a,b, Hypocotyl length measurements. WT
(n = 23/20, P < 0.001),
pif4 (n = 27/22, P
< 0.001), pif5 (n = 22/24,
P < 0.001), pif7
(n = 23/24, P < 0.001),
pif45 (n = 27/26, P
< 0.001), pif47 (n = 26/28,
P < 0.001), pif57
(n = 25/25, P < 0.001),
pif457 (n = 25/24, P =
0.275), pif1345 (n = 21/16, P
< 0.001) and pif13457 (n = 23/22,
P = 0.494) seedlings grown in WL or in response to low R:FR
(a) and WT (n = 22), phyB
(n = 22, P < 0.001), phyB
pif4 (n = 21, P < 0.001),
phyB pif5 (n = 26, P
< 0.001), phyB pif7 (n = 22,
P < 0.001), phyB pif45
(n = 20, P < 0.001), phyB
pif47 (n = 22, P < 0.001),
phyB pif57 (n = 26, P
< 0.001) and phyB pif457 (n = 27,
P = 0.218) seedlings grown in WL (b). Stars
denote statistically significant differences between light conditions
(a) or between WT and mutants (b) (two-way
analysis of variance (ANOVA) (a) or one-way ANOVA (b),
Tukey’s multiple comparisons test, n.s. (not significant)
P > 0.05, *P ≤ 0.05,
**P ≤ 0.01, ***P ≤ 0.001).
c, Immunoblot of pif457 PIF7:PIF7:4xMYC in LD.
Six-day-old seedlings continued to grow in WL or where exposed to low R:FR at
ZT0. Marks next to cropped blots represent 50 kDa (PIF7:4xMyC) or 37 kDa
(ACTIN), respectively. d, Immunoblot analysis of pif457
PIF7:PIF7:4xMYC and phyB pif457 PIF7:PIF7:4xMYC
seedlings (6-day-old, LD, WL). Marks next to cropped blots represent 50 kDa
(PIF7:4xMyC) or 37 kDa (ACTIN), respectively. e, Quantification of
percentages of nuclei with PIF7:4xMyc nuclear bodies as shown in f.
Five seedlings from each condition with at least 15 randomly selected nuclei per
seedling were examined. f, Subnuclear localization patterns of
PIF7:4xMyc and phyB in cotyledon epidermal nuclei of 6-day-old pif457
PIF7:PIF7:4xMYC and phyB pif457 PIF7:PIF7:4xMYC
seedlings grown in LD. Seedlings were fixed at ZT4 (WL) or after a subsequent 30
min low R:FR treatment. PIF7:4xMyC (red) and phyB (green) were labeled by
immunolocalization using anti-MyC and anti-phyB antibodies, respectively. Nuclei
were stained with DAPI (blue). The percentage of nuclei with the indicated
pattern, including s.e.m., is shown in merged images; n
indicates the number of nuclei analyzed. Scale bar, 5 μm. In
a, b and e, boxes extend from the
25th to 75th percentiles. Middle lines represent medians. Whiskers extend to the
smallest and largest values, respectively.