object recognition, and 8-radial arm maze test were employed to assess the neurobehavioral performance of mice. Neurogenesis-related gene expression (dcx, nestin, Hes5, and NFIA) was quantified through qPCR analysis. Spatial learning and memory in mice declined with MSG treatment. SNEDDS PGV-0 effectively alleviates mice’s cognitive and memory deficits, as shown in improvements in behavioral parameters, including the Discrimination Index, Recognition Index, and Memory Score. It also restores mRNA expression of several essential neurogenesis-related genes, including dcx, Hes5, and NFIA. SNEDDS PGV-0 demonstrates promising potential as a neurogenesis promoter, making it a viable drug candidate for AD or other neurodegenerative pathologies.
Frequent and asymmetric cell division in endosymbiotic bacteria of cockroaches
Authors: Noda Tomohito , Mizutani Masaki , Harumoto Toshiyuki , Katsuno Tatsuya , Koga Ryuichi , Fukatsu Takema
Published: May 2024

Many insects are obligatorily associated with and dependent on specific microbial species as essential mutualistic partners. In the host insects, such microbial mutualists are usually maintained in specialized cells or organs, called bacteriocytes or symbiotic organs. Hence, potentially exponential microbial growth cannot be realized but must be strongly constrained by spatial and resource limitations within the host cells or tissues. How such endosymbiotic bacteria grow, divide and proliferate is important for understanding the interactions and dynamics underpinning intimate host-microbe symbiotic associations. Here we report thatBlattabacterium, the ancient and essential endosymbiont of cockroaches, exhibits unexpectedly high rates of cell division (20-58%) and, in addition, the cell division is asymmetric (average asymmetry index > 1.5) when isolated from the German cockroachBlattella germanica. The asymmetric division of endosymbiont cells at high frequencies was observed irrespective of host tissues (fat bodies vs. ovaries) or developmental stages (adults vs. nymphs vs. embryos) ofB. germanica, and also observed in several different cockroach species. By contrast, such asymmetric and frequent cell division was observed neither inBuchnera, the obligatory bacterial endosymbiont of aphids, nor inPantoea, the obligatory bacterial gut symbiont of stinkbugs. Comparative genomics of cell division-related genes uncovered that theBlattabacteriumgenome lacks the Min system genes that determine the cell division plane, which may be relevant to the asymmetric cell division. These observations combined with comparative symbiont genomics provide insight into what processes and regulations may underpin the growth, division and proliferation of such bacterial mutualists continuously constrained under within-host conditions.

IMPORTANCE

Diverse insects are dependent on specific bacterial mutualists for their survival and reproduction. Due to the long-lasting coevolutionary history, such symbiotic bacteria tend to exhibit degenerative genomes and suffer uncultivability. Because of their microbiological fastidiousness, the cell division patterns of such uncultivable symbiotic bacteria have been poorly described. Here, using fine microscopic and quantitative morphometric approaches, we report that, although bacterial cell division usually proceeds through symmetric binary fission,Blattabacterium, the ancient and essential endosymbiont of cockroaches, exhibits frequent and asymmetric cell division. Such peculiar cell division patterns were not observed with other uncultivable essential symbiotic bacteria of aphids and stinkbugs. Gene repertoire analysis revealed that the molecular machineries for regulating the bacterial cell division plane are lost in theBlattabacteriumgenome, suggesting the possibility that the general trend toward the reductive genome evolution of symbiotic bacteria may underpin their bizarre cytological/morphological traits.
Unraveling the impact of host genetics and factors on the urinary microbiome in a young population
Authors: Zou Leying , Zhang Zhe , Chen Junhong , Guo Ruijin , Tong Xin , Ju Yanmei , Lu Haorong , Yang Huanming , Wang Jian , Zong Yang , Xu Xun , Jin Xin , Xiao Liang , Jia Huijue , Zhang Tao , Liu Xiaomin
Published: May 2024

