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| Open AccessFluorescent fatty acid conjugates for live cell imaging of peroxisomes
The array of tools to image peroxisome regulation is still limited. Here, the authors develop improved fatty acid-based probes with high peroxisome specificity and bright fluorescence in the red/far-red spectrum, which makes them ideal to study peroxisomes in live cells and whole organisms.
- Daria Korotkova
- , Anya Borisyuk
- & Triana Amen
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Article
| Open AccessThree stepwise pH progressions in stratum corneum for homeostatic maintenance of the skin
Previous studies of the skin have suggested a neutral-to-acidic pH gradient in the outermost layer. Here, the authors perform quantitative intravital pH imaging of the stratum corneum, the outermost epidermal layer, and demonstrate three distinct pH zones - rather than a gradient - with different biological functions.
- Keitaro Fukuda
- , Yoshihiro Ito
- & Masayuki Amagai
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Article
| Open AccessImplication of thermal signaling in neuronal differentiation revealed by manipulation and measurement of intracellular temperature
The role of subcellular thermogenesis during neuronal differentiation remains poorly understood. Here, the authors employ methods to monitor local intracellular temperature, and they investigate the effects of non-invasive temperature changes on cell differentiation using neuron-like cells.
- Shunsuke Chuma
- , Kazuyuki Kiyosue
- & Yoshie Harada
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Article
| Open AccessPrecise immunofluorescence canceling for highly multiplexed imaging to capture specific cell states
Multiplexed imaging to study cellular pathways can be hampered by lack of antibody specificity, especially when assessing signal activation. Here, the authors present Precise Emission Canceling Antibodies (PECAbs), which enable high-specificity sequential imaging and the study of signaling pathways.
- Kosuke Tomimatsu
- , Takeru Fujii
- & Yasuyuki Ohkawa
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Article
| Open AccessA fast-acting lipid checkpoint in G1 prevents mitotic defects
Lipid synthesis increases during the cell cycle to ensure sufficient membrane mass. Here, authors identify a lipid checkpoint in G1 phase that prevents cells from starting the cell cycle if lipid synthesis is low, thereby preventing mitotic defects.
- Marielle S. Köberlin
- , Yilin Fan
- & Tobias Meyer
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Article
| Open AccessImportin 13-dependent axon diameter growth regulates conduction speeds along myelinated CNS axons
Myelinated axons vary in diameter by over 100-fold. Here, the authors identify a role for the nuclear transport receptor importin 13 in axon diameter growth and corresponding increases to conduction speed along myelinated axons.
- Jenea M. Bin
- , Daumante Suminaite
- & David A. Lyons
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Article
| Open AccessALLO-1- and IKKE-1-dependent positive feedback mechanism promotes the initiation of paternal mitochondrial autophagy
Allophagy eliminates paternal mitochondria in C. elegans embryos. This study reveals that an autophagy adaptor, ALLO-1, recognizes cargos and initiates autophagosome formation by gathering with the ULK complex in an IKKE-1 kinase-dependent manner.
- Taeko Sasaki
- , Yasuharu Kushida
- & Miyuki Sato
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| Open AccessIntraocular liver spheroids for non-invasive high-resolution in vivo monitoring of liver cell function
Longitudinal monitoring of liver function in vivo is hindered by the lack of high-resolution non-invasive imaging techniques. Here, the authors show a crucial and unique tool for longitudinal in vivo imaging of liver spheroids at cellular resolution to study liver physiology and disease.
- Francesca Lazzeri-Barcelo
- , Nuria Oliva-Vilarnau
- & Per-Olof Berggren
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| Open AccessSingle-molecule localization microscopy reveals STING clustering at the trans-Golgi network through palmitoylation-dependent accumulation of cholesterol
Stimulator of interferon genes (STING) is known to translocate from ER to Golgi while orchestrating type I interferon responses to DNA. Here, the authors show single-molecule resolution images of STING clustering at the trans-Golgi network, where it acts as a scaffold to trigger downstream signaling.
- Haruka Kemmoku
- , Kanoko Takahashi
- & Tomohiko Taguchi
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Article
| Open AccessAI-driven projection tomography with multicore fibre-optic cell rotation
Conventional optical tomography can have disadvantages, including anisotropic resolution and incomplete imaging of cellular structures. Here, the authors propose an AI-driven 3D cell imaging system with a cell rotator, which offers improved resolution and automated processing.
