scholarly journals Mapping the Emergent Spatial Organization of Mammalian Cells using Micropatterns and Quantitative Imaging

Author(s):  
Darren Wisniewski ◽  
Sally Lowell ◽  
Guillaume Blin
2015 ◽  
Vol 18 (4) ◽  
pp. 478-488 ◽  
Author(s):  
Kristen A. Earle ◽  
Gabriel Billings ◽  
Michael Sigal ◽  
Joshua S. Lichtman ◽  
Gunnar C. Hansson ◽  
...  

2014 ◽  
Vol 25 (22) ◽  
pp. 3610-3618 ◽  
Author(s):  
Robert Mahen ◽  
Birgit Koch ◽  
Malte Wachsmuth ◽  
Antonio Z. Politi ◽  
Alexis Perez-Gonzalez ◽  
...  

Fluorescence tagging of proteins is a widely used tool to study protein function and dynamics in live cells. However, the extent to which different mammalian transgene methods faithfully report on the properties of endogenous proteins has not been studied comparatively. Here we use quantitative live-cell imaging and single-molecule spectroscopy to analyze how different transgene systems affect imaging of the functional properties of the mitotic kinase Aurora B. We show that the transgene method fundamentally influences level and variability of expression and can severely compromise the ability to report on endogenous binding and localization parameters, providing a guide for quantitative imaging studies in mammalian cells.


2018 ◽  
Vol 115 (21) ◽  
pp. 5438-5443 ◽  
Author(s):  
Anerudh Kannan ◽  
Zhenbin Yang ◽  
Minyoung Kevin Kim ◽  
Howard A. Stone ◽  
Albert Siryaporn

Bacteria colonize environments that contain networks of moving fluids, including digestive pathways, blood vasculature in animals, and the xylem and phloem networks in plants. In these flow networks, bacteria form distinct biofilm structures that have an important role in pathogenesis. The physical mechanisms that determine the spatial organization of bacteria in flow are not understood. Here, we show that the bacteriumP. aeruginosacolonizes flow networks using a cyclical process that consists of surface attachment, upstream movement, detachment, movement with the bulk flow, and surface reattachment. This process, which we have termed dynamic switching, distributes bacterial subpopulations upstream and downstream in flow through two phases: movement on surfaces and cellular movement via the bulk. The model equations that describe dynamic switching are identical to those that describe dynamic instability, a process that enables microtubules in eukaryotic cells to search space efficiently to capture chromosomes. Our results show that dynamic switching enables bacteria to explore flow networks efficiently, which maximizes dispersal and colonization and establishes the organizational structure of biofilms. A number of eukaryotic and mammalian cells also exhibit movement in two phases in flow, which suggests that dynamic switching is a modality that enables efficient dispersal for a broad range of cell types.


2005 ◽  
Vol 95 (15) ◽  
Author(s):  
Chinmay Pangarkar ◽  
Anh Tuan Dinh ◽  
Samir Mitragotri

2015 ◽  
Vol 26 (13) ◽  
pp. 2519-2534 ◽  
Author(s):  
Mini Jose ◽  
Sylvain Tollis ◽  
Deepak Nair ◽  
Romain Mitteau ◽  
Christophe Velours ◽  
...  

The coupling of endocytosis and exocytosis underlies fundamental biological processes ranging from fertilization to neuronal activity and cellular polarity. However, the mechanisms governing the spatial organization of endocytosis and exocytosis require clarification. Using a quantitative imaging-based screen in budding yeast, we identified 89 mutants displaying defects in the localization of either one or both pathways. High-resolution single-vesicle tracking revealed that the endocytic and exocytic mutants she4∆ and bud6∆ alter post-Golgi vesicle dynamics in opposite ways. The endocytic and exocytic pathways display strong interdependence during polarity establishment while being more independent during polarity maintenance. Systems analysis identified the exocyst complex as a key network hub, rich in genetic interactions with endocytic and exocytic components. Exocyst mutants displayed altered endocytic and post-Golgi vesicle dynamics and interspersed endocytic and exocytic domains compared with control cells. These data are consistent with an important role for the exocyst in coordinating endocytosis and exocytosis.