The significance of the urinary microbiome in maintaining health and contributing to disease development is increasingly recognized. However, a comprehensive understanding of this microbiome and its influencing factors remains elusive. Utilizing whole metagenomic and whole-genome sequencing, along with detailed metadata, we characterized the urinary microbiome and its influencing factors in a cohort of 1579 Chinese individuals. Our findings unveil the distinctiveness of the urinary microbiome from other four body sites, delineating five unique urotypes dominated byGardnerella vaginalis, Sphingobium fluviale, Lactobacillus iners, Variovorax sp PDC80andAcinetobacter junii, respectively. We identified 108 host factors significantly influencing the urinary microbiome, collectively explaining 12.92% of the variance in microbial composition. Notably, gender-related factors, including sex hormones, emerged as key determinants in defining urotype groups, microbial composition and pathways, with the urinary microbiome exhibiting strong predictive ability for gender (AUC=0.843). Furthermore, we discovered 43 genome-wide significant associations between host genetic loci and specific urinary bacteria,Acinetobacterin particular, linked to 8 host loci (p< 5×10-8). These associations were also modulated by gender and sex hormone levels. In summary, our study provides novel insights into the impact of host genetics and other factors on the urinary microbiome, shedding light on its implications for host health and disease.
Prediction of Conformational Ensembles and Structural Effects of State-Switching Allosteric Mutants in the Protein Kinases Using Comparative Analysis of AlphaFold2 Adaptations with Sequence Masking and Shallow Subsampling
Authors: Raisinghani Nishank , Alshahrani Mohammed , Gupta Grace , Tian Hao , Xiao Sian , Tao Peng , Verkhivker Gennady
Published: May 2024

Despite the success of AlphaFold2 approaches in predicting single protein structures, these methods showed intrinsic limitations in predicting multiple functional conformations of allosteric proteins and have been challenged to accurately capture of the effects of single point mutations that induced significant structural changes. We systematically examined several implementations of AlphaFold2 methods to predict conformational ensembles for state-switching mutants of the ABL kinase. The results revealed that a combination of randomized alanine sequence masking with shallow multiple sequence alignment subsampling can significantly expand the conformational diversity of the predicted structural ensembles and capture shifts in populations of the active and inactive ABL states. Consistent with the NMR experiments, the predicted conformational ensembles for M309L/L320I and M309L/H415P ABL mutants that perturb the regulatory spine networks featured the increased population of the fully closed inactive state. On the other hand, the predicted conformational ensembles for the G269E/M309L/T334I and M309L/L320I/T334I triple ABL mutants that share activating T334I gate-keeper substitution are dominated by the active ABL form. The proposed adaptation of AlphaFold can reproduce the experimentally observed mutation-induced redistributions in the relative populations of the active and inactive ABL states and capture the effects of regulatory mutations on allosteric structural rearrangements of the kinase domain. The ensemble-based network analysis complemented AlphaFold predictions by revealing allosteric mediating centers that often directly correspond to state-switching mutational sites or reside in their immediate local structural proximity, which may explain the global effect of regulatory mutations on structural changes between the ABL states. This study suggested that attention-based learning of long-range dependencies between sequence positions in homologous folds and deciphering patterns of allosteric interactions may further augment the predictive abilities of AlphaFold methods for modeling of alternative protein sates, conformational ensembles and mutation-induced structural transformations.
Ensemble Meta-Learning using SVM for Improving Cardiovascular Disease Risk Prediction
Authors: Punn Narinder Singh , Dewangan Deepak Kumar
Published: May 2024

Cardiovascular diseases (CVDs) remain a leading cause of mortality worldwide, posing a significant public health challenge. Early identification of individuals at high risk of CVD is crucial for timely intervention and prevention strategies. Machine learning techniques are increasingly being applied in healthcare for their ability to uncover complex patterns within large, multidimensional datasets. This study introduces a novel ensemble meta-learning framework designed to enhance cardiovascular disease (CVD) risk prediction. The framework strategically combines the predictive power of diverse machine learning algorithms – logistic regression, K nearest neighbors, decision trees, gradient boosting, gaussian Naive Bayes and XGBoost. Predicted probabilities from these base models are integrated using support vector machine as meta-learner. Rigorous performance evaluation over publicly available dataset demonstrates the improved performance of this ensemble approach compared to individual. This research highlights the potential of ensemble meta-learning techniques to improve predictive modeling in healthcare.
Comparison of the Vaginal and Labial Dimensions of Nullipara Ethnic Chinese and Western Women
Authors: Stevens Lisa , Masteling Mariana , Srinivasa Raju Kruthi , Mastrovito Sara , Ashton-Miller James A. , DeLancey John O. L.
Published: May 2024

Introduction and Hypothesis

Vaginal dimensions have clinical and surgical implications. We sought to quantify the differences between vaginal and labial dimensions in healthy ethnic Chinese and Western women with normal pelvic organ support.

