bluuebunny/crossref_metadata_2025_split huggingface dataset:
Rat primary cortical cell tri-culture to study effects of amyloid-beta on microglia function
Authors: Kim Hyehyun , Le Bryan , Goshi Noah , Zhu Kan , Grodzki Ana Cristina , Lein Pamela J. , Zhao Min , Seker Erkin 
Published: March 2024

AbstractIntroductionThe etiology and progression of sporadic Alzheimer’s Disease (AD) have been studied for decades. One proposed mechanism is that amyloid-beta (Aβ) proteins induce neuroinflammation, synapse loss, and neuronal cell death. Microglia play an especially important role in Aβ clearance, and alterations in microglial function due to aging or disease may result in Aβ accumulation and deleterious effects on neuronal function. However, studying these complex factorsin vivo, where numerous confounding processes exist, is challenging, and until recently,in vitromodels have not allowed sustained culture of microglia, astrocytes and neurons in the same culture. Here, we employ a tri-culture model of rat primary neurons, astrocytes, and microglia and compare it to co-culture (neurons and astrocytes) and mono-culture enriched for microglia to study microglial function (i.e., motility and Aβ clearance) and proteomic response to exogenous Aβ.MethodsWe established cortical co-culture (neurons and astrocytes), tri-culture (neurons, astrocytes, and microglia), and mono-culture (microglia) from perinatal rat pups. On daysin vitro(DIV) 7 – 14, the cultures were exposed to fluorescently-labeled Aβ (FITC-Aβ) particles for varying durations. Images were analyzed to determine the number of FITC-Aβ particles after specific lengths of exposure. A group of cells were stained for βIII-tubulin, GFAP, and Iba1 for morphological analysis via quantitative fluorescence microscopy. Cytokine profiles from conditioned media were obtained. Live-cell imaging with images acquired every 5 minutes for 4 hours was employed to extract microglia motility parameters (e.g., Euclidean distance, migration speed, directionality ratio).Results and discussionFITC-Aβ particles were more effectively cleared in the tri-culture compared to the co-culture. This was attributed to microglia engulfing FITC-Aβ particles, as confirmed via epifluorescence and confocal microscopy. Adding FITC-Aβ significantly increased the size of microglia, but had no significant effect on neuronal surface coverage or astrocyte size. Analysis of the cytokine profile upon FITC-Aβ addition revealed a significant increase in proinflammatory cytokines (TNF-α, IL-1α, IL-1β, IL-6) in tri-culture, but not co-culture. In addition, Aβ addition altered microglia motility marked by swarming-like motion with decreased Euclidean distance yet unaltered speed. These results highlight the importance of cell-cell communication in microglia function (e.g., motility and Aβ clearance) and the utility of the tri-culture model to further investigate microglia dysfunction in AD.
Contradictory behavioral effects of neuronal perturbations on behavioral responses to linearly polarized light in freely walkingDrosophila
Authors: Titova Anna V. , Straw Andrew D. 
Published: March 2024

AbstractMany insects can use the polarization of the skylight as a navigational cue. As shown previously, freely walkingDrosophilaorient along the e-vector of linearly polarized UV light presented both dorsally and ventrally. We are interested in the neuronal mechanisms leading to this behavior, and specifically how the central complex and its inputs are involved. We investigated the behavior of flies exposed to linearly polarized near-UV light (400 nm) presented dorsally. Flies walked freely in a circular, flat arena surrounded by a heat barrier. Using the GAL4-UAS genetic system, we drove the expression of the potassium inward rectifier KIR2.1 to perturb each of several different neuron types of the polarization vision pathway. Perturbing EPG compass neurons in the central complex slightly weakened average alignment and increased its variability. On the other hand, when two different GAL4 lines driving expression in the ER4m ring neurons, identified by connectomics as the major polarization inputs to the fly central complex, were perturbed, the alignment strength increased. A similar effect was observed when the inputs to ER4m, the TuBuaneurons, were perturbed. We did not predict EPG and ER4m perturbations to cause opposite effects. Further investigation would be required to understand the physiological mechanisms of these contradictory behavioral effects.
Structural basis for allosteric regulation of human phosphofructokinase-1
Authors: Lynch Eric M. , Hansen Heather , Salay Lauren , Cooper Madison , Timr Stepan , Kollman Justin M. , Webb Bradley A. 
Published: March 2024

