Reduced mRNA recovery in spleen cells of MDSC-Exo treated AA mice. Figure S1. Proteome analysis of ex vivo and in vitro generated MDCS and MDSC- ...
Supplement
Myeloid-derived suppressor cell exosomes have significant immunoregulatory effects in autoimmune disease as exemplified in mouse alopecia areata 1
1
1
1
2
3
Margot Zöller , Kun Zhao , Natalia Kutlu , Natalie Bauer , Jan Provaznik , Thilo Hackert , Martina 4 Schnölzer 1 2 Tumor Cell Biology, University Hospital of Surgery, Heidelberg, Gene Core Unit, EMBL Heidelberg, 3 4 Pancreas Section, University Hospital of Surgery, Heidelberg, Functional Proteome Analysis, German Cancer Research Center, Heidelberg, Germany
Table S1 Table S2 Table S3 S3A S3B S3C Table S4 S4A S4B S4C Table S5 S5A S5B Table S6 Table S7 S7A S7B
Antibodies and reagents Immunogenic Ha1 and K6irs keratinocyte peptides Comparison between in vitro versus ex vivo generated MDSC Top score recovered peptides Higher peptide recovery in in vitro than ex vivo MDSC Higher peptide recovery in ex vivo than in vitro MDSC Protein profile of MDSC-Exo from in vitro / ex vivo MDSC Top score recovered peptides Higher peptide recovery in Exosomes from in vitro MDSC Higher peptide recovery in Exosomes from ex vivo MDSC Differences in MDSC versus MDCS-Exo protein profile Preferential peptide recovery in MDSC-Exo Higher peptide recovery in MDSC than MDSC-Exo MDSC-Exo peptides recovered in pulldown from Sepharose-coupled lymph node cell membranes In vivo impact of MDSC-Exo on AA spleen cells Increased mRNA recovery in spleen cells of MDSC-Exo treated AA mice Reduced mRNA recovery in spleen cells of MDSC-Exo treated AA mice
Figure S1 Figure S2 Figure S3 Figure S4
Proteome analysis of ex vivo and in vitro generated MDCS and MDSC-exosomes Proteome analysis comparing ex vivo and in vitro generated MDCS with MDSC-exosomes MDSC-exosome membrane attached proteins Examples of hair regrowth in MDSC exosome-treated mice with AA partialis and incipient
1
Table S1 Antibodies and reagents Antibody specificity Akt (p)Akt Alix BAD pBAD BclXl Caspase 3 activ Caspase 8 Caspase9-clv. CCR3 (CD193) CCR4 (CD194) CCR5 (CD195) CCR6 (CD196) CCR7 (CD197) CCR8 (CDw198) CCR9 (CDw199) CD3 CD4 CD8 CD9 CD11b (ITM) (YBM6.6.10) CD11c (ItgX) CD14 CD16 (FcRIIIb) CD16/32 (NK) CD19 CD25 (PC61) CD28 CD32 (2.4G2) CD40 CD47 (IAP) CD54 (ICAM1) CD63 (Tspan30) CD69 CD80 CD81 (Tspan28) CD86 CD103 (ITE) CD115 (M-CSFR) CD117 (c-KIT) CD135 (FLT3) CD152 CD154 CD314 (NKG2D) CXCR3 (CD183) CXCR4 (CD184) Erk1/2 ERK1/2-p F4/80 FoxP3 GM-CSF Gr1 HIF1 IFN IB IB-p IgM IL1 IL4 IL6 IL10 IL12 JAK2 JAK2-p JAK3 JAK3-p JNK pJNK Jun Jun-p Ly6C Ly6G
donor mouse mouse rabbit hamster rabbit rabbit rabbit rabbit rabbit rat goat rat rat rat goat rat rabbit rat rat rat rat hamster rat rabbit rat rat rat monocl.hamster rat rat rat monocl.hamster rabbit monocl.hamster rat monocl.hamster rat rat rabbit rat rat monocl.hamster monocl.hamster rat rat/rabbit rat mouse mouse rat rat rat rat mouse rat rabbit mouse rabbit rat rat rat rat rat rabbit rabbit mouse goat rabbit mouse rabbit rabbit rat rat
supplier Becton Dickinson, HD, G Becton Dickinson, HD, G Santa Cruz, HD, G Becton Dickinson, HD, G Cell Signaling, Frankfurt, G Cell Signaling, Frankfurt, G Becton Dickinson, HD, G Becton Dickinson, HD, G Cell Signaling, Frankfurt, G R&D Systems, G abcam, Cambridge, UK Becton Dickinson, HD, G R&D Systems, G R&D Systems, G abcam, Cambridge, UK R&D Systems, G Abcam, Cambridge, UK EACCb EACCb Becton Dickinson, HD, G EACCb Becton Dickinson, HD, G Becton Dickinson, HD, G abcam, Cambridge, UK Becton Dickinson, HD, G ImmunoTools, Friesoythe, G EACCb Becton Dickinson, HD, G ATCCc Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Santa Cruz, HD, G ImmunoTools, Friesoythe, G Becton Dickinson, HD, G ImmunoTools, Friesoythe, G Becton Dickinson, HD, G Becton Dickinson, HD, G abcam, Cambridge, UK Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Santa Cruz, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G ImmunoTools, Friesoythe, G Novus Biologicals, Littletown, USA Becton Dickinson, HD, G Santa Cruz, HD, G Becton Dickinson, HD, G Dianova, Hamburg, G BioTrend, Cologne, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G SantaCruz, HD, G SantaCruz, HD, G Becton Dickinson, HD, G SantaCruz, HD, G SantaCruz, HD, G Becton Dickinson, HD, G SantaCruz, HD, G SantaCruz, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G
2
Table S1 continued Antibody specificity MHCI (K7-65) MHCII (K25-137) Myc-p MyD88 NFB NK (PK136) NOS2 P38 P38-p PI3K pPI3k Rab5 SCA1 (Ly6A) SCF (KIT ligand) Shc Shc-p Src Src-p STAT3-p STAT4-p STAT6-p Survivin Ter119 TGF1 TLR2 (CD282) TLR3 TLR4 (CD284) TNF TSG101 ZAP70 ZAP70-p dye or biotin labeled secondary antibodies /Streptavidin
donor mouse mouse rabbit rabbit rabbit rat rabbit rabbit rabbit rabbit rabbit mouse rat rabbit rabbit rabbit rabbit rabbit rabbit mouse rabbit rabbit rat rat rat rabbit rat rat rabbit rabbit rabbit