- Jiawei Sun
- , Bin Yang
- & Juergen W. Czarske
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Article
| Open AccessSingle-photon microscopy to study biomolecular condensates
The wide variety of cellular processes involving biomolecular condensation makes their quantification a challenging task. Here, the authors present an integrated platform based on single-photon microscopy to study complex biomolecular processes.
- Eleonora Perego
- , Sabrina Zappone
- & Giuseppe Vicidomini
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Article
| Open AccessiU-ExM: nanoscopy of organelles and tissues with iterative ultrastructure expansion microscopy
Current expansion microscopy approaches need further optimization to achieve the precision of nanoscopy techniques. Here, the authors develop an iterative ultrastructure expansion microscopy (iU-ExM) approach that achieves SMLM-level resolution.
- Vincent Louvel
- , Romuald Haase
- & Paul Guichard
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Article
| Open AccessAdaptive traits of cysts of the snow alga Sanguina nivaloides unveiled by 3D subcellular imaging
Sanguina nivaloides thrives in red snowfields worldwide. It cannot be cultivated. Using environmental samples, 3D electron microscopy combined with functional analyzes revealed unique cell architecture features adapted to life in a snowy environment.
- Jade A. Ezzedine
- , Clarisse Uwizeye
- & Eric Maréchal
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Article
| Open AccessLong-term cargo tracking reveals intricate trafficking through active cytoskeletal networks in the crowded cellular environment
Leveraging a label-free interferometric scattering microscope, scientists tracked numerous cargos within a crowded cellular environment. Intriguingly, cells employ effective strategies echoing human transportation systems to manage such transportation hurdles.
- Jin-Sung Park
- , Il-Buem Lee
- & Minhaeng Cho
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Article
| Open AccessVisualization of accessible cholesterol using a GRAM domain-based biosensor
Regulated cholesterol transport is essential for the maintenance of cellular cholesterol distribution and homeostasis, but tools to monitor this process are limited. Here, the authors develop a genetically encoded cholesterol biosensor and demonstrate its use for visualising cellular cholesterol distribution in various live cells in real time.
- Dylan Hong Zheng Koh
- , Tomoki Naito
- & Yasunori Saheki
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Article
| Open AccessRevealing invisible cell phenotypes with conditional generative modeling
Biological research relies on observing cell phenotypes, often obscured by natural variability. Here, the authors use generative modelling to unveil hidden changes triggered by infections, mutations, or drugs, allowing for accessible discovery of biomarkers.
- Alexis Lamiable
- , Tiphaine Champetier
- & Auguste Genovesio
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Article
| Open AccessFacile hermetic TEM grid preparation for molecular imaging of hydrated biological samples at room temperature
Electron microscopy of native hydrated biological samples close to physiological temperature is challenging. Here, authors encapsulate proteins and cells as an efficient, cost-effective complement to other microscopy technique in addressing biological questions at the molecular level.
- Lingli Kong
- , Jianfang Liu
- & Gang Ren
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Article
| Open AccessMulti-scale dynamic imaging reveals that cooperative motility behaviors promote efficient predation in bacteria
Myxococcus xanthus forages and moves collectively to prey and feed on other bacterial species. Here, the authors challenge the conventional idea that during Myxococcus xanthus predation, A- and S-motilities are limited to specific forager and swarm roles and reveal a synergistic interaction between these motilities to enhance predation efficiency.
- Sara Rombouts
- , Anna Mas
- & Marcelo Nollmann
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Article
| Open AccessSingle-cell lipidomics enabled by dual-polarity ionization and ion mobility-mass spectrometry imaging
Single-cell analysis provides unique insight into individual cell dynamics and cell-to-cell heterogeneity. Here, the authors utilize trapped ion mobility separation coupled with dual-polarity ionization mass spectrometry imaging to enable high-throughput in situ profiling of single-cell lipidome.
- Hua Zhang
- , Yuan Liu
- & Lingjun Li
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Article
| Open AccessSelf-assembly of CIP4 drives actin-mediated asymmetric pit-closing in clathrin-mediated endocytosis
CIP4 drives an asymmetric closing process in clathrin-mediated endocytosis by LLPS-driven self assembly and stereospecific interaction with the curved membrane and actin-regulating proteins which generates an actin-rich microenvironment near the pit.
