RAL-1 controls multivesicular body biogenesis and exosome secretion

Hyenne, Vincent; Apaydin, Ahmet; Rodriguez, David L.; Spiegelhalter, Coralie; Hoff-Yoessle, Sarah; Diem, Maxime; Tak, Saurabh; Lefèbvre, Olivier; Schwab, Yannick; Goetz, Jacky G.; Labouesse, Michel

doi:10.1083/jcb.201504136

Cited by 199 publications

(204 citation statements)

References 54 publications

Supporting

Mentioning

198

Contrasting

Unclassified

Order By: Relevance

“…the zebrafish lateral line primordium, are deposited through the assembly of microluminal structures that constrain FGF signaling (Durdu et al, 2014). Studies performed in C. elegans provide additional examples of laser-assisted targeted ultramicrotomy; these have helped to further clarify at high resolution the formation of excretory canals, as well as the contribution of exosomes to alae formation (Hyenne et al, 2015) (Fig. 4A).…”

Section: Ground-breaking In Vitro Clemmentioning

confidence: 99%

Seeing is believing: multi-scale spatio-temporal imaging towards in vivo cell biology

Follain

Mercier

Osmani

et al. 2016

Journal of Cell Science

Self Cite

View full text Add to dashboard Cite

Life is driven by a set of biological events that are naturally dynamic and tightly orchestrated from the single molecule to entire organisms. Although biochemistry and molecular biology have been essential in deciphering signaling at a cellular and organismal level, biological imaging has been instrumental for unraveling life processes across multiple scales. Imaging methods have considerably improved over the past decades and now allow to grasp the inner workings of proteins, organelles, cells, organs and whole organisms. Not only do they allow us to visualize these events in their most-relevant context but also to accurately quantify underlying biomechanical features and, so, provide essential information for their understanding. In this Commentary, we review a palette of imaging (and biophysical) methods that are available to the scientific community for elucidating a wide array of biological events. We cover the most-recent developments in intravital imaging, light-sheet microscopy, superresolution imaging, and correlative light and electron microscopy. In addition, we illustrate how these technologies have led to important insights in cell biology, from the molecular to the whole-organism resolution. Altogether, this review offers a snapshot of the current and state-of-the-art imaging methods that will contribute to the understanding of life and disease.

show abstract

Section: Ground-breaking In Vitro Clemmentioning

confidence: 99%

Seeing is believing: multi-scale spatio-temporal imaging towards in vivo cell biology

Follain

Mercier

Osmani

et al. 2016

Journal of Cell Science

Self Cite

View full text Add to dashboard Cite

show abstract

“…2), including proteins in the cytoskeleton 12,15 (vimentin, tubulin, and actin), gap junctions (Connexin43) 12 , chromatin (histone 2B) 12 , plasma membrane 14,16,17 , Golgi apparatus 18 , exosomes 19 , endosomes 14 , lipid droplets 14 , cytoplasm 14 , and a variety of mitochondrial sub-compartments (matrix, intermembrane space, and outer membrane) 12,15,20–26 . Because the DAB reaction product does not cross membranes, APEX2 can be used to elucidate the topology of transmembrane proteins, as we have demonstrated for several mitochondrial proteins 12,15,21 .…”

Section: Introductionmentioning

confidence: 99%

“…Although this Protocol focuses on EM imaging in cultured mammalian cells, APEX2 can be used in essentially any cell type, including bacteria 38 , the eukaryotic parasite Giardia lamblia 39 , and yeast 18 , as well as complex tissues and organisms, including zebrafish 14,28 , Caenorhabditis elegans 19 , Drosophila 40 , and mice 41 . APEX2 is especially useful for EM applications in vivo because, unlike existing genetic tags (see below), APEX2 does not require irradiation with light or exogenous delivery of large molecules such as antibodies and nanoparticles.…”

Section: Introductionmentioning

confidence: 99%

“…Although antibody-based techniques label the protein of interest with nanoparticles of defined dimensions, resulting in EM contrast that cannot spread, the antibody–nanoparticle conjugates themselves have dimensions >10 nm, which inherently limits the resolution. The requirement for the exogenous labeling reagents, DAB and H 2 O 2 , is an unavoidable aspect of APEX2 staining, but these reagents diffuse readily into cultured cells and even complex tissues 14,19,28,40,41 , indicating that reagent delivery should not limit the application of APEX2 in most systems. Another drawback inherent to APEX2 is that the electron-dense DAB reaction product can potentially obscure endogenous contrast from the labeled structure and its immediate surrounding environment.…”

Section: Introductionmentioning

confidence: 99%

“…This Protocol uses Durcupan ACM resin, but most alternative resins are compatible with APEX2 samples, including CY212 (ref. 31), Epon 19,24 , Spurr’s, or Procure 812 (ref. 42).…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Electron microscopy using the genetically encoded APEX2 tag in cultured mammalian cells

et al. 2017

View full text Add to dashboard Cite

Electron microscopy (EM) is the premiere technique for high-resolution imaging of cellular ultrastructure. Unambiguous identification of specific proteins or cellular compartments in electron micrographs, however, remains challenging because of difficulties in delivering electron-dense contrast agents to specific subcellular targets within intact cells. We recently reported enhanced ascorbate peroxidase 2 (APEX2) as a broadly applicable genetic tag that generates EM contrast on a specific protein or subcellular compartment of interest. This protocol provides guidelines for designing and validating APEX2 fusion constructs, along with detailed instructions for cell culture, transfection, fixation, heavy-metal staining, embedding in resin, and EM imaging. Although this protocol focuses on EM in cultured mammalian cells, APEX2 is applicable to many cell types and contexts, including intact tissues and organisms, and is useful for numerous applications beyond EM, including live-cell proteomic mapping. This protocol, which describes procedures for sample preparation from cell monolayers and cell pellets, can be completed in 10 d, including time for APEX2 fusion construct validation, cell growth, and solidification of embedding resins. Notably, the only additional steps required relative to a standard EM sample preparation are cell transfection and a 2- to 45-min staining period with 3,3′-diaminobenzidine (DAB) and hydrogen peroxide (H2O2).

show abstract

Biogenesis and Identification of Extracellular Vesicles

Ding,

Zhang,

Zhao

et al. 2023

Biomedical Applications of Extracellular Vesicles

View full text Add to dashboard Cite

RAL-1 controls multivesicular body biogenesis and exosome secretion

Abstract: Abbreviations used in this paper: CA, constitutively active; DN, dominant negative; ESC RT, endosomal sorting complex required for transport; EV, extracellular vesicle; HPF, high-pressure freezing; ILV, intraluminal vesicle; MVB, multivesicular body; sgRNA, single guide RNA; WT, wild type.

Cited by 199 publications

References 54 publications

Seeing is believing: multi-scale spatio-temporal imaging towards in vivo cell biology

Seeing is believing: multi-scale spatio-temporal imaging towards in vivo cell biology

Electron microscopy using the genetically encoded APEX2 tag in cultured mammalian cells

Biogenesis and Identification of Extracellular Vesicles

Contact Info

Product

Resources

About