supplier Ref 1d Ref 1d Santa Cruz, HD, G Santa Cruz, HD, G Cell Signaling, Franfurt, G ATCC SantaCruz, HD, G SantaCruz, HD, G SantaCruz, HD, G Cell Signaling, Frankfurt, G Cell Signaling, Frankfurt, G Becton Dickinson, HD, G Becton Dickinson, HD, G abcam, Cambridge, UK Cell Signaling, Franfurt, G Cell Signaling, Franfurt, G Cell Signaling, Franfurt, G Cell Signaling, Franfurt, G Cell Signaling, Franfurt, G Becton Dickinson, HD, G SantaCruz, HD, G SantaCruz, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G SantaCruz, HD, G Becton Dickinson, HD, G Becton Dickinson, HD, G SantaCruz, HD, G Cell Signaling, Frankfurt, G Cell Signaling, Frankfurt, G
Reagent AnnV-FITC / APC CFSE (carboxyfluoresceinsuccinimidylester) GM-CSF IL4 IL6 PGE2 PI Sulfobiotin-X-NHS
Dose 0.2µl-0.5µl
Supplier Becton Dickinson, HD, G
5µM (10-20ng/ml 2ng/ml 5ng/ml 1nM 0.08µg 1mg/ml
Invitrogen, Darmstadt, G ImmunoTools, Friesoythe, G ImmunoTools, Friesoythe, G ImmunoTools, Friesoythe, G BioTrend, Cologne, G Becton Dickinson, HD, G Calbiochem, Darmstadt, G
Dianova, Becton Dickinson, Amersham
a
all rat and mouse antibodies are monoclonal, monoclonal hamster antibodies are marked EACC: European Collection of Animal Cell Cultures ATCC: American Type Cell Culture Collection d Reference 1. Koch N, Hämmerling GJ, Tada N, Kimura S, Hämmerling U. Cross-blocking studies with monoclonal antibodies against IA molecules of haplotypes b, d and k. Eur J Immunol (1982) 12: 909-14. b c
3
Table S2 Immunogenic Ha1 and K6irs keratinocyte peptides MHCI
MHCII
Name-Position
Sequenz
Name-Position
Sequenz
Ha1(K31)-147
TELGLRQLV
Ha1-204
QLGDRLNVEVDAAPT
Ha1(K31)-211
VEVDAAPTV
Ha1-223
RVLNETRCQYEAMVE
Ha1-157
SDINGLRRI
Ha1-249
TQTEELNKQVVSSSE
Ha1-251
TEELNKQVV
Ha1-277
RTVNALEIELQAQHC
K6irs(K71)-205
SELRNVRDV
K6irs-296
LDLDSIIDEVRAQYE
K6irs-178
LEPILEGHI
K6irs-118
VELDPEIQKVRAQER
K6irs-193
LETLSGDRV
K6irs-382
QRGDSALKDARAKLD
K6irs-242
VDAAYANKV
K6irs-289
SMDNNRNLDLDSIID
4
Table S3 Comparison between in vitro versus ex vivo generated MDSC Table S3A Highest score recovered proteins in ex vivo and in vitro generated MDSC MDSC in vitro
MDSC ex vivo
Scorea
Prot.M.
Pept.M.
Scorea
Aconitate hydratase, mitochondrial
1252
78
20
1315
119
32
Actb
Actin, cytoplasmic 1
1972
1165
47
1995
1063
48
Actn1
actinin alpha 1
2631
141
35
2453
116
31
Actn4
actinin alpha 4
2575
128
36
2151
96
29
Aldh2
aldehyde dehydrogenase 2 family
1551
262
29
1555
245
33
Aldoa
aldolase, fructose-bisphosphate A
1401
182
18
1206
131
14
Anxa1
Annexin A1
2043
134
24
1963
134
24
Anxa2
Annexin A2
1679
180
21
1359
160
17
Anxa3
Annexin A3
1474
43
18
869
24
14
Anxa4
Annexin A4
1464
50
17
1018
29
12
Anxa5
Annexin A5
1574
44
20
963
31
13
Anxa6
Annexin A6
1727
80
18
1215
38
15
Atp1a1
ATPase Na+/K+ transporting subunit alpha 1
1058
49
11
1314
105
18
Atp2a2
ATPase Ca2+ transporting 2
1234
72
15
2257
155
26
Atp5a1
ATP synthase, H+ transporting, complex alpha 1
2090
331
25
2025
321
25
Atp5b
ATP synthase, H+ transporting complex, beta 5
2222
296
25
2205
301
25
Atp6v1a
ATPase H+ transporting V1 subunit A
1115
52
16
1807
104
24
Calr
Calreticulin
1533
159
22
1460
143
21
Canx
Calnexin
1538
87
19
1581
121
22
Cap1
Adenylyl cyclase-associated protein 1
1317
170
17
1179
76
16
Capn1
Calpain-1
1186
29
18
759
21
11
Cat
Catalase
1101
30
15
1347
58
21
CD11b
Integrin alpha-M
2995
244
38
2569
268
33
CD13
Alanyl aminopeptidase, membrane
1972
65
24
1874
67
23
CD177
CD177 antigen Cd177
1019
39
13
1283
43
18
CD18
Integrin beta-2
3098
178
39
3099
259
37
CD206
mannose receptor C-type 1
2224
48
27
1580
41
20
Ckap4
Cytoskeleton-associated protein 4
1713
54
20
1668
60
21
Cltc
Clathrin heavy chain 1
3779
99
49
3425
107
42
Cndp2
CNDP dipeptidase 2
1218
77
16
857
40
13
Copa
coatomer protein complex subunit alpha
1159
23
15
1033
34
15
Ctsb
Cathepsin B
1139
66
14
746
25
10
Cybb
Cytochrome b-245 beta chain
787
50
13
1053
130
14
Dync1h1
dynein cytoplasmic 1 heavy chain 1
2332
56
36
3537
89
53
Eef2
Elongation factor 2
2630
269
35
1538
167
21
Ehd4
EH domain-containing 4
884
45
12
1276
57
18
Eif4a1
Eukaryotic initiation factor 4A1
1254
135
15
1141
117
14
Eno1
enolase 1
2380
373
27
2238
280
25
Esyt1
Extended synaptotagmin-1
2753
217
37
3012
261
44
Ezr
Ezrin
1565
123
21
1370
91
17
Fasn
Fatty acid synthase
1018
25
15
1200
22
16
Fermt3
Fermitin family member 3
1519
111
21
1160
126
16
Flna
Filamin-A
5389
391
71
4448
314
54
Ganab
glucosidase II alpha subunit
1962
113
26
2094
115
29
Gapdh
Glyceraldehyde-3-phosphate dehydrogenase
1357
401
13
1158
243
11
Gdi2
GDP dissociation inhibitor 2
1335
92
15
1147
65
16
Glg1
Golgi apparatus protein 1
920
32
10
1423
50
18
Glud1
Glutamate dehydrogenase 1
1589
77
21
1793
131
26
Gnai2
G protein alpha inhibiting 2
1310
140
15
1340
221
17
Got2
Aspartate aminotransferase, mitochondrial
1177
93
19
1064
80
18
Gpi
Glucose-6-phosphate isomerase
1495
113
21
1125
67
15
Synonym
Protein Description
Aco2
5
Prot.M. Pept.M.