Methods

This is a cross-sectional study of a convenience sample of ethnic Chinese nullipara (n = 33) and Western nullipara (n= 33) women recruited for research purposes. For each subject, magnetic resonance imaging was used to quantify the vaginal and labial dimensions. Specifically, we identified the anterior and posterior vaginal wall, the outline of the cervix in the mid-sagittal and coronal planes, and the distance from the labia majora to the hymenal ring at the urethral meatus.

Results

The vaginal and labial dimensions of ethnic Chinese nullipara were 9-21 % smaller than those of Western nullipara; however, the relative orientation of the cervix to the vaginal opening was not different.

Conclusions

Significant group differences in vaginal and labial dimensions were found, with Chinese nullipara dimensions being up to 21% smaller than those of Western nullipara.

Brief Summary

Ethnic Chinese women have smaller vaginal and labial dimensions than Western women. These differences have clinical and surgical implications.
Auto QSAR-based Active learning docking for hit identification of potential inhibitors ofPlasmodium falciparumHsp90 as antimalarial agents
Authors: Matlhodi Thato , Makatsela Lisema Patrick , Dongola Tendamudzimu Harmfree ,   Mthokozisi Blessing Cedric Simelane, Shonhai Addmore , Gumede Njabulo Joyfull , Mokoena Fortunate
Published: May 2024

Malaria which is mainly caused byPlasmodium falciparumparasite remains a devastating public health concern, necessitating the need to develop new antimalarial agents.P. falciparumheat shock protein 90 (Hsp90), is indispensable for parasite survival and a promising drug target. Inhibitors targeting the ATP-binding pocket of the N-terminal domain have anti-Plasmodiumeffects. We proposed ade novoactive learning (AL) driven method in tandem with docking to predict inhibitors with unique scaffolds and preferential selectivity towards PfHsp90. Reference compounds, predicted to bind PfHsp90 at the ATP-binding pocket and possessing anti-Plasmodiumactivities, were used to generate 10,000 unique derivatives and to build the Auto-quantitative structures activity relationships (QSAR) models. Glide docking was performed to predict the docking scores of the derivatives and > 15,000 compounds obtained from the ChEMBL database. Re-iterative training and testing of the models was performed until the optimum Kennel-based Partial Least Square (KPLS) regression model with a regression coefficient R2 = 0.75 for the training set and squared correlation prediction Q2 = 0.62 for the test set reached convergence. Rescoring using induced fit docking and molecular dynamics simulations enabled us to prioritize 15 ATP/ADP-like design ideas for purchase. The compounds exerted moderate activity towardsP. falciparumNF54 strain with IC50values of ≤ 6μM and displayed moderate to weak affinity towards PfHsp90 (KDrange: 13.5-19.9μM) comparable to the reported affinity of ADP. The most potent compound was FTN-T5 (PfN54 IC50:1.44μM; HepG2/CHO cells SI≥ 29) which bound to PfHsp90 with moderate affinity (KD:7.7μM), providing a starting point for optimization efforts. Our work demonstrates the great utility of AL for the rapid identification of novel molecules for drug discovery (i.e., hit identification). The potency of FTN-T5 will be critical for designing species-selective inhibitors towards developing more efficient agents against malaria.
Endoplasmic Reticulum Alterations in Living Cells Expressing INF2 Variants Causing Glomerulosclerosis and Peripheral Neuropathy
Authors: Tran Quynh Thuy Huong , Kondo Naoyuki , Ueda Hiroko , Matsuo Yoshiyuki , Tsukaguchi Hiroyasu
Published: May 2024

The endoplasmic reticulum (ER) is a single dynamic, and continuous network, which mediates a variety of biological processes. The ER spreads throughout the cytoplasm as an interconnected network mainly consisting of flat (sheets) or reticular (tubules) networks. Such ER integrity is regulated by actin-microtubule interaction. INF2 is an actin assembly factor that exclusively located in ER and is mutated in hereditary form of glomerulopathy (focal segmental glomerulosclerosis, FSGS) and peripheral neuropathy (Charcot-Marie Tooth, CMT-DIE, MIM 614455). It remains unclear how INF2 variants could affect ER morphology.

