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Tracking Trafficking of Notch and Its Ligands in Mammalian Cells

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Notch Signaling

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1187))

Abstract

The Notch receptor and its ligands are cell surface transmembrane proteins that are internalized. Endocytosis and vesicle trafficking play key roles in Notch signaling activation and modulation. In mammalian cultured cells it is possible to track these cell surface molecules by pulse-labeling these proteins in vivo. One labeling protocol consists in the covalent linkage of membrane-impermeable biotin followed by western blotting. An alternative protocol consists of using high affinity antibodies against the extracellular domains of the proteins followed by immunofluorescence, thereby allowing monitoring of the fate of the labeled proteins. In this chapter, we will describe these two approaches to study the dynamics of receptor and ligand trafficking.

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Acknowledgments

We thank Loredana Puca for critical reading of the manuscript, J. Moretti, S. Heuss, and F. Logeat for discussion and materials. Financial support of Institut Pasteur, CNRS, and Ligue Nationale Contre le Cancer (LNCC RS11/75-21) is acknowledged.

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Authors and Affiliations

  1. Signalisation Moléculaire et Activation Cellulaire, Institut Pasteur and CNRS, 25, Rue du Docteur Roux, 75724, Paris Cedex 15, France

    Patricia Chastagner & Christel Brou

Authors
  1. Patricia Chastagner
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  2. Christel Brou
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Corresponding author

Correspondence to Christel Brou .

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Editors and Affiliations

  1. Dept.of Molecular and Human Genetics, Program in Developmental Biology, Dept. of Neuroscience, Jan and Dan Duncan Neurological Research Institute at Texas Children's Hospital, Howard Hughes Medical Institute, Baylor College of Medicine, Houston, Texas, USA

    Hugo J. Bellen

  2. Dept. of Molecular and Human Genetics, Program in Developmental Biology, Jan and Dan Duncan Neurological Research Institute at Texas Children's Hospital, Baylor College of Medicine, Houston, Texas, USA

    Shinya Yamamoto

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Chastagner, P., Brou, C. (2014). Tracking Trafficking of Notch and Its Ligands in Mammalian Cells. In: Bellen, H., Yamamoto, S. (eds) Notch Signaling. Methods in Molecular Biology, vol 1187. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1139-4_7

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  • DOI: https://doi.org/10.1007/978-1-4939-1139-4_7

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1138-7

  • Online ISBN: 978-1-4939-1139-4

  • eBook Packages: Springer Protocols

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