Table S3A cont MDSC in vitro Synonym
Protein Description
MDSC ex vivo
Scorea
Prot.M.
Pept.M.
Scorea
48
15
1044
50
12
Prot.M. Pept.M.
Gsn
Gelsolin
1257
Hbb-b1
hemoglobin, beta adult major chain
1295
503
14
960
184
12
Hbb-b2
hemoglobin, beta adult minor chain
1130
299
14
854
117
12
Hnrnpa2b1
Heterogeneous nuclear ribonucleoproteins A2/B1
1286
82
17
479
10
5
Hsd17b4
hydroxysteroid 17-beta dehydrogenase 4
464
9
5
1082
44
11
Hsp90aa1
Heat shock protein HSP 90-alpha
1900
255
27
1289
173
18
Hsp90ab1
Heat shock protein HSP 90-beta
2507
419
33
1973
309
27
Hsp90b1
heat shock protein 90, beta (Grp94), member 1
2758
457
35
3265
525
42
Hspa5
heat shock protein 5
3362
544
38
3586
610
38
Hspa8
Heat shock cognate 71 kDa protein
3021
436
38
2745
330
35
Hspd1
heat shock protein family D (Hsp60) member 1
1943
69
24
861
44
10
Hyou1
Hypoxia up-regulated protein 1
2463
169
30
2426
161
31
Idh2
Isocitrate dehydrogenase [NADP] 2
920
63
14
1081
77
18
Immt
inner membrane mitochondrial protein
435
7
5
1078
44
14
Iqgap1
IQ motif containing GTPase activating protein 1
3437
233
43
2981
239
36
Lcp1
lymphocyte cytosolic protein 1
3082
486
38
2690
400
37
LDHA
L-lactate dehydrogenase A chain
1329
147
16
1064
129
16
Lmna
Lamin-A/C Lmna
1994
102
25
Lmnb1
Lamin-B1 Lmnb1
693
21
10
2297
131
27
Lnpep
Leucyl-cystinyl aminopeptidase
1122
37
15
1132
34
15
Ltf
Lactotransferrin
3279
223
41
2903
192
37
Mdh2
Malate dehydrogenase 2
1365
114
16
1281
88
16
Mpo
Myeloperoxidase
1761
149
21
1271
68
17
Msn
Moesin
2728
221
36
2573
158
34
Mvp
Major vault protein
1750
45
24
1791
55
25
Myh9
myosin heavy chain 9
5150
207
69
263
3
2
Myof
Myoferlin
191
4
3
3636
186
52
Napa
NSF attachment protein alpha
655
16
8
1066
31
14
Ncl
Nucleolin
1729
87
23
816
51
12
Oat
Ornithine aminotransferase
711
16
9
1119
48
16
Ogdh
oxoglutarate dehydrogenase
1079
38
15
1201
68
20
P4hb
prolyl 4-hydroxylase subunit beta
2412
286
31
2351
312
31
Pdia3
Protein disulfide-isomerase A3
2747
263
37
2655
335
39
Pdia4
Protein disulfide-isomerase A4
2505
182
32
2519
215
35
Pdia6
Protein disulfide-isomerase A6
1286
113
15
1358
155
14
Pgk1
Phosphoglycerate kinase 1
1289
107
13
960
69
13
Pgls
6-phosphogluconolactonase
1191
156
15
1018
113
13
Pkm
Pyruvate kinase PKM
2580
403
31
2108
298
27
Plek
Pleckstrin
352
14
5
1959
53
25
Por
cytochrome P450 oxidoreductase
1104
67
14
1248
92
15
Ppib
Peptidyl-prolylisomerase B
1043
55
13
1130
60
14
Ptpn6
protein tyrosine phosphatase, non-receptor type 6
1348
110
17
1182
106
15
Ptprc
protein tyrosine phosphatase, receptor type C
1229
85
17
1247
113
20
Pygl
Glycogen phosphorylase L
1534
106
22
838
41
13
Rab7a
RAB7A, member RAS oncogene family
1229
149
15
1009
88
13
Rack1
Receptor of activated protein C kinase 1
1145
81
16
762
53
12
Rdx
Radixin
1183
108
17
1075
80
14
Rnh1
ribonuclease/angiogenin inhibitor 1
1332
52
14
1035
50
12
Rpn1
ribophorin I
1331
169
21
1783
250
23
Rpn2
ribophorin II
1212
95
15
1371
150
17
Rrbp1
Ribosome-binding protein 1
1444
46
19
1724
54
24
Sdha
succinate dehydrogenase complex subunit A
497
19
6
1089
66
13
Serpinb1a
Leukocyte elastase inhibitor A
1377
156
19
913
64
12
6
Table S3A cont. MDSC in vitro Synonym
Scorea
Protein Description
Prot.M.
MDSC ex vivo
Pept.M.
Scorea
89
17
Prot.M. Pept.M.
