TY - JOUR
T1 - A functional GFP-fusion for imaging clathrin-mediated endocytosis
AU - Rappoport, Joshua
AU - Simon, SM
PY - 2008/5/21
Y1 - 2008/5/21
N2 - The ability to localize proteins of interest in live cells through imaging inherently fluorescent protein tags has provided an unprecedented level of information on cellular organization. However, there are numerous cases where fluorescent tags alter the localization and/or function of the proteins to which they are appended. Clathrin-mediated endocytosis from the plasma membrane is a physiologically important process evolutionarily conserved from yeast to humans. Some proteins that are associated with the machinery of clathrin-mediated endocytosis have been tagged with fluorescent proteins. However, it has not yet been possible to study this process through a protein marker that is specific to this step and still fully functional when linked to a fluorescent protein. Here we present the first demonstration that one of these proteins, in this case a GFP fusion to alpha-adaptin, a marker of the AP-2 complex, functionally complements knockdown of endogenous protein through siRNA silencing. GFP-alpha-adaptin, as well as the techniques used to test the fusion protein, represents an important contribution to the Cell Biologist's Toolbox, which will permit a greater understanding vesicle trafficking in live cells.
AB - The ability to localize proteins of interest in live cells through imaging inherently fluorescent protein tags has provided an unprecedented level of information on cellular organization. However, there are numerous cases where fluorescent tags alter the localization and/or function of the proteins to which they are appended. Clathrin-mediated endocytosis from the plasma membrane is a physiologically important process evolutionarily conserved from yeast to humans. Some proteins that are associated with the machinery of clathrin-mediated endocytosis have been tagged with fluorescent proteins. However, it has not yet been possible to study this process through a protein marker that is specific to this step and still fully functional when linked to a fluorescent protein. Here we present the first demonstration that one of these proteins, in this case a GFP fusion to alpha-adaptin, a marker of the AP-2 complex, functionally complements knockdown of endogenous protein through siRNA silencing. GFP-alpha-adaptin, as well as the techniques used to test the fusion protein, represents an important contribution to the Cell Biologist's Toolbox, which will permit a greater understanding vesicle trafficking in live cells.
U2 - 10.1111/j.1600-0854.2008.00770.x
DO - 10.1111/j.1600-0854.2008.00770.x
M3 - Article
C2 - 18498437
SN - 1600-0854
SN - 1600-0854
SN - 1600-0854
SN - 1600-0854
SN - 1600-0854
JO - Traffic
JF - Traffic
ER -