Zimmerliet al.,Science 374 , eabd9776 (2021) 10 December 2021 8 of 15
3456
40
50
60
70
80
Nuclear volume (m^3 )
N
PC di
amete
r(n
m
)
40 50 60 70 80 90
12
16
20
24
28
NPC diameter (nm)
INM-ONM distanc
e
(nm)
r = 0.4803
p = <0.0001
Nup60-
mCherry
control 30min ED 60min ED >150min ED
CD
ABcontrol osmotic shock
I
F
JK
G
E
H
N
N
N
ctr
l
ED OS
OSr
ec
ove
ry
0
20
40
60
NPCs/μm
2
ns
ns
ns
ns
ctr
l
ED OS
OS
rec
ove
ry
0
10
20
30
IN
M-ONM di
stance (nm)
ns
ctrl 30 60 >150
0
5
10
15
Energy depletion (min)
Nuclear volume (
m
3)
ctrl OS
0
5
10
15
Nuclear volume (
m
3)
Fig. 7. NE tension is reduced during ED and OS and leads to NPC constriction.
(A) Maximum projections of Nup60-mCherry channel from live-cell imaging
confocal stacks of NLS-GFP-NLS and Nup60-mCherry–expressing cells used for
nuclear shape and volume analysis. Scale bar, 5mm. (B) Same as (A) but for
cells expressing ubiquitous sfGFP and Nup60-mCherry under OS conditions.
(CandD) Quantification of nuclear volume based on 3D reconstructions of Nup60-
mCherry signal corresponding to (A) and (B). Data are from three experiments,
and triangles indicate means of individual replicates (means ± SDs;n= 300 nuclei
per condition under ED andn= 300 nuclei per condition under OS conditions).
One-way ANOVA and Dunnett’s multiple comparison test; ***P< 0.001;
**P< 0.01. (E) Number of NPCs per estimated NE surface area under control,
ED, OS, and OS recovery conditions (see Materials and methods). Data are from
one or more experiments (means ± SDs;n= 71 tomograms under control,
n= 72 tomograms under ED,n= 33 tomograms under OS, andn= 53 tomograms
under OS recovery conditions). One-way ANOVA andŠidák’s multiple comparison
test; ns indicates not significant,P> 0.05. (FandG) Deconvolved slice through
representative tomograms under control (F) and OS conditions (G). N marks
the nucleus, and arrowheads mark NPCs. Scale bars, 200 nm. (H) Mean NPC
diameter (see also Fig. 6A) plotted against mean nuclear volume [same as (C) and
(D)] under and control and 60 min after ED and OS conditions. Linear regression
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