High-resolution, live-imaging of HeLa cells revealed that pathogenic INF2-CAAX variants disrupt peripheral ER complexity, generating focal clustering of polygonal tubules and preferential sheet-like appearance. G73D (causing CMT+FSGS) induced more remarkable alterations than T161N, N202S and R218W (leading to FSGS). Both actin and microtubule inhibitors shifted the ER balance towards sheet predominance and focal compaction of ER tubules, suggesting the role of cytoskeleton in shaping tubular ER networks. INF2 variants induced mitochondria fragmentation with peripheral mis-distribution. The mitochondrial alterations correlated with the degree of cytoskeletal disorganization, leading to defective respiratory function. Moreover, lysosomal trafficking was restricted by INF2 variants in the cell cortex. These organelle-cytoskeletal interactions were more remarkably impaired by CMT+FSGS variant than in FSGS variants.

Our observations underscore that INF2 variants disrupt ER integrity by disorganizing cytoskeletons, which leads to defective mitochondria function and vesicle trafficking in INF2 disorders. INF2 CMT+FSGS variants impair ER-organelles interaction more prominent than FSGS variants, suggesting the existence of specific mediators for CMT+FSGS variants.
CD44/Integrin β1 association drives fast motility on HA substrates
Authors: Roy Tanusri , Dutta Sarbajeet , Sthanam Lakshmi Kavitha , Sen Shamik
Published: May 2024

In addition to proteins such as collagen (Col) and fibronectin, the extracellular matrix (ECM) is enriched with bulky proteoglycan molecules such as hyaluronic acid (HA). However, how ECM proteins and proteoglycans collectively regulate cellular processes has not been adequately explored. Here, we address this question by studying cytoskeletal and focal adhesion organization and dynamics on cells cultured on polyacrylamide hydrogels functionalized with Col, HA and a combination of Col and HA (Col/HA). We show that fastest migration on HA substrates is attributed to the presence of smaller and weaker focal adhesions. Integrinβ1 co-localization and its association with CD44—which is the receptor for HA, and insensitivity of cell spreading to RGD on HA substrates suggests that focal adhesions on HA substrates are formed via integrin association with HA bound CD44. Consistent with this, adhesion formation and cell motility were inhibited when CD44 was knocked out. Collectively, our results suggest that association of integrinβ1 with CD44 drives fast motility on HA substrates.
Elastic tethers remain functional during anaphase arrest in partially-lysed crane-fly spermatocytes: a possible approach for studying mitotic tethers
Authors: Aidil Aisha , Malick Samir A. , Forer Arthur
Published: May 2024

Mitotic tethers connect partner telomeres of all segregating anaphase chromosomes in all animal cells that have been tested, as detected by laser-cutting chromosome arms during anaphase and seeing that the arm fragments move rapidly across the equator to their partner chromosome moving to the opposite pole, telomere moving towards telomere. Tethers exert anti-poleward forces on the poleward separating telomeres, but tether elasticity (that produces the backwards forces) diminishes during anaphase: as determined by the behavior of arm fragments; short tethers (early anaphase) are elastic, long tethers (late anaphase) are not elastic, and medium-length tethers transition between the two states. We developed a procedure in which the tethers still functioned after we partially-lysed anaphase crane-fly spermatocytes. The partial lysis consistently arrested chromosome movements, after which the tethers moved the chromosomes backwards, potentially allowing the elastic tethers to be studied biochemically. To ensure that tether function was not altered by the partial cell-lysis procedure, we compared backward chromosome movements in partially-lysed cells with arm fragment movements in control cells. In the partially-lysed cells the backward chromosomal movements had characteristics identical to those of arm fragments in non-lysed (control) cells. In particular, in both control and partially-lysed cells shorter tethers caused backward movements more often than did longer tethers; shorter tethers caused backward movements over greater fractional distances (of the tether) than did longer tethers; and velocities of the backwards movements were the same for tethers of different lengths. We also compared the effects of Calyculin A (an inhibitor of Protein Phosphatase1) in controlversusin partially-lysed cells. Calyculin A (CalA) added to control cells in early anaphase blocks dephosphorylation, thereby maintaining tether elasticity throughout anaphase: after the chromosomes reach the poles they move backwards when the usual poleward forces are reduced. Partial lysis preserves this tether functionality: after partial lysis of CalA-treated cells the chromosomes move backward and reach the partner telomeres at even very long tether lengths. We conclude that partial cell-lysis arrests anaphase chromosome poleward movement but does not affect tether function.
Rational design of HER2-targeted combination therapies to reverse drug resistance in fibroblast-protected HER2+ breast cancer cells
Authors: Poskus Matthew D. , McDonald Jacob , Laird Matthew , Li Ruxuan , Norcoss Kyle , Zervantonakis Ioannis K.
Published: May 2024

Introduction

Fibroblasts, an abundant cell type in the breast tumor microenvironment, interact with cancer cells and orchestrate tumor progression and drug resistance. However, the mechanisms by which fibroblast-derived factors impact drug sensitivity remain poorly understood. Here, we develop rational combination therapies that are informed by proteomic profiling to overcome fibroblast-mediated therapeutic resistance in HER2+ breast cancer cells.