2021 ◽  
Author(s):  
Romain Lopez ◽  
Baoguo Li ◽  
Hadas Keren-Shaul ◽  
Pierre Boyeau ◽  
Merav Kedmi ◽  
...  

The function of mammalian cells is largely influenced by their tissue microenvironment. Advances in spatial transcriptomics open the way for studying these important determinants of cellular function by enabling a transcriptome-wide evaluation of gene expression in situ. A critical limitation of the current technologies, however, is that their resolution is limited to niches (spots) of sizes well beyond that of a single cell, thus providing measurements for cell aggregates which may mask critical interactions between neighboring cells of different types. While joint analysis with single-cell RNA-sequencing (scRNA-seq) can be leveraged to alleviate this problem, current analyses are limited to a discrete view of cell type proportion inside every spot. This limitation becomes critical in the common case where, even within a cell type, there is a continuum of cell states that cannot be clearly demarcated but reflects important differences in the way cells function and interact with their surroundings. To address this, we developed Deconvolution of Spatial Transcriptomics profiles using Variational Inference (DestVI), a probabilistic method for multi-resolution analysis for spatial transcriptomics that explicitly models continuous variation within cell types. Using simulations, we demonstrate that DestVI is capable of providing higher resolution compared to the existing methods and that it can estimate gene expression by every cell type inside every spot. We then introduce an automated pipeline that uses DestVI for analysis of single tissue slices and comparison between tissues. We apply this pipeline to study the immune crosstalk within lymph nodes to infection and explore the spatial organization of a mouse tumor model. In both cases, we demonstrate that DestVI can provide a high resolution and accurate spatial characterization of the cellular organization of these tissues, and that it is capable of identifying important cell-type-specific changes in gene expression - between different tissue regions or between conditions. DestVI is available as an open-source software package in the scvi-tools codebase (https://scvi-tools.org).


Author(s):  
John D. Finan ◽  
Farshid Guilak

The spatial organization of the genome influences its function [1]. Therefore, physical signals that deform the nucleus and the genome within may directly affect gene transcription and translation. In articular chondrocytes, nuclear deformation in response to osmotic stress is not sensitive to actin organization [2]. However, articular chondrocytes differ from most mammalian cells in that they remain round with cortically organized actin in monolayer culture. Adherent cells such as adipose stem cells (ASCs) spread in monolayer culture, forming a more typical, highly bundled actin cytoskeleton. These actin bundles exert tensile stress on the nucleus so we hypothesized that the osmotic sensitivity of the cell nucleus would be modulated by actin organization in ASCs. The osmotic sensitivity of the nucleus was quantified by measuring changes in the size and shape of the nucleus and the spatial arrangement of the chromatin within using 3D confocal microscopy.


2006 ◽  
Vol 290 (1) ◽  
pp. R11-R26 ◽  
Author(s):  
Erich Gulbins ◽  
Pin Lan Li

Activation of cells by receptor- and nonreceptor-mediated stimuli not only requires a change in the activity of signaling proteins but also requires a reorganization of the topology of the signalosom in the cell. The cell membrane contains distinct domains, rafts that serve the spatial organization of signaling molecules in the cell. Many receptors or stress stimuli transform rafts by the generation of ceramide. These stimuli activate the acid sphingomyelinase and induce a translocation of this enzyme onto the extracellular leaflet of the cell membrane. Surface acid sphingomyelinase generates ceramide that serves to fuse small rafts and to form large ceramide-enriched membrane platforms. These platforms cluster receptor molecules, recruit intracellular signaling molecules to aggregated receptors, and seem to exclude inhibitory signaling factors. Thus ceramide-enriched membrane platforms do not seem to be part of a specific signaling pathway but may facilitate and amplify the specific signaling elicited by the cognate stimulus. This general function may enable these membrane domains to be critically involved in the induction of apoptosis by death receptors and stress stimuli, bacterial and viral infections of mammalian cells, and the regulation of cardiovascular functions.


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