AbstractPhosphofructokinase-1 (PFK1) catalyzes the rate-limiting step of glycolysis, committing glucose to conversion into cellular energy. PFK1 is highly regulated to respond to the changing energy needs of the cell. In bacteria, the structural basis of PFK1 regulation is a textbook example of allostery; molecular signals of low and high cellular energy promote transition between an active R-state and inactive T-state conformation, respectively. Little is known, however, about the structural basis for regulation of eukaryotic PFK1. Here, we determine structures of the human liver isoform of PFK1 (PFKL) in the R- and T-state by cryoEM, providing insight into eukaryotic PFK1 allosteric regulatory mechanisms. The T-state structure reveals conformational differences between the bacterial and eukaryotic enzyme, the mechanisms of allosteric inhibition by ATP binding at multiple sites, and an autoinhibitory role of the C-terminus in stabilizing the T-state. We also determine structures of PFKL filaments that define the mechanism of higher-order assembly and demonstrate that these structures are necessary for higher-order assembly of PFKL in cells.
Bottlenecks in the Implementation of Genome Scale Metabolic Model Based Designs for Bioproduction from Aromatic Carbon Sources
Authors: Banerjee Deepanwita , Menasalvas Javier , Chen Yan , Gin Jennifer W. , Baidoo Edward E. K. , Petzold Christopher J. , Eng Thomas , Mukhopadhyay Aindrila 
Published: March 2024

ABSTRACTGenome scale metabolic models (GSMM) are commonly used to identify gene deletion sets that result in growth coupling, pairing product formation with substrate utilization. While such approaches can improve strain performance beyond levels typically accessible using targeted strain engineering approaches, sustainable feedstocks often pose a challenge for GSMM-based methods due to incomplete underlying metabolic data. Specifically, we address a four-gene deletion design for the lignin-derived non-sugar carbon source,para-coumarate, that proved challenging to implement. We examine the performance of the fully implemented design forp-coumarate to glutamine, a useful biomanufacturing intermediate. In this study glutamine is then converted to indigoidine, an alternative sustainable pigment and a model heterologous product. Through omics, promoter-variation and growth characterization of a fully implemented gene deletion design, we provide evidence that aromatic catabolism in the completed design is rate-limited by fumarate hydratase activity in the citrate cycle and required careful optimization of the final fumarate hydratase protein (PP_0897) expression to achieve growth and production. A metabolic cross-feeding experiment with the completed design strain also revealed an unanticipated nutrient requirement suggesting additional functions for the fumarate hydratase protein. A double sensitivity analysis confirmed a strict requirement for fumarate hydratase activity in the strain where all genes in the growth coupling design have been implemented. While a complete implementation of the design was achieved, this study highlights the challenge of precisely inactivating metabolic reactions encoded by under-characterized proteins especially in the context of multi-gene edits.
Redefining the role of Hypoxia-inducible factors (HIFs) in oxygen homeostasis
Authors: Arias Clemente F. , Acosta Francisco J. , Bertocchini Federica , Fernańdez-Arias Cristina 
Published: March 2024