Methods

Drug sensitivity to the HER2 kinase inhibitor lapatinib was characterized under conditions of monoculture and exposure to breast fibroblast-conditioned medium. Protein expression was measured using reverse phase protein arrays. Candidate targets for combination therapy were identified using differential expression and multivariate regression modeling. Follow-up experiments were performed to evaluate the effects of HER2 kinase combination therapies in fibroblast-protected cancer cell lines and fibroblasts.

Results

Compared to monoculture, fibroblast-conditioned medium increased the expression of plasminogen activator inhibitor-1 (PAI1) and cell cycle regulator polo like kinase 1 (PLK1) in lapatinib-treated breast cancer cells. Combination therapy of lapatinib with inhibitors targeting either PAI1 or PLK1, eliminated fibroblast-protected cancer cells, under both conditions of direct coculture with fibroblasts and protection by fibroblast-conditioned medium. Analysis of publicly available, clinical transcriptomic datasets revealed that HER2-targeted therapy fails to suppress PLK1 expression in stroma-rich HER2+ breast tumors and that high PAI1 gene expression associates with high stroma density. Furthermore, we showed that an epigenetics-directed approach using a bromodomain and extraterminal inhibitor to globally target fibroblast-induced proteomic adaptions in cancer cells, also restored lapatinib sensitivity.

Conclusions

Our data-driven framework of proteomic profiling in breast cancer cells identified the proteolytic degradation regulator PAI1 and the cell cycle regulator PLK1 as predictors of fibroblast-mediated treatment resistance. Combination therapies targeting HER2 kinase and these fibroblast-induced signaling adaptations eliminates fibroblast-protected HER2+ breast cancer cells.
Elastic mitotic tethers remain functional after microtubules are inhibited
Authors: Adil Aisha , Janan Maral , Forer Arthur
Published: May 2024

During normal anaphase in animal cells, elastic tethers connect partner telomeres of segregating chromosomes and exert backward (anti-poleward) forces on those chromosomes. The experiments reported herein test whether microtubules need to be present in order for tethers to produce backwards forces. We disassembled spindle microtubules by treating anaphase crane-fly primary spermatocytes separately with nocodazole, colcemid, or podophyllotoxin. The drug treatments caused anaphase chromosomes to stop moving poleward; almost immediately thereafter they moved backward. The characteristics of the backward movements of the half-bivalents match those of the backwards movements of arm fragments formed by cutting chromosome arms during anaphase – for example the occurrence and lengths of backward movements were a function of tether length. The only difference from movement of arm fragments is that the chromosomes in the treated cells moved backwards slower than arm fragments did. Immunofluorescence of spindle tubulin after the drug treatments indicated that acetylated kinetochore microtubules were not depolymerized by the drugs, though the non-kinetochore spindle microtubules were depolymerized. Our data indicate that tethers move anaphase chromosomes backwards in the absence of functioning spindle microtubules. We suggest that the backward movements that take place when poleward forces are absent are due to tethers, and that the backward movements are slowed by the presence of acetylated kinetochore microtubules.
Brain Extracellular Matrix implications in multiple neurological disorders are revealed through a meta-analysis of transcriptional changes
Authors: Sadis Hagit , Peles David , Hussein Yara , Stern Shani
Published: May 2024

Neurological disorders comprise a wide range of illnesses that may affect the central and peripheral nervous systems. Despite diverse etiologies, patients with these disorders may share symptoms.

