Patch-Based Nonlocal Functional for Denoising Fluorescence Microscopy Image Sequences

Boulanger, Jérôme; Kervrann, Charles; Bouthémy, Patrick; Elbau, Peter; Sibarita, Jean‐Baptiste; Salamero, Jean

doi:10.1109/tmi.2009.2033991

Cited by 267 publications

(276 citation statements)

References 65 publications

(70 reference statements)

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“…FLIP experiments were performed using the same system; however, the images were recorded every 250 ms. FRAP images were denoised using the ND-Safir software on 10 cells for each condition. This procedure, developed by Boulanger et al (47), improves the signal to noise ratio and is crucial for the quantitative analysis of FRAP data. Once denoised, the images were normalized by the fluorescence intensity of the bleached area just before the bleach.…”

Section: Methodsmentioning

confidence: 99%

Severe osmotic compression triggers a slowdown of intracellular signaling, which can be explained by molecular crowding

Miermont

Waharte

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

180

169

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Regulation of the cellular volume is fundamental for cell survival and function. Deviations from equilibrium trigger dedicated signaling and transcriptional responses that mediate water homeostasis and volume recovery. Cells are densely packed with proteins, and molecular crowding may play an important role in cellular processes. Indeed, increasing molecular crowding has been shown to modify the kinetics of biochemical reactions in vitro; however, the effects of molecular crowding in living cells are mostly unexplored. Here, we report that, in yeast, a sudden reduction in cellular volume, induced by severe osmotic stress, slows down the dynamics of several signaling cascades, including the stress-response pathways required for osmotic adaptation. We show that increasing osmotic compression decreases protein mobility and can eventually lead to a dramatic stalling of several unrelated signaling and cellular processes. The rate of these cellular processes decreased exponentially with protein density when approaching stalling osmotic compression. This suggests that, under compression, the cytoplasm behaves as a soft colloid undergoing a glass transition. Our results shed light on the physical mechanisms that force cells to cope with volume fluctuations to maintain an optimal protein density compatible with cellular functions.

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Section: Methodsmentioning

confidence: 99%

Severe osmotic compression triggers a slowdown of intracellular signaling, which can be explained by molecular crowding

Miermont

Waharte

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

180

169

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“…4B, contours were normalized to remove size differences and centered via their centers of mass. Filament orientations and lengths were quantified under different conditions by using images of sperm from peel-1 promotor strains (EG5897) denoised using the program Safir (22) and analyzed manually in Metamorph. For length measurements, fibers were assumed to be straight.…”

Section: Methodsmentioning

confidence: 99%

Membrane tension regulates motility by controlling lamellipodium organization

Batchelder

Hollopeter

Campillo

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

132

114

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Many cell movements proceed via a crawling mechanism, where polymerization of the cytoskeletal protein actin pushes out the leading edge membrane. In this model, membrane tension has been seen as an impediment to filament growth and cell motility. Here we use a simple model of cell motility, the Caenorhabditis elegans sperm cell, to test how membrane tension affects movement and cytoskeleton dynamics. To enable these analyses, we create transgenic worm strains carrying sperm with a fluorescently labeled cytoskeleton. Via osmotic shock and deoxycholate treatments, we relax or tense the cell membrane and quantify apparent membrane tension changes by the membrane tether technique. Surprisingly, we find that membrane tension reduction is correlated with a decrease in cell displacement speed, whereas an increase in membrane tension enhances motility. We further demonstrate that apparent polymerization rates follow the same trends. We observe that membrane tension reduction leads to an unorganized, rough lamellipodium, composed of short filaments angled away from the direction of movement. On the other hand, an increase in tension reduces lateral membrane protrusions in the lamellipodium, and filaments are longer and more oriented toward the direction of movement. Overall we propose that membrane tension optimizes motility by streamlining polymerization in the direction of movement, thus adding a layer of complexity to our current understanding of how membrane tension enters into the motility equation. I n crawling cells, motility is mainly driven by actin polymerization, which forms filaments beneath the leading edge cell membrane to make protrusions (1). In this scenario, polymerization is inhibited by the presence of the cell membrane, and membrane tension is thus commonly seen as an impediment to cell motility (2). Experiments show that lamellipodial extension rate is indeed inversely correlated with membrane tension (3), but steady-state, whole cell translocation has not been studied. Here we use a simplified system of crawling cell motility, the Caenorhabditis elegans sperm cell, in order to address the question of how membrane tension affects whole cell translocation. The sperm cell contains only cytoskeleton, mitochondria, and nuclear material and is incapable of de novo protein synthesis or classical exo-and endocytosis, thus representing a useful model for exploring the interplay between membrane tension and cell motility. The sperm cell translocates by adhering to the substrate and emitting a dynamic lamellipodia, in a manner that is morphologically similar to actomyosin containing motile cells, despite the fact that the movement of sperm cells is powered by the dynamics of the major sperm protein (MSP) cytoskeleton in the absence of actin and known molecular motors (4, 5).The production of fluorescently labeled actin in living cells was a watershed in the understanding of how actin structures assemble, flow, and disassemble to produce cell motility. However, due to efficient gene silencing mechanisms in th...

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“…A total of 180 3D stack images were first denoised using Nd Safir (Boulanger et al, 2010) before being treated within the pipeline. First, the extraction of the volumes from individualized OBs through the pipeline is based on the watershed procedure dedicated to the separation of touching objects (Soille, 2003).…”

Section: Quantitative Analyses Of Ob Size Distribution and Dispersionmentioning

confidence: 99%

Specialization of Oleosins in Oil Body Dynamics during Seed Development in Arabidopsis Seeds

et al. 2014

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Oil bodies (OBs) are seed-specific lipid storage organelles that allow the accumulation of neutral lipids that sustain plantlet development after the onset of germination. OBs are covered with specific proteins embedded in a single layer of phospholipids. Using fluorescent dyes and confocal microscopy, we monitored the dynamics of OBs in living Arabidopsis (Arabidopsis thaliana) embryos at different stages of development. Analyses were carried out with different genotypes: the wild type and three mutants affected in the accumulation of various oleosins (OLE1, OLE2, and OLE4), three major OB proteins. Image acquisition was followed by a detailed statistical analysis of OB size and distribution during seed development in the four dimensions (x, y, z, and t). Our results indicate that OB size increases sharply during seed maturation, in part by OB fusion, and then decreases until the end of the maturation process. In single, double, and triple mutant backgrounds, the size and spatial distribution of OBs are modified, affecting in turn the total lipid content, which suggests that the oleosins studied have specific functions in the dynamics of lipid accumulation.The seed is a complex, specific structure that allows a quiescent plant embryo to cope with unfavorable germinating conditions and also permits dissemination of the species. To achieve these functions, seeds accumulate reserve compounds that will ensure the survival of the embryo and fuel the growth of the plantlet upon germination. Accumulation of lipids occurs in many eukaryotic cells and is a rather common means of storing carbon and energy. Lipid droplets (LDs) can be found in all eukaryotes, such as yeast (Saccharomyces cerevisiae;

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Patch-Based Nonlocal Functional for Denoising Fluorescence Microscopy Image Sequences

Cited by 267 publications

References 65 publications

Severe osmotic compression triggers a slowdown of intracellular signaling, which can be explained by molecular crowding

Severe osmotic compression triggers a slowdown of intracellular signaling, which can be explained by molecular crowding

Membrane tension regulates motility by controlling lamellipodium organization

Specialization of Oleosins in Oil Body Dynamics during Seed Development in Arabidopsis Seeds

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