ABSTRACTHypoxia-inducible factors (HIFs) are key regulators of intracellular oxygen homeostasis. The marked increase in HIFs activity in hypoxia as compared to normoxia, together with their transcriptional control of primary metabolic pathways, motivated the widespread view of HIFs as responsible for the cell’s metabolic adaptation to hypoxic stress. In this work, we suggest that this prevailing model of HIFs regulation is misleading. We propose an alternative model focused on understanding the dynamics of HIFs’ activity within its physiological context. Our model suggests that HIFs would not respond to but rather prevent the onset of hypoxic stress by regulating the traffic of electrons between catabolic substrates and oxygen. The explanatory power of our approach is patent in its interpretation of the Warburg effect, the tendency of tumor cells to favor anaerobic metabolism over respiration, even in fully aerobic conditions. This puzzling behavior is currently considered as an anomalous metabolic deviation. Our model predicts the Warburg effect as the expected homeostatic response of tumor cells to the abnormal increase in metabolic demand that characterizes malignant phenotypes. This alternative perspective prompts a redefinition of HIFs’ function and underscores the need to explicitly consider the cell’s metabolic activity in understanding its responses to changes in oxygen availability.
Mapping the breast tumor microenvironment: proximity analysis reveals spatial relationships between macrophage subtypes and metastasis-initiating cancer cells
Authors: Grasset Eloise M. , Desphande Atul , Lee Jae W. , Cho Yeonju , Shin Sarah M. , Coyne Erin M. , Hernandez Alexei , Yuan Xuan , Zhang Zhehao , Cimino-Matthews Ashley , Ewald Andrew J. , Ho Won Jin 
Published: March 2024

AbstractThe development of metastasis, responsible for the majority of cancer-related fatalities, is the most dangerous aspect of breast cancer, the predominant malignancy affecting women. We previously identified specific cancer cell populations responsible for metastatic events which are cytokeratin-14 (CK14) and E-cadherin positive in luminal tumors, and E-cadherin and vimentin positive in triple-negative tumors. Since cancer cells evolve within a complex ecosystem comprised of immune cells and stromal cells, we sought to decipher the spatial interactions of these aggressive cancer cell populations within the tumor microenvironment (TME). We used imaging mass cytometry to detect 36 proteins in tumor microarrays containing paired primary and metastatic lesions from luminal or triple-negative breast cancers (TNBC), resulting in a dataset of 1,477,337 annotated cells. Focusing on metastasis-initiating cell populations, we observed close proximity to specific fibroblast and macrophage subtypes, a relationship maintained between primary and metastatic tumors. Notably, high CK14 in luminal cancer cells and high vimentin in TNBC cells correlated with close proximity to specific macrophage subtypes (CD163intCD206intPDL1intHLA-DR+or PDL1highARG1high). Our in-depth spatial analysis elucidates that metastasis-initiating cancer cells exhibit with distinct cell populations within the TME, implicating the role of these cell-cell interactions in promoting metastasis.
Myocardin-related transcription factor drives epithelial fibrogenesis in polycystic kidney disease
Authors: Lichner Zsuzsanna , Ding Mei , Khare Tarang , Dan Qinghong , Benitez Raquel , Praszner Mercédesz , Song Xuewen , Saleeb Rola , Hinz Boris , Pei York , Szászi Katalin , Kapus András 
Published: March 2024

AbstractPolycystic kidney disease (PKD) is characterized by extensive cyst formation and progressive fibrosis. However, the molecular mechanisms whereby the loss/loss-of-function of Polycystin 1 or 2 (PC1/2) provokes fibrosis are largely unknown. The small GTPase RhoA has been recently implicated incystogenesis, and we have shown that the RhoA/cytoskeleton/myocardin-related transcription factor (MRTF) pathway is a key mediator of epithelium-induced fibrogenesis. Therefore, we hypothesized that MRTF is activated by PC1/2 loss and plays a critical role in fibrogenic reprogramming of the epithelium. Loss of PC1 or PC2 induced by siRNA in vitro activated RhoA, caused cytoskeletal remodeling and robust nuclear MRTF translocation and overexpression. These phenomena were also manifest in PKD1 (RC/RC) and PKD2 (WS25/-) mice, with MRTF translocation and overexpression occurring predominantly in dilated tubules and in cyst-lining epithelium, respectively. In epithelial cells, a large cohort of PC1/PC2 downregulation-induced genes was MRTF-dependent, including cytoskeletal, integrin-related, and matricellular/fibrogenic proteins. Epithelial MRTF was necessary for paracrine priming of fibroblast-myofibroblast transition. Thus, MRTF is a critical novel mediator of the PC1/2 loss-induced profibrotic epithelial phenotype, and consequently PKD-related fibrosis.
Transposable element methylation state predicts age and disease
Authors: Morandini Francesco , Lu Jinlong Y. , Rechsteiner Cheyenne , Shadyab Aladdin H. , Casanova Ramon , Snively Beverly M. , Seluanov Andrei , Gorbunova Vera 
Published: March 2024