634
40
10
1297
solute carrier family 25 member 4
868
109
13
1169
171
16
solute carrier family 25 member 5
1193
204
15
1328
280
18
Snd1
Staphylococcal nuclease and tudor domain-1
1459
51
21
483
17
5
Sun2
SUN domain-containing protein 2
1682
125
20
1539
107
19
Thbs1
Thrombospondin-1
1206
64
16
268
10
3
Tkt
Transketolase
1676
163
20
1625
113
20
Tln1
Talin-1
4912
297
57
5199
342
61
Tuba1b
Tubulin alpha-1B
1440
238
19
1360
157
18
Tuba1c
Tubulin alpha-1C
1357
222
18
1295
137
17
Tuba4a
Tubulin alpha-4A
1352
189
18
1189
122
16
Tubb2a
Tubulin beta-2A
1173
166
15
Tubb4b
Tubulin beta-4B
1454
203
17
1332
160
17
Tubb5
Tubulin beta-5
1628
247
20
1459
179
19
Uba1
Ubiquitin-like modifier-activating enzyme 1
2006
101
23
1243
67
14
Uggt1
UDP-glucose glycoprotein glucosyltransferase 1
3039
116
39
2906
124
38
Vcp
valosin containing protein
2350
132
29
1970
173
24
Vdac1
Voltage-dependent anion channel 1
1431
64
15
1545
73
16
Vim
Vimentin
3056
371
42
2495
153
37
Slc25a12
solute carrier family 25 member 12
Slc25a4 Slc25a5
a
Score ≥1000: bold, ≥2-fold difference in significant peptide hits: italics
7
Table S3B Peptides recovered at a significantly higher frequency in in vitro than ex vivo MDSC
Synonym Ap1b1 Ap1m1 Ap2b1 Arg1 Arsb Ca1 Ca2 Cbr2 CD41 CD61 Celf2 Chp1 Cmpk2 Cpd Ddx39a Ddx3x Elavl1 Ethe1 Fam3c Glo1 Gmfg Grb2 Gstm1 Gsto1 Hadh Hck Hdac1 Hnrnpa1 Hnrnpa2b1 Hnrnph1 Hnrnpl Hprt1 Hsd17b10 Ifi44 Ifit1
MDSC in vitro MDSC ex vivo Prot.M. Pept.M. Prot.M. Pept.M. 33 12 17 2 5 4 23 6 35 10 5 3 6 3 21 7 6 2 5 4 23 12 14 6 10 5 5 3 37 12 6 3 80 14 4 4 17 7 13 6 6 3 10 4 12 6 1 1 13 6 2 1 13 7 1 1 26 12 1 1 13 6 7 4 12 4 9 6 95 15 2 1 82 17 10 5 67 13 10 4 31 11 5 3 9 5 6 3 27 9 40 12
Protein Description AP-1 complex subunit beta-1 AP-1 complex subunit mu-1 AP-2 complex subunit beta Arginase-1 Arylsulfatase B Carbonic anhydrase 1 Carbonic anhydrase 2 Carbonyl reductase 2 Integrin alpha-IIb Integrin beta-3 CUGBP Elav-like family member 2 calcineurin like EF-hand protein 1 cytidine/uridine monophosph kinase 2 Carboxypeptidase D DEAD-box helicase 39A DEAD-box helicase 3, X-linked ELAV like RNA binding protein 1 ETHE1, persulfide dioxygenase family with similarity 3 member C glyoxalase I Glia maturation factor gamma Growth factor receptor-bound prot. 2 Glutathione S-transferase Mu 1 Glutathione S-transferase omega-1 hydroxyacyl-CoA dehydrogenase HCK protooncogene, Src family TK Histone deacetylase 1 Heterogeneous ribonucleoprotein A1 Heterogeneous ribonucleoprot. A2/B1 Heterogeneous ribonucleoprotein H1 Heterogeneous ribonucleoprotein L Hypoxanthine phosphoribosyltransf. 1 hydroxysteroid 17- dehydrogenase 10 IFN-induced protein 44 IFN-induc. prot. w tetratricopep.rep.1
Protein Family membrane traffic. ECM receptor membrane traffic., prot. transporter cysteine protease Hydrolase dehyratase dehyratase dehydrogenase Adhesion Adhesion nucleic acid & RNA binding phosphatase nucleotide kinase metalloprotease RNA helicase GTPase activity protein & RNA & nucleic acid binding hydrolase lyase signaling non-receptor tyrosine kinase transferase cytoskel., transferase, elong. factor dehydrogenase tyrosine kinase reductase RNA-, nucleic acid binding RNA-, miRNA binding protein & RNA & nucleic acid binding protein, mRNA binding glycoyltransferase dehydrogenase, reductase protein & RNA binding
8
Reactome pathway protein transporter & binding vesicle biogenesis & transport vesicle med. transport, endocyt., signaling scavenger receptor metabolism metabolism reductase, protein binding adhesion and signaling adhesion and signaling metabolism, hyaluronan degradation vesicle biogenesis mRNA processing protein-, RNA-, DNA-binding splicing ion binding platelet activation & signaling citric acid cycle, electron transport Arp2/3 complex binding multiple signaling pathways metabolism metabolism protein kinase, transferase protein binding, transcription factor activity signaling splicing and signaling splicing metabolism metabolism molecular function
Table S3B cont. Synonym Ifit2 Ifit3 Ilf2 Iqgap2 Irgm1 Isg15 Kpnb1 Mcm4 Mcm7 Mcpt8 Mrps34 Msra Mta2 Mybbp1a Myh9 Myl12b Nipsnap3b Npc2 Nsf Nudt21 Ostf1 Park7 Pcmt1 Pddc1 Pds5a Pf4 Pfdn2 Ppil1 Ppp1ca Ppp1cb Ppp1cc Ppp2ca Ppp2r1a Prdx6 Psma2 Psma3 Psma5
MDSC in vitro MDSC ex vivo Prot.M. Pept.M. Prot.M. Pept.M. 19 6 20 4 5 3 4 3 7 3 52 4 55 13 20 4 7 4 13 3 11 4 5 4 5 3 10 6 29 12 2 1 207 69 3 2 27 8 8 4 7 4 0 0 7 4 13 7 9 6 9 4 5 3 8 3 6 3 4 3 6 4 1 1 4 3 36 13 7 4 19 9 25 11 12 7 1 1 15 7 3 2 30 13 14 8 1 1 14 6 1 1 19 8
Protein Description IFN-induc. prot. w tetratricopep. rep. 2 IFN-induc. prot. W tetratricopep. rep. 3 Interleukin enhancer-binding factor 2 IQ motif contain. GTPase active. prot.2 Immunity-relat. GTPase family M prot.1 ISG15 ubiquitin-like modifier karyopherin subunit beta 1 minichromosome mainten. complex 4 minichromosome mainten. complex 7 Mast cell protease 8 Mcpt8 mitochondrial ribosomal protein S34 Methionine sulfoxide reductase 1 metastasis assoc. 1 family member 2 Myb-binding protein 1A myosin heavy chain 9 myosin light chain 12B NipSnap homolog 3B NPC intracell. cholesterol transporter 2 Vesicle fusing ATPase nudix hydrolase 21 Osteoclast-stimulating factor 1 Parkinsonism assoc. deglycase, DJ-1 protein-L-isoaspartate methyltransfer. Parkinson disease 7 domain-cont. 1 PDS5 cohesin associated factor A Platelet factor 4 Prefoldin subunit 2 peptidylprolyl isomerase like 1 protein phosphatase 1 subunit alpha protein phosphatase 1 subunit beta protein phosphatase 1 subunit gamma protein phosphatase 2 subunit alpha Protein phosphatase 2A subunit A Peroxiredoxin-6 Proteasome subunit alpha 2 Proteasome subunit alpha 3 Proteasome subunit alpha 5