In this study, we aimed to explore potential common mechanisms between seven neurological disorders spanning three categories: neurodegenerative diseases, neuropsychiatric disorders, and neurodevelopmental disorders, by comparing gene expression profiles and focusing on the most prominent dysregulated genes consistently reported within and across disorders. Our results demonstrate 31 genes that are commonly differentially expressed in brain cells and tissues derived from human disease models when compared to healthy controls. These genes were enriched in brain Extracellular Matrix (ECM) pathways, Growth factor binding, Response to acid chemical, and External encapsulating structure. Remarkedly, dysregulation of ECM genes was evident separately in each of the three categories of disorders. This suggests a notable distinction in the brain ECM in disease states. Furthermore, we identified that the most frequently reported genes among all disorders wereGFAP, andIFITM3.

Key Points

Analysis of 41 human studies revealed 31 significantly dysregulated genes shared among seven neurological disorders when compared to healthy controls, spanning three distinct categories: Neurodegenerative diseases, Neuropsychiatric disorders, and Neurodevelopmental disorders.

These shared Differentially Expressed Genes (DEGs) demonstrated significant enrichment for Extracellular Matrix (ECM) pathways, Growth factor binding, Response to acid chemical, Blood vessel development, and External encapsulating structure. Particularly,SSTandBCL6were the most frequently reported shared DEGs.

Notably, each of the three categories of neurological disorders exhibited significant cellular component enrichment for ECM pathways.

In order to distinguish noise genes (false-positive genes) from disease-relevant genes, we identified the DEGs that were reported the highest number of times per disorder.GFAP, followed byIFITM3, were found to be the most reported genes.

Furthermore, due to partially shared symptoms, we explored commonalities between Autism Spectrum Disorders (ASD) and Schizophrenia. DEGs shared between both disorders were specifically enriched with ECM pathways, External encapsulating structure, Growth factor binding, Cell adhesion molecule binding, and PI3K-Akt signaling pathway. Noteworthy,IFITM2, HSPB1, IFITM3, HSPA1A, MKNK2, GFAPandCOL4A1were among the most frequently reported shared DEGs.

The central aspects of our findings suggest a substantial distinction between the Central Nervous System (CNS) ECM in health and disease.
The PVD neuron has male-specific structure and mating function inC. elegans
Authors: Iosilevskii Yael , Hall David H. , Katz Menachem , Podbilewicz Benjamin
Published: May 2024

Neurons display unique shapes and establish intricate networks, which may differ between sexes. In complex organisms, studying sex differences in structure and function of individual neurons is difficult. The nematodeCaenorhabditis eleganshermaphrodites and males present an exceptional model for studying neuronal morphogenesis in a simple, sexually-dimorphic system. We focus on the polymodal sensory bilateral neuron pair PVD, which forms a complex but stereotypic dendritic tree composed of multiple subunits that resemble candelabra. PVD is well studied in hermaphrodites, but not in males. We show here that during larval development, male PVDs extend a similar architecture to the hermaphrodite utilizing the sexually-shared Menorin patterning mechanism. In early adulthood, however, male PVD develops a unique extension into the copulatory tail structure. Alongside established tail ray neurons RnA and RnB, we show PVD is a third, previously unrecognized, neuron within the tail rays. Unlike RnA and RnB, PVD extends anterogradely, branches and turns within the ray hypodermis, and is non-ciliated. This PVD sexually-dimorphic arborization is absent in mutant backgrounds which perturb the Menorin guidance complex. SAX-7/L1CAM, a hypodermal component of this complex, shows a male-specific expression pattern which precedes PVD extension, and its presence allows PVD to enter the tail rays. Further, our results reveal that genetically altered arborization or ablation of the PVD result in male mating behavioral defects, particularly as males turn around the hermaphrodite. These results uncover an adult-stage sexual dimorphism of dendritic branching and uncover a function for PVD in male sexual behavior.

Significance Statement

Neurons form intricate shapes and networks, which may display sexual differences. Pinpointing these changes at the single cell level, however, is challenging. InC. elegans, the PVD neuron is a powerful model for stereotypical neuron shaping, yet most studies concern only one of two sexes. Our research focuses on the understudied male, where we show an adult-stage extension of PVD into the male copulatory tail organ. We further show how this stems from a sex-shared patterning complex, progressing independently and in a different environment compared with nearby male-specific neurons. We further find PVD has a role in the male adult-specific mating behavior. PVD thus presents a unique example of a highly arborized neuron showing sexually dimorphic behavioral function and structure.
Intra-amygdala circuits of sleep disruption-induced anxiety in female mice
Authors: Peng Beibei , Foilb Allison R. , Manasian Yunona , Li Yan , Deng Xin , Meloni Edward G. , Ressler Kerry J. , Carlezon William A. , Bolshakov Vadim Y.
Published: May 2024