AbstractTransposable elements (TEs) are DNA sequences that expand selfishly in the genome, possibly causing severe cellular damage. While normally silenced, TEs have been shown to activate during aging. DNA methylation is one of the main mechanisms by which TEs are silenced and has been used to train highly accurate age predictors. Yet, one common criticism of such predictors is that they lack interpretability. In this study, we investigate the changes in TE methylation that occur during human aging. We find that evolutionarily young LINE1s (L1s), the only known TEs capable of autonomous transposition in humans, undergo the fastest loss of methylation, suggesting an active mechanism of de-repression. We then show that accurate age predictors can be trained on both methylation of individual TE copies and average methylation of TE families genome wide. Lastly, we show that while old L1s gradually lose methylation during the entire lifespan, demethylation of young L1s only happens late in life and is associated with cancer.
Curating models from BioModels: Developing a workflow for creating OMEX files
Authors: Xu Jin , Smith Lucian 
Published: March 2024

AbstractThe reproducibility of computational biology models can be greatly facilitated by widely adopted standards and public repositories. We examined 50 models from the BioModels Database and attempted to validate the original curation and correct some of them if necessary. For each model, we reproduced these published results using Tellurium. Once reproduced we manually created a new set of files, with the model information stored by the Systems Biology Markup Language (SBML), and simulation instructions stored by the Simulation Experiment Description Markup Language (SED-ML), and everything included in an Open Modeling EXchange (OMEX) file, which could be used with a variety of simulators to reproduce the same results. On the one hand, the reproducibility procedure of 50 models developed a manual workflow that we would use to build an automatic platform to help users more easily curate and verify models in the future. On the other hand, these exercises allowed us to find the limitations and possible enhancement of the current curation and tooling to verify and curate models.
A feedback-driven brain organoid platform enables automated maintenance and high-resolution neural activity monitoring
Authors: Voitiuk Kateryna , Seiler Spencer T. , Pessoa de Melo Mirella , Geng Jinghui , van der Molen Tjitse , Hernandez Sebastian , Schweiger Hunter E. , Sevetson Jess L. , Parks David F. , Robbins Ash , Torres-Montoya Sebastian , Ehrlich Drew , Elliott Matthew A. T. , Sharf Tal , Haussler David , Mostajo-Radji Mohammed A. , Salama Sofie R. , Teodorescu Mircea 
Published: March 2024

AbstractThe analysis of tissue cultures, particularly brain organoids, requires a sophisticated integration and coordination of multiple technologies for monitoring and measuring. We have developed an automated research platform enabling independent devices to achieve collaborative objectives for feedback-driven cell culture studies. Our approach enables continuous, communicative, non-invasive interactions within an Internet of Things (IoT) architecture among various sensing and actuation devices, achieving precisely timed control ofin vitrobiological experiments. The framework integrates microfluidics, electrophysiology, and imaging devices to maintain cerebral cortex organoids while measuring their neuronal activity. The organoids are cultured in custom, 3D-printed chambers affixed to commercial microelectrode arrays. Periodic feeding is achieved using programmable microfluidic pumps. We developed a computer vision fluid volume estimator used as feedback to rectify deviations in microfluidic perfusion during media feeding/aspiration cycles. We validated the system with a set of 7-day studies of mouse cerebral cortex organoids, comparing manual and automated protocols. The automated protocols were validated in maintaining robust neural activity throughout the experiment. The automated system enabled hourly electrophysiology recordings for the 7-day studies. Median neural unit firing rates increased for every sample and dynamic patterns of organoid firing rates were revealed by high-frequency recordings. Surprisingly, feeding did not affect firing rate. Furthermore, performing media exchange during a recording showed no acute effects on firing rate, enabling the use of this automated platform for reagent screening studies.
Human cortical neurons rapidly generated by direct ES cell programming integrate into stroke-injured rat cortex
Authors: Martinez-Curiel Raquel , Hayj Mazin , Tsupykov Oleg , Jansson Linda , Avaliani Natalia , Martín Berta Coll-San , Monni Emanuela , Skibo Galyna , Lindvall Olle , Palma-Tortosa Sara , Kokaia Zaal 
Published: March 2024