Protein Family
kinase activator G-protein modulator hydrolase protein binding transfer / carrier protein DNA helicase, hydrolase hydrolase, DNA helicase serine protease reductase chromatin binding transcription factor G-protein modul., actin binding binding cholesterol binding hydrolase hydrolase SH3 domain binding protease, transcription methyltransferase RNA binding, transcription factor chromatin binding chemokine chaperone isomerase phosphatase phosphatase phosphatase phosphatase phosphatase peroxidase endopeptidase peptidase, hydrolase peptidase, hydrolase
9
Reactome pathway RNA binding protein binding Rho GTPase signaling cytokine signaling apoptosis execution cell cycle cell cycle proteolysis protein synthesis, ribosome constituent metabolism DNA & protein binding protein & RNA-binding protein binding Rho GTPase signaling vesicular trafficking cholesterol transport COPI-,COPII-mediated vesicle transport mRNA splicing bone resorption scaffold protein binding
cell cycle CXCR3 signaling protein folding splicing GPCR signailng Rho GTPase signaling, cell cycle protein binding, phosphatase activity TLR4 cascade, MAPK signaling, TLR & multiple signaling peroxidase, hydrolase, binding multiple signaling pathways signaling signaling
Table S3B cont. Synonym Psma6 Psma7 Psmb10 Psmb3 Psmb4 Psmb8 Psmb9 Psme1 Ptbp3 S100a13 Sept7 Sept9 Set Sf3a1 Sf3b3 Siglec1 Skap2 Smarca5 Snd1 Snrpa Snrpa1 Snx5 Sptan1 Srsf10 Ssb Ssr4 Stip1 Tfrc Tgm2 Thbs1 Tln2 Tmed4 Tpr Tpt1 Trex1 Tubb2a U2af2
MDSC in vitro MDSC ex vivo Prot.M. Pept.M. Prot.M. Pept.M. 18 8 3 2 17 10 6 3 11 5 20 7 14 6 15 5 3 1 38 11 4 3 19 8 9 3 5 3 15 3 9 4 16 4 6 4 1 1 32 9 6 5 0 0 5 3 12 5 51 21 17 5 8 3 10 5 13 4 1 1 7 5 4 3 14 7 2 2 29 5 13 6 30 9 21 6 64 16 10 3 26 6 12 5 2 1 14 9 30 4 14 5 166 15 9 5
Protein Description Proteasome subunit alpha 6 Proteasome subunit alpha 7 Proteasome subunit beta 10 Proteasome subunit beta 3 Proteasome subunit beta 4 Proteasome subunit beta 8 Proteasome subunit beta 9 proteasome activator subunit 1 Polypyrimidine tract-binding protein 3 S100 calcium binding protein A13 Septin-7 Septin-9 SET nuclear proto-oncogene Splicing factor 3A subunit 1 Splicing factor 3B subunit 3 Sialoadhesin Src kinase-assoc. phosphoprotein 2 regulator of chromatin, subfamily a5 Staphyloc. nuclease & tudor dom.1 small nuclear ribonucleoprotein A small nuclear ribonucleoprotein A' Sorting nexin-5 Spectrin alpha, non-erythrocytic 1 Serine/arginine-rich splicing factor 10 Sjogren syndrome antigen B Stomatostatin receptor type 4 Stress-induced-phosphoprotein 1 Transferrin receptor transglutaminase 2 Thrombospondin-1 Talin-2 transmembrane p24 trafficking prot. 4 translocated promoter tumor protein, translation.-control. 1 Three-prime repair exonuclease 1 Tubulin beta-2A U2 snRnP auxiliary factor 2
Protein Family peptidase, hydrolase endopeptidase endopeptidase peptidase peptidase endopeptidase endopeptidase endopeptidase NA & RNA binding calmodulin cytoskeleton cytoskeleton chaperon, phosphatase inhibitor RNA & protein binding RNA & protein binding Ig superfam. adhesion, signaling protein binding DNA helicase transcription cofactor RNA binding RNA bindingr trafficking regulator non-motor actin binding RNA & nucleic acid binding tRNA / RNA binding GPCR chaperone receptor acyltransferase integrin-, fibrinogen-, collagen V-bind. actin family (cytoskeleton) transfer protein, vesicle coat RNA-, protein-binding microtubule binding nuclease, hydrolase, protein binding GTP binding RNA & protein binding
10
Reactome pathway signaling multiple signaling pathways multiple signaling pathways
GPCR signaling multiple signailng pathways regulator of differentiation binding & signaling binding binding cell cycle cell cycle splicing factor protein & carbohydrate binding signal regulator biological processes protein, NA, RNA binding splicing factor splicing factor vesicle biogenesis, -mediated transport multiple signaling splicing factor splicing factor signal transduction axon guidance clathrin-mediated endocytosis apoptosis platelet aggreg., aniogenesis, inflamm., matrix adhesion actin filament assembly signal transduction SLC-mediated transmembrane transport regulator of growth and differentiation mismatch repair membrane trafficking, vesicle-mediated transport splicing factor
Table S3B cont. Synonym Trex1 Tubb2a U2af2 U2surp Vps29 Xpo2
MDSC in vitro MDSC ex vivo Prot.M. Pept.M. Prot.M. Pept.M. 14 5 166 15 9 5 4 3 4 3 9 4
Protein Description Three-prime repair exonuclease 1 Tubulin beta-2A U2 snRnP auxiliary factor 2 U2 snRNP-associated SURP motif vacuolar protein sorting 29 homolog Exportin-2
Protein Family GTP binding splicing factor mRNA processing vesicle coat GTPase, transporter
Reactome pathway nuclease, hydrolase, protein binding membrane trafficking, vesicle-mediated transport protein-, RNA-binding splicing protein & ion binding protein binding, Ran GTPase binding
Table S3C
Proteins recovered at a significantly higher level in ex vivo than in vitro MDSC Synonym Atp6ap2 Atp6v0a1 Capzb Casp6 Chmp2a Cpt2 D1Pas1 Dnajc10 Emc1 Erlin2 Fundc2 Galnt7 Gas7 Glrx Igf2r Lmna Lmnb2 Mmp12 Mrpl41 Mrpl49 Mtch2 Myof Nono Nup155 Nup210 Olfm4
MDSC in vitro MDSC ex vivo Prot.M. Pept.M. Prot.M. Pept.M. 9 5 23 8 17 4 6 4 5 3 1 1 10 7 8 4 2 1 13 5 16 8 15 6 7 4 1 1 7 6 6 5 8 4 5 3 102 25 17 3 1 1 18 6 7 4 3 3 16 5 4 3 186 52 6 3 8 5 3 1 45 12 8 5
