A) Ear 21.2
ILC2s 23.7
98.1
Lineage
B)
iCD3𝜀
15.0% 26.0% 4.1% 46.5% 8.5%
ILC1s ILC2s ILC3s TrN Ungated
Total=4617.94
ILC3s 6.72
ROR𝛾t
GATA-3
GATA-3
IL-7R⍺
IL-7R⍺
ILC1s 19.5 TrN 39.5
T-bet
Lineage Gate: B220, CD11b, CD11c, CD5, CD3, CD19, Ter119, CD123, Gr1, F4/80, Fc𝜀R1 and CD49b
Supplementary Fig1
G)
CD3𝜀i
Lineage
60
Ear auLN
IL-7R⍺
IL-7R⍺
Id2+ 1.7
CD3𝜀i
Lineage
Id2-GFP
C) auLN Id2-GFP
IL-7R⍺
IL-7R⍺
IL-7R⍺
Id2+ 89.7
CD3𝜀i
Lineage
Id2-GFP Id2+ 1.6
IL-7R⍺
IL-7R⍺
WT
IL-7R⍺
D)
80
40
Id2-GFP
WT
IL-7R⍺
B)
IL-7R⍺
IL-7R⍺
IL-7R⍺
Id2+ 89.9
100
% of Id2GFP+ ILCs
A) Ear Id2-GFP
CD3𝜀i
Lineage
Id2-GFP
E) NK control Id2-GFP
CD45
NK1.1
IL-7R⍺
Id2+ 92.5
NKp46
CD3𝜀
Id2-GFP
F)
Id2+ 2.6
CD45
NK1.1
IL-7R⍺
WT
CD3𝜀
NKp46
Id2-GFP
Supplementary Fig2
Ear – Ear Digestion Protocol (EDP)
A)
CD45
Lineage
ILC3s 3.2
iCD3𝜀
GATA-3
IL-7R⍺
9.5
GATA-3
v
IL-7R⍺
Live/Dead
9.5
ILC2s 7.1 TrN 60.1 ILC1s 17.3
T-bet
ROR𝛾t
B) Lung – Lung Digestion Protocol (LDP) ILC2s 44.0
CD45
Lineage
ILC3s 3.8
iCD3𝜀
TrN 34.0
GATA-3
GATA-3
58.6
IL-7R⍺
IL-7R⍺
Live/Dead
1.1
ILC1s 9.6
T-bet
ROR𝛾t
Lung – Ear Digestion Protocol (EDP)
C)
ILC2s 20.5
Lung
80 70 60
10
7
6
EDP LDP
4 3 2 1
105
50
EDP LDP
Lung Number of ILCs
% of ILCs
80 60 40 20
15,324
104
ILC2
ILC3
TrN
GATA-3
82,535 37,712
EDP LDP 68,278
16,610
3,612
12,828 2,890 1,430
1,288
103 102
EDP LDP EDP LDP EDP LDP EDP LDP ILC1
10
5
Lung 105
100
0
10
6
104
0
EDP LDP
ILC1s 1.7
Lung
5
3,640,967
TrN 66.5
T-bet
ROR𝛾t Lung
E)
5,379,522
% of ILCs
Number of Live CD45+ cells
10
90
iCD3𝜀
Lung
100
% of Live CD45+ cells
F)
Lineage
ILC3s 3.9
Number of ILCs
CD45
D)
GATA-3
76.0
IL-7R⍺
IL-7R⍺
Live/Dead
1.4
EDP LDP EDP LDP EDP LDP EDP LDP ILC1
ILC2
ILC3
TrN
Supplementary Fig3
28.7
Lung -EDP 25.3
IL-7R⍺
61.7
Lung -LDP
B)
50
% CD3+ LiveCD45+
Ear -EDP
A)
Lung
40 30 20 10 0 EDP LDP
CD3𝜀 27.6
IL-7R⍺
32.4
% CD5+LiveCD45+
50
35.2
40 30 20 10 0 EDP LDP
CD5 0.8
IL-7R⍺
0.7
% CD49b+LiveCD45+
5
21.7
2 1 0
9.5
IL-7R⍺
11.2
% F4/80+LiveCD45+
50
18.8
40 30 20 10 0 EDP LDP
F4/80 3.8
IL-7R⍺
4.5
% Fc𝜀RI+LiveCD45+
30
32.5
20
10
0 EDP LDP
Fc𝜀RI 0.08
IL-7R⍺
11.1
% CD4+LiveCD45+
20
0.6
15 10 5 0 EDP LDP
CD4 9.1
0.02
% CD8+LiveCD45+
20
0.06
IL-7R⍺
3
EDP LDP
CD49b
CD8
4
15 10 5 0 EDP LDP
Supplementary Fig4
WT
60
**
40 20 0
ICOS
WT
ICOS
ILC1
9,378 7,664
104
103
WT
ICOS
% of ILC subsets
mLN
WT
ICOS
ILC2
WT
ICOS
WT
ILC3
80
105
C)
*
80
ICOS
105
***
202,975 172,408 102,446 25,893 70,660 24,285 9.411 10,202
104 103 102 WT
ICOS
ILC1
104
*
40
106
TrN
***
60
Number of ILC subsets
105
B)
Number of ILC subsets
145,012 127,878
% of ILC subsets
106
Number of ILCs
SI LP
Number of ILCs
A)
WT
ICOS
ILC2
WT
ICOS
WT
ILC3
ICOS TrN
*** 4,790 1,863 2,403
1,454
2,457 1,767
923 611
103
20
102
0 WT
ICOS
ILC1
WT
ICOS
ILC2
WT
ICOS
ILC3
WT
ICOS TrN
WT
ICOS
ILC1
WT
ICOS
ILC2
WT
ICOS
ILC3
WT
ICOS TrN
Supplementary Fig5
