K. Hirschberg et al., Kinetic analysis of secretory protein traffic and characterization of Golgi to plasma membrane transport intermediates in living cells, J CELL BIOL, 143(6), 1998, pp. 1485-1503
Quantitative time-lapse imaging data of single cells expressing the transme
mbrane protein, vesicular stomatitis virus ts045 G protein fused to green f
luorescent protein (VSVG-GFP), were used for kinetic modeling of protein tr
affic through the various compartments of the secretory pathway. A series o
f first order rate laws was sufficient to accurately describe VSVG-GFP tran
sport, and provided compartment residence times and rate constants for tran
sport into and out of the Golgi complex and delivery to the plasma membrane
. For ER to Golgi transport the mean rate constant (i.e., the fraction of V
SVG-GFP moved per unit of time) was 2.8% per min, for Golgi to plasma membr
ane transport it was 3.0% per min, and for transport from the plasma membra
ne to a degradative site it was 0.25% per min. Because these rate constants
did not change as the concentration of VSVG-GFP in different compartments
went from high (early in the experiment) to low (late in the experiment), s
ecretory transport machinery was never saturated during the experiments.
The processes of budding, translocation, and fusion of post-Golgi transport
intermediates carrying VSVG-GFP to the plasma membrane were also analyzed
using quantitative imaging techniques. Large pleiomorphic tubular structure
s, rather than small vesicles, were found to be the primary vehicles for Go
lgi to plasma membrane transport of VSVG-GFP, These structures budded as en
tire domains from the Golgi complex and underwent dynamic shape changes as
they moved along microtubule tracks to the cell periphery. They carried up
to 10,000 VSVG-GFP molecules and had a mean life time in COS cells of 3.8 m
in. In addition, they fused with the plasma membrane without intersecting o
ther membrane transport pathways in the cell. These properties suggest that
the post-Golgi intermediates represent a unique transport organelle for co
nveying large quantities of protein cargo from the Golgi complex directly t
o the plasma membrane.