- Yiming Yu
- & Shige H. Yoshimura
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Article
| Open AccessHighly specific and non-invasive imaging of Piezo1-dependent activity across scales using GenEPi
Stretch-activated ion channel Piezo1 contribures to mechanotransduction in many tissues, but its output is mostly measured indirectly. Here, the authors introduce GenEPi, a fluorescent reporter for directly visualizing Piezo1 activation-dependent calcium influx.
- Sine Yaganoglu
- , Konstantinos Kalyviotis
- & Periklis Pantazis
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Article
| Open AccessNuclear lamina strain states revealed by intermolecular force biosensor
In this work, a nanobody based intermolecular strain sensor was used to follow the mechanical strain in the nuclear lamina. The results indicate that mechanical state of the nuclear lamina is not only affected by the cell contractility, but also chromatin packing.
- Brooke E. Danielsson
- , Bobin George Abraham
- & Teemu O. Ihalainen
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Article
| Open AccessMuscleblind-like proteins use modular domains to localize RNAs by riding kinesins and docking to membranes
RNA localization is mediated by kinesin motors and anchoring. However, mechanisms underlying specificity are unclear. Here, the authors find that MBNL protein’s zinc fingers prefer specific kinesins, and its unstructured tail mediates membrane anchoring.
- Ryan P. Hildebrandt
- , Kathryn R. Moss
- & Eric T. Wang
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Article
| Open AccessQuantitative structured illumination microscopy via a physical model-based background filtering algorithm reveals actin dynamics
Quantitative live-cell superresolution imaging that maintains the linearity of fluorescence signals remains difficult. Here, the authors propose a physical model-based background filtering method for 2D-SIM, which allows for quantitative imaging and high signal completeness.
- Yanquan Mo
- , Kunhao Wang
- & Liangyi Chen
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Article
| Open AccessAutophagy receptor NDP52 alters DNA conformation to modulate RNA polymerase II transcription
An autophagy receptor, NDP52, is recruited to the nucleus where it can bind DNA. The authors show this promotes changes in chromatin accessibility which supports transcription initiation, providing a direct link between autophagy and transcription regulation.
- Ália dos Santos
- , Daniel E. Rollins
- & Christopher P. Toseland
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Article
| Open AccessRAD-TGTs: high-throughput measurement of cellular mechanotype via rupture and delivery of DNA tension probes
Mechanical forces drive critical cellular processes, but methods to study such cellular forces are typically low-throughput. Here the authors present a method using “Rupture And Deliver” Tension Gauge Tethers, where flow cytometry or sequencing can be used to record the mechanical history of thousands of individual cells.
- Matthew R. Pawlak
- , Adam T. Smiley
- & Wendy R. Gordon
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Article
| Open AccessThe CellPhe toolkit for cell phenotyping using time-lapse imaging and pattern recognition
Approaches for temporal analysis and quantitative characterisation of single cell morphology and dynamics remain in high demand. Here authors present CellPhe, a pattern recognition toolkit for the unbiased characterisation of cellular phenotypes within time-lapse videos.
- Laura Wiggins
- , Alice Lord
- & Julie Wilson
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| Open AccessCryo-tomography reveals rigid-body motion and organization of apicomplexan invasion machinery
In this study, the authors use cryo-focused-ion-beammilling and cryo-electron tomography to image the apical complex of parasites in their native states. They report insights into the parasite invasion machinery in its protruded and retracted states in three dimensions, including all cytoskeletal assemblies, secretory organelles, and membranes intact.
- Long Gui
- , William J. O’Shaughnessy
- & Daniela Nicastro
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Article
| Open AccessSingle-molecule localization microscopy reveals the ultrastructural constitution of distal appendages in expanded mammalian centrioles
The authors have combined direct stochastic optical reconstruction microscopy with expansion microscopy to describe the 3-dimensional molecular organization of centriolar distal appendages.
- Ting-Jui Ben Chang
- , Jimmy Ching-Cheng Hsu
- & T. Tony Yang
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Article
| Open AccessMultiplexed analysis of EV reveals specific biomarker composition with diagnostic impact
Multiplexed analyses of near single EVs is currently challenging. Here the authors report the method MASEV, multiplexed analysis of EVs, to interrogate thousands of individual EVs during 5 cycles of multi-channel fluorescence staining for 15 EV biomarkers.
