extended
binding
studies
confirmed
above
rab5-binding
site
vivo
cell
monitoring
rab5-mediated
appl1
recruitment
early
endosomes
cell
via
confocal
microscopy
case
rfp
dsred-monomer
rab5-q79l
fusion
protein
expressed
pc12
cells
targeted
early
endosomes
recruited
effectively
coexpressed
gfp
green
fluorescence
protein
appl1
early
endosomes
figure
5c
importantly
appl1
bar-ph
domain
sufficient
target
rab5-q79l
containing
early
endosomes
figure
5c
contrast
one
rab5-binding
defective
mutants
a318d
failed
target
early
endosomes
exhibited
diffused
pattern
throughout
cytoplasm
cell
figure
5c
mutational
effects
appl1
targeting
rab5-positive
early
endosomes
cell
rfp
rab5-q79l
coexpressed
gfp
appl1
full
length
fl
bar-ph
domain
bar-ph
mutant
pc12
cells
indicated
followed
confocal
fluorescence
microscopy
shown
typical
confocal
microscopic
images
indicating
rfp
rab5-q79l
labeled
early
endosomes
red
colocalization
gfp
appl1
mutants
green
cells
scale
bartobacco
leaf
epidermal
cells
from
agroinfiltration
examined
under
zeiss
lsm
inverted
confocal
laser-scanning
microscope
zeiss
jena
germany
following
settings
described
goodin
et
al.
minor
modifications
single
optical
sections
scanned
resulting
images
each
transient
expression
each
plasmid
construct
cells
imaged
similar
results
gfp
fluorescence
excited
nm
filtered
primary
dichroic
uv/488/543
secondary
dichroic
nm
subsequently
bp505
nm
emission
filters
photomultiplier
tube
pmt
detector
dsred
fluorescence
excited
nm
emission
passed
similar
primary
secondary
dichroic
mirrors
finally
bp560
nm
emission
filter
pmt
detector
fluorescence
resonance
energy
transfer
fret
pairs
gfp/dsred
analysed
using
confocal
laser-scanning
microscope
zeiss
lsm510
meta
fret
measurements
dsred
emission
zero
contribution
from
gfp
accomplished
described
erickson
et
al.
using
following
settings
excitation
nm
emission
filters
bp
nm
gfp
bp
nm
dsred
following
agroinfiltration
tobacco
leaves
both
acbp4
dsred
gfp
atebp
observations
carried
out
confocal
microscopy
using
green
filter
investigate
fluorescence
pattern
gfp
red
filter
visualize
fluorescence
dsred
fig.
gfp
atebp
located
mainly
nucleus
signals
cytosol
plasma
membrane
fig.
2a
acbp4
dsred
localized
predominantly
cytosol
inclusive
signals
detected
cytosol
surrounding
nucleus
fig.
2b
signals
both
fusion
proteins
common
cytosol
confocal
images
indicating
co-localization
acbp4
dsred
gfp
atebp
fusion
proteins
transiently-expressed
tobacco
leaves
representative
tobacco
leaf
epidermal
cells
shown
laser-scanning
confocal
microscopy
following
agroinfiltration
plasmid
pat282
pat225