protein
immunoprecipitated
using
antibodies
against
brca1
antibody
against
flag
epitope
immunoprecipitate
flag
epitope
tagged
pp1α
brca1
coimmunoprecipitated
three
pp1
isoforms
conversely
pp1
coimmunoprecipitated
brca1
figure
indicating
interaction
between
brca1
pp1
specific293t
cells
transfected
ha-uxt
transfection
cells
treated
ng/ml
tnf-α
indicated
times
fractionated
cytoplasmic
nuclear
fractions
immunoprecipitated
immunoblotted
indicated
antibodies
respectively
293t
cells
treated
ng/ml
tnf-α
indicated
times
whole
cell
lysates
immunoprecipitated
immunoblotted
indicated
antibodies
bar
explore
uxt-binding
region
within
p65
constructed
series
p65
deletion
mutants
fig.
found
loss
amino
acids
terminus
p65
resulted
complete
inability
interact
uxt
fig.
top
contrast
p65
fragments
spanning
amino
acids
fully
retained
binding
capability
interacted
uxt
well
wild
type
fig.
middle
rhd
p65
consisted
two
ig-like
domains
chen
et
al.
made
two
additional
deletion
mutants
p65
amino
acids
each
contained
only
one
ig-like
domain
immunoprecipitation
assays
revealed
neither
able
interact
uxt
fig.
bottom
tagged
full-length
uxt
transfected
293t
cells
along
p65
deletion
mutants
indicated
whole
cell
lysates
immunoprecipitated
immunoblotted
indicated
antibodies
293t
cells
transfected
flag-uxt
together
ha-p50
myc-crel
ha
lymphoid
enhancer
binding
factor
cell
lysates
immunoprecipitated
immunoblotted
indicated
antibodiestherefore
whether
cop1
co
interact
vitro
tested
using
co-immunoprecipitation
assay
figure
cop1
attached
gal4
activation
domain
gad
cop1
co
made
vitro
transcription/translation
system
combined
gad
cop1
precipitated
anti-gad
antibody
co
co-precipitated
gad
cop1
figure
vitro
precipitation
experiments
demonstrated
coδb-box
co-immunoprecipitated
gad
cop1
whereas
coδcct
therefore
n-terminal
region
containing
b-boxes
required
interaction
cop1
suggesting
interaction
cop1
mediated
c-terminal
region
co
contains
cct
domain
cop1
directly
interacts
target
proteins
directs
degradation
hoecker
jiao
et
al
co
composed
three
domains
zinc-finger
b-boxes
central
domain
c-terminal
cct
domain
wenkel
et
al
co
cop1
interact
directly
vitro
demonstrated
immunoprecipitation
experiments
interaction
almost
abolished
c-terminal
part
co
removed
suggesting
cop1
interacts
c-terminal
region
co
previously
observed
interactions
between
cop1
col3
between
co
spa1
datta
et
al
laubinger
et
al
vitro
interaction
between
co
cop1
detected
co-immunoprecipitation
35s-methionine-labeled
co
coδb-box
coδcct
incubated
35s-methionine-labeled
gad
cop1
gad
co-immunoprecipitated
anti-gad
antibodies
supernatant
fractions
pellet
fractions
resolved
sds
page
visualized
autoradiography
using
phosphorimager
quantification
fractions
prey
proteins
co-immunoprecipitated
indicated
bait
proteins
gad
cop1
gad
error
bars
denote
standard
error
mean
two
replicate
experimentsimmunoprecipitation
experiments
using
tagged
tbk1
suggested
interaction
ddx3x
transcription
factor
irf3
significantly
weaker
interaction
between
tbk1
tank
therefore
detected
coimmunoprecipitation
under
stringent
conditions
supplementary
figure
1awild-type
mdc1
derivative
efficiently
co-immunoprecipitated
mrn
physiological
salt
concentrations
whereas
only
low
levels
mrn
recovered
immunoprecipitates
mdc1sdtdδ
mutant
fig
4b
consistent
sdtd
region
principal
mrn
interaction
interface
indicated
expression
constructs
transfected
human
embryonic
kidney
cells
extracts
prepared
immunoprecipitated
ip
gfp
antibodies
immunoblotted
immunoprecipitations
washes
performed
salt
inp
input
indicated
osteosarcoma
u2os
cell
lines
treated
two
rounds
control
cntl
mdc1-targeting
sirna
h.
cells
treated
gy
x-rays
processed
immunofluorescence
later
mdc1
nbs1
53bp1
antibodies
non-irradiated
cells
shown
supplementary
fig
s4b
onlineco-ip
experiments
repeated
using
hct116
sirt3
expressing
stable
cell
lines
foxo3a
found
interact
sirt3
both
whole
cell
fig.
1b
mitochondrial
extracts
fig.
1c
fractions
samples
co-ip
experiments
checked
presence
sirt3
using
sirt3
specific
antibody
biomol
plymouth
meeting
pa
well
tubulin
santa
cruz
biotechnology
santa
cruz
ca
cytochrome
mitosciences
eugene
ensure
fraction
purity
data
shown
experiments
demonstrate
both
wild
type
mutant
sirt3
form
physical
interaction
foxo3a
mitochondrial
extractscorroborate
interaction
from
yeast
two-hybrid
analysis
co-immunoprecipitation
studies
performed
according
mongiat
et
al.
constructs
used
interaction
assays
derivatives
vector
pbluescriptii
ks
pks
hindiii-saci
fragment
from
pbi-771
carrying
gal4
ta
amino
acids
cloned
corresponding
restriction
sites
pks
gal4
ta
acbp4
fusion
construct
prepared
inserting
acbp4
cdna
from
pat181
kb
ecori-bamhi
fragment
ecori-bglii
sites
pks-ta
ta-acbp4
adjacent
t3
promoter
co-immunoprecipitation
monoclonal
anti-gal4
ta
antibody
clontech
usa
performed
following
mongiat
et
al.
co-immunoprecipitation
acbp4
atebp
using
anti-gal4
ta
monoclonal
antibody
autoradiograph
sds-page
left
panel
showing
vitro
transcribed
translated
adf3
atebp
gal4
ta
acbp4
respectively
indicated
right
panel
shows
co-immunoprecipitation
equimolar
amounts
gal4
ta
acbp4
adf3
atebp
using
anti-gal4
ta
antibody
arrows
indicate
positions
proteins
co-immunoprecipitation
vitro
transcription/translation
products
gal4
ta
acbp4
fusion
protein
immobilized
protein
a/agarose
beads
using
monoclonal
antibody
against
gal4
ta
showed
gal4
ta
acbp4
fusion
protein
significantly
binds
atebp
fig.
1b
however
binding
gal4
ta
acbp4
adf3
observed
fig.
1b
perhaps
due
lack
cofactors
must
present
vitro
interaction
interaction
atebp
acbp4
substantiated
co-immunoprecipitation
using
autofluorescent
protein
fusions
transient
expression
tobacco
leaf
epidermal
cells
acbp4
atebp
showed
overlapping
expression
patterns
leaves
stems
both
inducible
acc
meja
treatment
infection
fungal
pathogen
botrytis
cinerea