- Joshua D. Spitzberg
- , Scott Ferguson
- & Ralph Weissleder
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Article
| Open AccessVisualizing orthogonal RNAs simultaneously in live mammalian cells by fluorescence lifetime imaging microscopy (FLIM)
No multi-color RNA fluorescent tags are currently available for use in live cells. Here, the authors show that fluorescence lifetime imaging microscopy is advantageous for multiplexed RNA visualization while achieving robust cellular contrast.
- Nadia Sarfraz
- , Emilia Moscoso
- & Esther Braselmann
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Article
| Open AccessA conformational switch in clathrin light chain regulates lattice structure and endocytosis at the plasma membrane of mammalian cells
Conformational changes within proteins are thought to regulate clathrin-mediated endocytosis. Here, authors develop a correlative FRET and EM imaging method and uncover a structural switch in clathrin light chain that controls endocytosis in cells.
- Kazuki Obashi
- , Kem A. Sochacki
- & Justin W. Taraska
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Article
| Open AccessExocytosis of the silicified cell wall of diatoms involves extensive membrane disintegration
Exocytosis is a fundamental cellular process. Here, the authors report an unusual exocytosis mechanism in the silicified cell wall of diatoms, in which membrane patches are discarded.
- Diede de Haan
- , Lior Aram
- & Assaf Gal
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Article
| Open AccessRevealing the tissue-level complexity of endogenous glucagon-like peptide-1 receptor expression and signaling
Visualizing endogenous GPCRs is challenging. Here the authors generate mice with an enzyme self-label genome-edited into the endogenous glucagon-like peptide-1 receptor locus, design fluorescent dyes for specific labelling in complex tissue, and reveal tissue-level organisation and dynamics of an endogenous class B GPCR.
- Julia Ast
- , Daniela Nasteska
- & David J. Hodson
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Article
| Open AccessMachine learning assisted interferometric structured illumination microscopy for dynamic biological imaging
Structured Illumination Microscopy allows for the visualization of biological structures at resolutions below the diffraction limit, but this imaging modality is still hampered by high experimental complexity. Here, the authors present a combination of interferometry and machine learning to construct a structured illumination microscope for super resolution imaging of dynamic sub-cellular biological structures in multiple colors.
- Edward N. Ward
- , Lisa Hecker
- & Clemens F. Kaminski
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Article
| Open AccessSTREAMING-tag system reveals spatiotemporal relationships between transcriptional regulatory factors and transcriptional activity
Using the newly developed STREAMING-tag system, the authors find that clusters of RNA polymerase II and BRD4 are formed specifically in the transcriptionally active state near the Nanog gene in mouse embryonic stem cells.
- Hiroaki Ohishi
- , Seiru Shimada
- & Hiroshi Ochiai
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Article
| Open AccessAchieving single nucleotide sensitivity in direct hybridization genome imaging
Visualisation of point mutations in situ is informative for studying genetic diseases. Here the authors report single guide genome oligopaint via local denaturation fluorescence in situ hybridisation, sgGOLDFISH, a direct hybridisation genome imaging method with single-nucleotide sensitivity.
- Yanbo Wang
- , W. Taylor Cottle
- & Taekjip Ha
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Article
| Open AccessSepN is a septal junction component required for gated cell–cell communication in the filamentous cyanobacterium Nostoc
The filamentous cyanobacterium Nostoc regulates communication between sister cells via a conformational change in septal junctions. Here, the authors identify and characterize protein SepN as a component of septal junctions, and highlight the importance of controlling molecular diffusion between cells to ensure the survival of a multicellular organism.
- Ann-Katrin Kieninger
- , Piotr Tokarz
- & Iris Maldener
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Article
| Open AccessFlexible hyperspectral surface plasmon resonance microscopy
The authors developed a pristine hyperspectral SPR microscopy that enables monochromatic and polychromatic SPR imaging with flexible field-of-view option, single-pixel spectral SPR sensing and 2D quantification of thin films with resonant wavelength images.
- Ziwei Liu
- , Jingning Wu
- & Zhi-mei Qi
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Article
| Open AccessFibrocytes boost tumor-supportive phenotypic switches in the lung cancer niche via the endothelin system
Fibrocytes are monocyte-derived cells implicated in wound healing. Here, the authors utilise single cell RNA-seq, genetic ablation and multiplexed imaging to identify a fibrocyte population in lung cancer models, and use human lung cancer coculture systems to highlight their potential to modulate microenvironmental niche and sensitivity to endothelin blockade.
