Abstract
Elevation in the cytosolic Ca2+ concentration triggers exocytosis of lysosomes in many cell types. This chapter describes a method to detect lysosomal exocytosis in mammalian cells, which takes advantage of the presence of an abundant glycoprotein, Lamp1, on the membrane of lysosomes. Lamp1 is a transmembrane protein with a large, heavily glycosylated region that faces the lumen of lysosomes. When lysosomes fuse with the plasma membrane, epitopes present on the luminal domain of Lamp1 are exposed on the cell surface. The Lamp1 luminal epitopes can then be detected on the surface of live, unfixed cells using highly specific monoclonal antibodies and fluorescence microscopy. The main advantage of this method is its sensitivity, and the fact that it provides spatial information on lysosomal exocytosis at the single cell level.
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References
de Duve C (2005) The lysosome turns fifty. Nat Cell Biol 7(9):847–849. doi:10.1038/ncb0905–847
Saftig P, Klumperman J (2009) Lysosome biogenesis and lysosomal membrane proteins: trafficking meets function. Nat Rev Mol Cell Biol 10(9):623–635. doi:10.1038/nrm2745
Rodriguez A, Webster P, Ortego J, Andrews NW (1997) Lysosomes behave as Ca2 + −regulated exocytic vesicles in fibroblasts and epithelial cells. J Cell Biol 137:93–104
Andrews NW (2000) Regulated secretion of conventional lysosomes. Trends Cell Biol 10(8):316–321
Griffiths GM (1996) Secretory lysosomes - a special mechanism of regulated secretion in haemopoietic cells. Trends Cell Biol 6:329–332
Reddy A, Caler E, Andrews N (2001) Plasma membrane repair is mediated by Ca2+−regulated exocytosis of lysosomes. Cell 106:157–169
Andrews NW, Almeida PE, Corrotte M (2014) Damage control: cellular mechanisms of plasma membrane repair. Trends Cell Biol 24(12):734–742. doi:10.1016/j.tcb.2014.07.008
Jaiswal JK, Andrews NW, Simon SM (2002) Membrane proximal lysosomes are the major vesicles responsible for calcium-dependent exocytosis in nonsecretory cells. J Cell Biol 159(4):625–635. doi:10.1083/jcb.200208154
Castro-Gomes T, Corrotte M, Tam C, Andrews NW (2016) Plasma membrane repair is regulated extracellularly by proteases released from lysosomes. PLoS One 11(3):e0152583. doi:10.1371/journal.pone.0152583
Miller H, Castro-Gomes T, Corrotte M, Tam C, Maugel TK, Andrews NW, Song W (2015) Lipid raft-dependent plasma membrane repair interferes with the activation of B lymphocytes. J Cell Biol 211(6):1193–1205. doi:10.1083/jcb.201505030
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Andrews, N.W. (2017). Detection of Lysosomal Exocytosis by Surface Exposure of Lamp1 Luminal Epitopes. In: Öllinger, K., Appelqvist, H. (eds) Lysosomes. Methods in Molecular Biology, vol 1594. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6934-0_13
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DOI: https://doi.org/10.1007/978-1-4939-6934-0_13
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