ABSTRACTStroke is a major cause of long-term disability in adult humans, the neuronal loss leading to motor, sensory, and cognitive impairments. Replacement of dead neurons by intracerebral transplantation of stem cell-derived neurons for reconstruction of injured neuronal networks has potential to become a novel therapeutic strategy to promote functional recovery after stroke. Here we describe a rapid and efficient protocol for the generation of cortical neurons via direct programming of human embryonic stem (hES) cells. Our results show that 7 days overexpression of the transcription factor neurogenin 2 (NGN2) in vitro was enough to generate hES-induced cells with cortical phenotype, as revealed by immunocytochemistry and RT-qPCR, and electrophysiological properties of neurons in an intermediate stage of maturity. At 3 months after translantation into the stroke-injured rat cortex, the hES-induced neurons (hES-iNs) showed immunocytochemical markers of mature layer-specific cortical neurons and sent widespread axonal projections to several areas in both hemispheres of the host brain. Their axons became myelinated and formed synaptic contacts with host neurons, as shown by immunoelectron microscopy. Our findings demonstrate for the first time that direct transcription factor programming of hES cells can efficiently and rapidly produce cortical neurons with capacity to integrate into the stroke-injured brain.
Random forest machine-learning algorithm classifies white- and brown-rot fungi according to the number of Carbohydrate-Active enZyme genes
Authors: Hasegawa Natsuki , Sugiyama Masashi , Igarashi Kiyohiko 
Published: March 2024

AbstractWood-rotting fungi play an important role in the global carbon cycle because they are only known organisms that digest wood, the largest carbon stock in nature. In the present study, we used linear discriminant analysis and random forest (RF) machine learning algorithms to predict white- or brown-rot decay modes from the numbers of genes encoding Carbohydrate-Active enZymes (CAZymes) with over 98% accuracy. Unlike other algorithms, RF identified specific genes involved in cellulose and lignin degradation, including auxiliary activities (AA) family 9 lytic polysaccharide monooxygenases, glycoside hydrolase family 7 cellobiohydrolases, and AA family 2 peroxidases, as critical factors. This study sheds light on the complex interplay between genetic information and decay modes and underscores the potential of RF for comparative genomics studies of wood-rotting fungi.ImportanceWood-rotting fungi are categorized as either white- or brown-rot modes based on the coloration of decomposed wood. The process of classification can be influenced by human biases. The random forest machine learning algorithm effectively distinguishes between white- and brown-rot fungi based on the presence of Carbohydrate-Active enZyme genes. These findings not only aid in the classification of wood-rotting fungi but also facilitate the identification of the enzymes responsible for degrading woody biomass.
Anti-Immune Complex Antibodies are Elicited During Repeated Immunization with HIV Env Immunogens
Authors: Brown Sharidan , Antanasijevic Aleksandar , Sewall Leigh M. , Garcia Daniel Montiel , Brouwer Philip J. M. , Sanders Rogier W. , Ward Andrew B. 
Published: March 2024

AbstractVaccination strategies against HIV-1 aim to elicit broadly neutralizing antibodies (bnAbs) using prime-boost regimens with HIV envelope (Env) immunogens. Early antibody responses to easily accessible epitopes on these antigens are directed to non-neutralizing epitopes instead of bnAb epitopes. Autologous neutralizing antibody responses appear upon boosting once immunodominant epitopes are saturated. Here we report another type of antibody response that arises after repeated immunizations with HIV Env immunogens and present the structures of six anti-immune complexes discovered using polyclonal epitope mapping. The anti-immune complex antibodies target idiotopes composed of framework regions of antibodies bound to Env. This work sheds light on current vaccine development efforts for HIV, as well as for other pathogens, in which repeated exposure to antigen is required.One Sentence SummaryPolyclonal epitope mapping reveals anti-immune complex antibodies which target idiotopes on antibodies bound to HIV Env.