Abstract
In order to understand the mechanisms through which apical and basolateral membrane proteins achieve their subcellular distributions in polarized epithelial cells, it is critical to develop techniques that permit the selective observation of newly synthesized populations of these proteins. The SNAP tag system permits the detection and visualization of distinct spatially and temporally defined cohorts of tagged proteins. Thus, this technique is especially well suited to studying the trafficking routes pursued by newly synthesized proteins. The SNAP tag can be applied in the setting of fixed or live cell fluorescence microscopic analysis and can also be used in the context of various biochemical approaches. Here, we describe the use of the SNAP tag in association with confocal microscopy and SDS-PAGE to follow the biosynthetic pool of a membrane protein as it exits from the trans-Golgi network and makes its way to the plasma membrane.
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Acknowledgements
The authors would like to thank Dr. Ivan Correa for helpful discussions and support. This material is based upon work supported by NIH training grant 5T32GM007223-35 and National Science Foundation Graduate Research Fellowship Grant No. DGE-1122492 (E.H. Stoops) and NIH grants DK17433 and DK072612 (M.J. Caplan).
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Stoops, E.H., Farr, G.A., Hull, M., Caplan, M.J. (2014). SNAP-Tag to Monitor Trafficking of Membrane Proteins in Polarized Epithelial Cells. In: Ivanov, A. (eds) Exocytosis and Endocytosis. Methods in Molecular Biology, vol 1174. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0944-5_11
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DOI: https://doi.org/10.1007/978-1-4939-0944-5_11
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