Combining mouse genetics, electrophysiology, and behavioral training and testing, we explored how sleep disruption may affect the function of anxiety-controlling circuits, focusing on projections from the basolateral nucleus of the amygdala (BLA) to CRF-positive cells in the lateral division of the central amygdala (CeL). We found in Crh-IRES-Cre::Ai14(tdTomato) reporter female mice that 6 hours of sleep disruption during their non-active (light) cycle may be anxiogenic. Notably, the AMPAR/NMDAR EPSC amplitude ratio at the BLA inputs to CRF-CeL cells (CRFCeL), assessed with whole-cell recordings inex vivoexperiments, was enhanced in slices from sleep-disrupted mice, whereas paired-pulse ratio (PPR) of the EPSCs induced by two closely spaced presynaptic stimuli remained unchanged. These findings indicate that sleep disruption-associated synaptic enhancements in glutamatergic projections from the BLA to CRF-CeL neurons may be postsynaptically expressed. We found also that the excitation/inhibition (E/I) ratio in the BLA to CRFCeLinputs was increased in sleep-disrupted mice, suggesting that the functional efficiency of excitation in BLA inputs to CRFCeLcells has increased following sleep disruption, thus resulting in their enhanced activation. The latter could be translated into enhanced anxiogenesis as activation of CRF cells in the CeL was shown to promote anxiety-like behaviors.
Differential Control of Inhibitory and Excitatory Nerve Terminal Function by Mitochondria
Authors: Bredvik Kirsten , Ryan Timothy A.
Published: May 2024

Inhibitory neurons shape the brain’s computational landscape and rely on different cellular architectures and intrinsic properties than excitatory neurons. Maintenance of the overall balance of excitatory (E) versus inhibitory (I) drive is essential, as disruptions can lead to neuropathological conditions, including autism and epilepsy. Metabolic perturbations are a common driver of E/I imbalance but differential sensitivity of these two neuron types to metabolic lesions is not well understood. Here, we characterized differences in presynaptic bioenergetic regulation between excitatory and inhibitory nerve terminals using genetically encoded indicators expressed in primary dissociated neuronal cultures. Our experiments showed that inhibitory nerve terminals sustain higher ATP levels than excitatory nerve terminals arising from increased mitochondrial metabolism. Additionally, mitochondria in inhibitory neurons play a greater role in buffering presynaptic Ca2+and inhibitory mitochondrial Ca2+handling is differentially regulated by TMEM65-mediated acceleration of mitochondrial Ca2+extrusion following bursts of activity. These experiments thus identify differential reliance on mitochondrial function across two major neuron types.
Sustainable admixtures to enhance long-term strength and durability properties of eco-concrete: an innovative use of Saudi agro-industrial by-products
Authors: Amran Mugahed , Onaizi Ali
Published: May 2024

Purpose

Low-carbon concrete represents a new direction in mitigating the global warming effects caused by clinker manufacturing. Utilizing Saudi agro-industrial by-products as an alternative to cement is a key support in reducing clinker production and promoting innovation in infrastructure and circular economy concepts, toward decarbonization in the construction industry. The use of fly ash (FA) as a cement alternative has been researched and proven effective in enhancing the durability of FA-based concrete, especially at lower replacement levels. However, at higher replacement levels, a noticeable impediment in mechanical strength indicators limits the use of this material.

Design/methodology/approach

In this study, low-carbon concrete mixes were designed by replacing 50% of the cement with FA. Varying ratios of nano-sized glass powder (4 and 6% of cement weight) were used as nanomaterial additives to enhance the mechanical properties and durability of the designed concrete. In addition, a 10% of the mixing water was replaced with EMs dosage.

Findings

The results obtained showed a significant positive impact on resistance and durability properties when replacing 10% of the mixing water with effective microorganisms (EMs) broth and incorporating nanomaterial additives. The optimal mix ratios were those designed with 10% EMs and 4–6% nano-sized glass powder additives. However, it can be concluded that advancements in eco-friendly concrete additive technologies have made significant contributions to the development of sophisticated concrete varieties.

Originality/value

This study focused at developing nanomaterial additives from Saudi industrial wastes and at presenting a cost-effective and feasible solution for enhancing the properties of FA-based concrete. It has also been found that the inclusion of EMs contributes effectively to enhancing the concrete's resistance properties.