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Bio Protoc. 2016 Feb 20;6(4). pii: e1739.

VAMP8-3xHA Uptake Assay in HeLa Cells.

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Département d'anatomie et de biologie cellulaire, Faculté de Médecine et des Sciences de la Santé. Université de Sherbrooke, Sherbrooke, Canada.
Division of Biological Sciences, University of California, San Diego, USA.


Transmembrane proteins are rarely exclusively localized to a specific vesicle or an organelle. Most transmembrane proteins undergo complicated trafficking routes. Thus, transmembrane proteins are under constant flux, and at steady state, found on a variety of vesicles or organelles. This characteristic makes the study of their trafficking routes complex, since at any given moment, different molecules are often being trafficked in opposing directions. Pulse-chase experiments can temporally track a specific pool of a transmembrane protein of interest, allowing for the kinetic description of its trafficking route. This type of technique has been used extensively to follow a large array of plasma membrane localized proteins (Diril et al., 2006; Jean et al., 2010). Here, we describe a method that allows the study of VAMP8 trafficking from the plasma membrane to endolysosomal compartments. This method was used to describe a role for MTMR13 and RAB21 in the regulation of VAMP8 trafficking to endolysosomes (Jean et al., 2015).


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