- Andreas Weigert
- , Xiang Zheng
- & Rajkumar Savai
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Article
| Open AccessSolving the MCM paradox by visualizing the scaffold of CMG helicase at active replisomes
For several decades the MCM2-7 proteins, the core of the DNA replicative helicase, eluded detection at DNA replication sites. Here, the authors solve this conundrum by gene editing, which enables visualization of replication dynamics in living cells.
- Hana Polasek-Sedlackova
- , Thomas C. R. Miller
- & Jiri Lukas
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Article
| Open AccessTwo Rac1 pools integrate the direction and coordination of collective cell migration
Previous studies suggested a chemokine receptor governed gradient of Rac1 activity is essential for collective guidance of Drosophila border cells. Here, Zhou et al. report that two distinct Rac1 pools at protrusions and cables, not Rac1 activity gradient, integrate the direction and coordination for collective guidance.
- Sijia Zhou
- , Peng Li
- & Xiaobo Wang
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Article
| Open AccessLive imaging and conditional disruption of native PCP activity using endogenously tagged zebrafish sfGFP-Vangl2
Planar cell polarity (PCP) is critical for tissue-wide coordination and successful development. Here Jussila et al. generate a GFP-Vangl2 fusion for live imaging and discover a surprising directionality to the intercellular propagation of cell polarity, and ultimately link PCP defects with idiopathic scoliosis.
- Maria Jussila
- , Curtis W. Boswell
- & Brian Ciruna
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Article
| Open AccessA multi-adenylate cyclase regulator at the flagellar tip controls African trypanosome transmission
Trypanosomes can sense signal molecules and coordinate their movement in response to such signals, a phenomenon termed social motility (SoMo). Here, Bachmaier et al show that cyclic AMP response protein 3 (CARP3) localization to the flagellar tip and its interaction with a number of different adenylate cyclases is essential for migration to tsetse fly salivary glands and for SoMo, therewith linking SoMo and cAMP signaling to trypanosome transmission.
- Sabine Bachmaier
- , Giacomo Giacomelli
- & Michael Boshart
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Article
| Open AccessLife at high temperature observed in vitro upon laser heating of gold nanoparticles
Studying microorganisms at high temperatures is challenging on conventional optical microscopes. Here, the authors introduce the concept of microscale laser heating over the full field of view by using gold nanoparticles as light absorbers, and study thermophile species up to 80 °C.
- Céline Molinaro
- , Maëlle Bénéfice
- & Guillaume Baffou
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Article
| Open AccessCtIP-dependent nascent RNA expression flanking DNA breaks guides the choice of DNA repair pathway
RNA has been implicated in DNA repair. This work shows that the interplay of RNAPII-generated nascent RNA, RNA:DNA hybrids and the resection factor CtIP guide DNA double strand break repair pathway choice towards error-free homologous recombination.
- Daniel Gómez-Cabello
- , George Pappas
- & Jiri Bartek
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Article
| Open AccessIsoform-specific and ubiquitination dependent recruitment of Tet1 to replicating heterochromatin modulates methylcytosine oxidation
A short isoform of the Tet1 enzyme (Tet1s) that oxidizes the DNA 5-methylcytosine (5mC) mark is overexpressed in tumors. Here the authors show Tet1s, but not full length Tet1, changes localization over the cell cycle upon ubiquitination and Uhrf1 interaction and is targeted to heterochromatin during S-phase. This leads to 5mC oxidation and loss of DNA methylation in heterochromatin.
- María Arroyo
- , Florian D. Hastert
- & M. Cristina Cardoso
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Article
| Open AccessNanoscale segregation of channel and barrier claudins enables paracellular ion flux
Meshworks of claudin polymers control the paracellular transport and barrier properties of epithelial tight junctions. Here, the authors show different claudin nanoscale organization principles, finding that claudin segregation enables barrier formation and paracellular ion flux across tight junctions.
- Hannes Gonschior
- , Christopher Schmied
- & Martin Lehmann
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Article
| Open AccessMultiscale light-sheet organoid imaging framework
Live imaging of organoid growth remains a challenge: it requires long-term imaging of several samples simultaneously and dedicated analysis pipelines. Here the authors report an experimental and image processing framework to turn long-term light-sheet imaging of intestinal organoids into digital organoids.
- Gustavo de Medeiros
- , Raphael Ortiz
- & Prisca Liberali