Protein Description ATPase H+ transporting 2, Renin receptor ATPase, H+ transporting, V0 subunit A1 capping protein muscle Z-line, beta Caspase-6 Charged multivesicular body protein 2a Carnitine O-palmitoyltransferase 2 DNA segment, Chr 1, Pasteur Institute 1 DnaJ Hsp family (Hsp40) member C10 ER membrane protein complex subunit 1 ER lipid raft associated 2 FUN14 domain-containing 2 N-acetylgalactosaminyltransferase 7 Growth arrest-specific 7 Glutaredoxin insulin-like growth factor 2 receptor Lamin-A/C Lmna Lamin-B2 matrix metallopeptidase 12 mitochondrial ribosomal protein L41 mitochondrial ribosomal protein L49 Mitochondrial carrier 2 Myoferlin Non-POU domain-con. octamer-binding nucleoporin 155 nucleoporin 210 Olfactomedin-4
Protein Family ATP synthase, hydrolase non-motor actin binding protease, protease inhibitor . transferase RNA helicase . . . . transferase actin family oxidoreductase receptor intermediate filament intermediate filament . ribosomal protein ribosomal protein transfer / carrier protein membrane traffic mRNA splicing . membrane traffic protein receptor, structural protein
11
Reactome pathway angiotensin metabolism InsR signal., ROS, transmembr. & ion transp. b-tubulin-, actin-, protein-binding apoptotic execution phase vesicle biogenesis, ECSRT, traffic, budding lipid / lipidprotein metabolism ATP-, RNA-, DNA-binding chaperone binding, Hsp70 binding molecular function transmembrane transport molecular function glycosylation membrane trafficking regulator electron carrier, nucleotide metabolism clathrin endocytos., memb. traffic, vesicle biogen. apoptotic execution, cell cycle structural molecule ECM organization, ECM degradation mitochondrial biogenesis and maintenance mitochondrial biogenesis and maintenance molecular function phospholipid binding protein-, RNA-, DNA-binding heat stress regulation, SLC-mediated transport heat stress response, transmembrane transport protein binding
Table S3C cont. MDSC in vitro MDSC ex vivo Synonym Prot.M. Pept.M. Prot.M. Pept.M. Parp1 4 3 Pdha1 13 6 Pfkp 2 1 13 6 Ptbp1 18 5 Samm50 6 3 Sel1l 7 4 23 13 Slc25a13 47 8 Snap29 5 3 Snx12 4 3 Ssr2 0 0 46 4 Timm50 6 4 Tomm40 14 5 Trpv2 2 1 18 5 Uqcrc1 7 2 46 7
Protein Description poly (ADP-ribose) polymerase family 1 pyruvate dehydrogenase alpha 1 phosphofructokinase, platelet Polypyrimidine tract-binding protein 1 SAMM50 sorting and assembly machinery SEL1L ERAD E3 ligase adaptor subunit solute carrier family 25 member 13 Synaptosomal-associated protein 29 Sorting nexin-12 signal sequence receptor subunit 2 translocase of inner mitoch. membrane 50 translocase of outer mitoch. membrane 40 Transient receptor cation channel V 2 ubiquinol-cytochrome c reductase I
Protein Family DNA ligase dehydrogenase carbohydrate kinase . protein binding and sorting enzyme modulator AA transporter, protein transfer SNARE protein phosphatidyl & lipid bindingr G-protein-coupled receptor phosphatase membrane trafficking ion channel metalloprotease
12
Reactome pathway TGFbR complex signaling RA signaling, electron transport, sign. transduct. glucose metabolism mRNA splicing, FGFR signaling protein assembly transmembrane transport, NOTCH signaling glucose metabolism membrane traffic, vesicle-mediated transport membrane traffick regulato GPCR signaling IL2R binding protein channel, transmembrane transport transmembrane transport electron transport
Table S4 Protein profile of MDSC-Exosomes from in vitro / ex vivo MDSC Table S4A Top score peptides of MDSC-Exosomes from in vitro / ex vivo MDSCa MDSC-Exo in vitro
MDSC-Exo ex vivo
Synonym
Protein Description
Prot.M.
Pept.M.
Prot.M.
Pept.M.
Actb
Actin, cytoplasmic 1
477
42
589
38
Actr3
Actin-related protein 3
34
12
47
13
Ahcy
Adenosylhomocysteinase
21
7
53
10
Anxa1
Annexin A1
88
18
23
11
Anxa11
Annexin A11
53
10
29
6
Anxa2
Annexin A2
108
18
34
15
Anxa4
Annexin A4
56
14
23
10
Anxa5
Annexin A5
43
17
34
13
Anxa6
Annexin A6
64
19
10
3
Ap2m1
AP-2 complex subunit mu
21
10
2
1
Arpc2
Actin-related protein 2/3 complex subunit 2
24
12
C3
Complement C3
119
16
152
15
Cap1
Adenylyl cyclase-associated protein 1
19
11
38
10
Ccdc47
Coiled-coil domain-containing 47
33
14
Cct2
chaperonin containing TCP1 subunit 2
39
12
18
9
Cct3
chaperonin containing TCP1 subunit 3
32
10
4
4
Cct4
chaperonin containing TCP1 subunit 4
32
10
7
4
CD11b
Integrin alpha-M
83
21
39
16
CD13
Alanyl aminopeptidase, membrane
36
16
CD18
Integrin beta-2
78
27
32
17
Clic1
Chloride intracellular channel 1
26
10
26
7
Cltc
Clathrin heavy chain 1
208
50
14
9
Coro1a
Coronin-1A
41
10
24
8
Ctsb
Cathepsin B
9
5
47
10
Ctsd
Cathepsin D
25
6
52
10
Dera
Deoxyribose-phosphate aldolase
11
6
20
12
Eea1
Early endosome antigen 1
113
13
91
12
Eif3a
Eukaryotic translation initiation factor 3 A
127
23
Eif3b
Eukaryotic translation initiation factor 3 B
42
17
Eif3c
Eukaryotic translation initiation factor 3 C
128
19
Eif3e
Eukaryotic translation initiation factor 3 E
49
18
13
8
Eif3l
Eukaryotic translation initiation factor 3 L
32
10
Eif4a1
Eukaryotic initiation factor 4A1
46
15
26
12
Emilin2
elastin microfibril interfacer 2
20
10
Eno1
enolase 1
194
20
152
23
Ezr
Ezrin
33
12
Fasn
Fatty acid synthase
27
16
2
1
Fermt3
Fermitin family member 3
33
14
65
18
Flna
Filamin-A
27
16
23
10
Fn1
Fibronectin 1
580
71
71
25
Fth1
Ferritin heavy chain
171
15
132
12
Ftl1
Ferritin light chain 1
226
14
271
13
Gapdh
Glyceraldehyde-3-phosphate dehydrogenase
210
13
215
13
Gdi2
GDP dissociation inhibitor 2
21
12
13
8
Gsn
Gelsolin
86
11
H2-Ab1
histocompatibility 2, class II antigen A, beta 1
53
10
37
9
Hbb-b1
hemoglobin, beta adult major chain
205
10
507
13
Hbb-b2
hemoglobin, beta adult minor chain
122
10
306
13
Hmgb1
High mobility group box 1
95
11
16
4
Hmgb2
High mobility group box 2
52
10
26
7
Hsp90ab1
Heat shock protein HSP 90-beta
133
17
102
18
13
.
