92 128
Nkx2.5-GFP FITC
64.9%
76.4%
81.8%
Day 5.3
WT
cnbp
KO
#1
cnbp
KO
#2
10 0 10 1 10 2 10 3 10 4 10 5
0
20
40
60
80
100
70.3%
90.9%
86.0%
cTnT APC
Day 10
WT
cnbp
KO
#1
cnbp
KO
#2
10 0 10 1 10 2 10 3 10 4 10 5
0
20
40
60
80
100
Percentage of Max
10 0 10 1 10 2 10 3 10 4 10 5
10 0
10 1
10 2
10 3
10 4
10 5
10 0 10 1 10 2 10 3 10 4 10 5
10 0
10 1
10 2
10 3
10 4
10 5
10 0 10 1 10 2 10 3 10 4 10 5
10 0
10 1
10 2
10 3
10 4
10 5
Flk1 APC
PdgfR
α
PE
cnbp
KO
#1
cnbp
KO
#2
WT
Day 4
13.4% 75.6%
3.93% 7.03%
4.16% 61.8%
10.2% 23.8%
5.43% 55.5%
11.8% 27.2%
A
Zn-1 RGG Zn-2 Zn-3
Zn-6
Zn-5
Zn-7
Zn-4
1
sgRNA-1
CNBP
: CCHC-type zinc finger,
C
ellular
N
ucleic acid
B
inding
P
rotein
170 AA
α
-CNBP
α
-GAPDH
#1 #2
cnbp
KO
WT
bvht
dAGIL
#1 #2
cnbp
KO
;
bvht
dAGIL
B
C
D
E
F
cTnT Myh6 Myh7
0.0
0.5
1.0
1.5
2.0
2.5
WT
cnbp
KO
#1
cnbp
KO
#2
**
**
***
Relative levels of transcript
Day 10
G
Relative levels of transcript
Gata4
Gata6
Hand1
Hand2
Nkx2.5
Mef2c
Tbx5
0
2
4
6
8
WT
cnbp
KO
#1
cnbp
KO
#2
Day 5.3
* *
**
***
****
*
H
63.5%
33.2%
cTnT APC
Percentage of Max
Nkx2.5-GFP FITC
Day 5.3
Day 10
10 0 10 1 10 2 10 3 10 4 10 5
0
20
40
60
80
100
10 0 10 1 10 2 10 3 10 4 10 5
0
20
40
60
80
100
63.9%
50.6%
WT;pEF_Empty
WT;pEF_CNBP-FLAG
WT;pEF_Empty
WT;pEF_CNBP-FLAG
10 0 10 1 10 2 10 3 10 4 10 5
10 0
10 1
10 2
10 3
10 4
10 5
10 0 10 1 10 2 10 3 10 4 10 5
10 0
10 1
10 2
10 3
10 4
10 5
PdgfR
α
PE
Flk1 APC
Day 4
WT;pEF_Empty
WT;pEF_CNBP-FLAG
4.65% 66.6%
8.59% 20.2%
9.87% 55.2%
7.85% 27.0%
Relative levels of transcript
Gata4
Gata6
Hand1
Hand2
Nkx2.5
Mef2c
Tbx5
0.0
0.5
1.0
1.5
Day 5.3
* *
** ****
*** ****
WT;pEF_Empty
WT;pEF_CNBP-FLAG
Relative levels of transcript
cTnT Myh6 Myh7
0.0
0.5
1.0
1.5
Day 10
** ** ****
WT;pEF_Empty
WT;pEF_CNBP-FLAG
I
CNBP-FLAG
CNBP
GAPDH
α
-CNBP
WT;pEF_Empty
WT;pEF_CNBP-FLAG
ESC
(legend on next page)
44
Molecular Cell
64
, 37–50, October 6, 2016