Table S4A cont. MDSC-Exo in vitro
MDSC-Exo ex vivo
Synonym
Protein Description
Prot.M.
Pept.M.
Prot.M.
Pept.M.
Hspa5
heat shock protein 5
43
11
22
4
Hspa8
Heat shock cognate 71 kDa protein
217
25
208
25
Ighm
Ig mu chain C region
9
5
86
14
Iqgap1
IQ motif containing GTPase activating protein 1
46
15
2
1
Klhdc4
Kelch domain-containing 4
27
11
Krt73
Keratin 73
31
4
67
13
LDHA
L-lactate dehydrogenase A chain
49
9
38
10
Lgals3bp
Galectin-3-binding protein Lgals3bp
149
21
116
19
Lpl
Lipoprotein lipase
174
12
59
9
Ltf
Lactotransferrin
98
29
89
26
Mfge8
Lactadherin
120
18
50
14
Mmp12
matrix metallopeptidase 12
324
22
229
20
Mmp19
matrix metallopeptidase 19
56
14
39
10
Mpo
Myeloperoxidase
15
9
56
16
Msn
Moesin
52
19
22
8
Mvp
Major vault protein
26
14
Myh1
myosin heavy chain 1
54
13
Myh4
myosin heavy chain 4
.
63
15
Myh9
myosin heavy chain 9
235
60
69
20
Ncl
Nucleolin
302
30
13
7
Pdcd6ip
Programmed cell death 6-interacting protein
118
27
12
6
Pgam1
Phosphoglycerate mutase 1
19
9
19
10
Pkm
Pyruvate kinase PKM
186
24
Ppib
Peptidyl-prolylisomerase B
39
12
26
9
Prdx1
Peroxiredoxin-1
79
10
16
6
Prkcd
protein kinase C delta
222
25
Psma1
Proteasome subunit alpha 1
34
9
38
11
Psma5
Proteasome subunit alpha 5
29
8
33
10
Psma6
Proteasome subunit alpha 6
34
10
39
10
Psma7
Proteasome subunit alpha 7
30
11
43
10
Psmb8
Proteasome subunit beta 8
34
8
49
11
Psmd11
proteasome 26S subunit, non-ATPase 11
37
13
7
4
Psmd2
proteasome 26S subunit, non-ATPase 2
32
19
3
3
Psmd3
proteasome 26S subunit, non-ATPase 3
24
13
2
1
Psmd6
proteasome 26S subunit, non-ATPase 6
35
16
19
10
Rab11b
RAB11B, member RAS oncogene family
35
11
30
9
Rack1
Receptor of activated protein C kinase 1
33
12
50
14
Rplp0
ribosomal protein P0
36
7
38
10
Ruvbl2
RuvB like AAA ATPase 2
20
10
4
3
Ssrp1
structure specific recognition protein 1
48
10
Tcp1
T-complex 1
42
14
2
1
Thbs1
Thrombospondin-1
60
17
104
23
Tln1
Talin-1
24
14
Tuba1b
Tubulin alpha-1B
127
18
210
36
Tuba1c
Tubulin alpha-1C
115
17
145
17
Tuba4a
Tubulin alpha-4A
91
16
144
18
Tubb4b
Tubulin beta-4B
107
19
120
19
Tubb5
Tubulin beta-5
115
20
124
19
Uba1
Ubiquitin-like modifier-activating enzyme 1
33
10
12
5
Vcp
valosin containing protein
226
41
37
14
a
.
.
Only proteins are shown with ≥10 significant peptides by at least one MDSC-Exo population
14
Table S4B Higher recovery in Exosomes from in vitro MDSC Synonym Actn1 Adam10 Anp32b Anxa6 Ap2a2 Ap2b1 Ap2m1 Arrb2 Ass1 Bin2 Brox Capg Capns1 Cbx3 Ccdc47 Ccl9 Cct6a Cct7 Cct8 CD13 CD316 CD36 Cltc Copb2 Csnk2a1 Cyp4f3 Ddost Dek Dhx9 Dip2b Dnaja2 Dync1h1 Eftud2 Eef1d Eif3a Eif3b
Exo - in vitro Prot.M. Pept.M. 8 4 9 5 5 3 64 19 10 4 28 6 21 10 13 6 16 8 17 4 4 3 7 4 9 3 11 5 33 14 13 3 20 7 5 3 13 7 36 16 24 8 14 7 208 50 10 5 6 4 17 6 6 3 38 9 5 4 4 3 6 3 32 21 14 8 3 3 127 23 42 17
Exo - ex vivo Prot.M. Pept.M. Protein Description actinin alpha 1 Disintegrin and metalloproteinase 10 Acidic nuclear phosphoprotein 32B 10 3 Annexin A6 AP-2 complex subunit alpha-2 AP-2 complex subunit beta 2 1 AP-2 complex subunit mu arrestin beta 2 Argininosuccinate synthase 1 Bridging integrator 2 BRO1 domain and CAAX motif capping actin protein, gelsolin like Calpain small subunit 1 Chromobox 3 Coiled-coil domain-containing 47 C-C motif chemokine 9 7 2 chaperonin TCP1 subunit 6A chaperonin containing TCP1 subunit 7 2 1 chaperonin TCP1 subunit 8 Alanyl aminopeptidase 13 3 2 Immunoglobulin superfamily 8 1 1 CD36, platelet glycoprotein 4 14 9 Clathrin heavy chain 1 coatomer complex beta 2 2 1 Casein kinase II alpha 1 3 1 cytochrome P450 family 4F3 DDoligosaccharide-glycosyltransferase DEK proto-oncogene DEAH-box helicase 9 Disco-interacting protein 2 homolog B DnaJ heat shock protein member A2 3 2 dynein cytoplasmic 1 heavy chain 1 elongation factor Tu GTP binding 2 Elongation factor 1-delta Eukaryotic translation initiation 3 A Eukaryotic translation initiation 3 B
Molecular function integrin, actin, vinculin binding metallopeptidase RNA & histone & protein binding protein complex & cholesterol binding lipid & protein binding transporter binding, signaling enzyme modulator binding, ligase phospholipid binding,kinase protein binding, molecular function actin binding, protein complex binding endopeptidase transcription factor ion & RNA binding cytokine & chemokine activity chaperonin ATP & protein binding chaperonin metalloprotease protein binding lipoprotein binding vesicle coat vesicle coat, structural molecule PTK, phosphatase regulator oxygenase DNA & histone binding hydrolase, protein binding molecular function, catalytic activity glycoprotein & HSP binding microtubule activity hydrolase, translation initiat & elong. translation elongation translation initiation translation initiation
15
Reactome pathway cell junction organization, cell-cell communication NOTCH, Ephrin, EGFR signaling apoptosis, differentiation contraction clathrin endocytosis, lipid transport, vesicle transport lipid & vesicle transport, clathrin endocytosis, signaling lipid transport, Gap junction traffic, clathrin endoc. clathrin endocyt., vesic.-med.transport, GPCR signaling AA & polyamine metabolism brassinosteroid signaling pathway actin filament capping epigenetic regul. of gene expression ER organization, osteopblast differention GPCR signaling protein folding chaperon- & protein folding protein folding angiogenesis, signaling, proteolysis CD81 partner, cell component movement TLR cascade, vesicle-mediated transport lipid transport, vesicle transport & biog., clathrin endoc. transport, vesicle-med.trasport,, membrane traffic. signaling, cell cycle Lipid metabolism T cell activ., neutrophil degranulation, cytokine response regulation of gene expression & AP-2 transcription MyD88, TLR, type I IFN signaling cascade cochaperone activity, protein refolding Vesicle & vesicle-med. transport, organelle biogenesis splicing translation, protein metabolism
Table S4B cont. Exo - in vitro Synonym Prot.M. Pept.M. Eif3c 128 19 Eif3i 11 6 Eif3l 32 10 Emilin2 20 10 Esyt1 22 9 Ezr 33 12 Fasn 27 16 Fgl2 11 5 Gca 4 3 Glg1 5 3 Gm2a 6 3 Gpi 5 3 Gsn 86 11 Hnrnpk 18 5 Iqgap1 46 15 Klhdc4 27 11 Lamp2 5 3 Lcp1 20 9 M6pr 11 4 Man2a1 12 8 Mvp 26 14 Myadm 10 3 Mybbp1a 10 3 Myh9 235 60 Ncl 302 30 Nrdc 10 7 Opn 20 3 Pabpc1 15 8 Pcbp1 5 3 Pdcd6ip 118 27 Pfkp 4 3 Pfn1 82 7 Prkcd 222 25 Psma2 21 8 Psmb5 7 3 Psmc3 7 3 Psmd1 12 7
Exo - ex vivo Prot.M. Pept.M. Protein Description Eukaryotic translation initiation 3 C 1 1 Eukaryotic translation initiation 3 I Eukaryotic translation initiation 3 L elastin microfibril interfacer 2 1 1 Extended synaptotagmin-1 Ezrin 2 1 Fatty acid synthase Fibroleukin Grancalcin Golgi apparatus protein 1 GM2 ganglioside activator Glucose-6-phosphate isomerase Gelsolin 4 1 Heterogeneous ribonucleoprotein K 2 1 IQ motif GTPase activating protein 1 Kelch domain-containing 4 Lysosome-associated glycoprotein 2 3 2 lymphocyte cytosolic protein 1 1 1 mannose-6-phosphate receptor mannosidase 2, alpha 1 Major vault protein Myeloid-assoc. differentiation marker Myb-binding protein 1A 69 20 myosin heavy chain 9 13 7 Nucleolin nardilysin convertase secreted phosphoprotein 1,Osteopontin poly(A) binding protein 1 Poly(rC)-binding protein 1 12 6 Programmed cell death 6-interact phosphofructokinase, platelet Profilin-1 protein kinase C delta Proteasome subunit alpha 2 Proteasome subunit beta 5 proteasome 26S subunit, ATPase 3 2 1 proteasome 26S subunit 1
Molecular function translation initiation translation initiation, protein binding translation initiation ECM protein ion & lipid binding cytoskeletal protein binding hyrolysae, transferase, ligase peptidase, signaling dimerization, metal ion binding fibroblast growth factor binding enzyme activator, hydrolase growth factor activity actin & myosin binding enzyme modulator G-protein modulator molecular function protein domain specific binding actin & protein binding GPCR activity hydrolase, binding phosphatase & kinase binding transcription corepressor, prot.binding actin & protein & RNA binding RNA & protein binding metalloprotease ECM binding, cytokine activity RNA & protein binding DNA & protein binding protein binding transferase and kinase activity binding non-receptor kinase endopeptidase endopeptidase hydrolase enzyme regulator
16
Reactome pathway
cell adhesion lipid metabolism Netrin1 signaling lipid metabolism signaling membrane fusion, neutrophil degranulation hemostasis lipid metabolism carbohydrate metabolism apoptosis, programmed cell death splicing Rho GTPase effector signaling platelet activation & signaling T cell activation, IL12 & PKA signaling clathrin endocytosis, vesicle transport protein transport & modification InsR signaling, transport, endocytosis raft organization, migration epigenetic regulation membrane traffic, Rho GTPase signaling LIF response, angiogenesis migration, proliferation ECM organization, signal transduction splicing, RNA silencing, translation splicing protein binidng Glucose metabolism Rho GTPase signaling IFN & cytokine & multiple signaling pathways ABC transporter, multiple signaling pathways multiple signaling pathways multiple signaling pathways multiple signaling cascades
Table S4B cont. Exo - in vitro Synonym Prot.M. Pept.M. Psmd11 37 13 Psmd13 19 7 Psmd2 32 19 Psmd3 24 13 Pygl 14 7 Rab10 26 3 Rap2b 4 3 Rdx 29 10 Rnh1 6 4 Rpl3 9 4 Rtn4 28 8 Ruvbl1 17 8 Ruvbl2 20 10 Scamp3 8 4 Sf3b3 12 6 Slc25a5 9 4 Smarca5 15 5 Snrpb 10 3 Ssrp1 48 10 Stk24 17 6 Tcp1 42 14 Tln1 24 14 Tm9sf2 15 9 Tm9sf3 8 3 Tmed4 7 4 Tmed9 11 5 Tra2b 36 4 U2surp 16 6 Vat1 10 5 Vcp 226 41 Vim 20 7 Vrk1 7 4 Wdr5 10 5 Ybx1 16 5 Ypel5 7 4
Exo - ex vivo Prot.M. Pept.M. 7 4 1 1 3 3 2 1
6 4 4
2 2 3
1
1
2
1
2 1
1 1
37
14
2 1 1
1 1 1
Protein Description proteasome 26S subunit 11 proteasome 26S subunit 13 proteasome 26S subunit 2 proteasome 26S subunit 3 Glycogen phosphorylase L RAB10, member RAS oncogene family RAP2B, member of RAS oncogene fam. Radixin ribonuclease/angiogenin inhibitor 1 ribosomal protein L3 Reticulon-4 RuvB like AAA ATPase 1 RuvB like AAA ATPase 2 secretory carrier membrane protein 3 Splicing factor 3B subunit 3 solute carrier family 25 member 5 Regulator of chromatin a5 Small nuclear ribonucleoprotein B & B1 structure specific recognition protein 1 Serine/threonine-protein kinase 24 T-complex 1 Talin-1 Transmembrane 9 member 2 Transmembrane 9 superfamily 3 transmembrane p24 trafficking protein 4 transmembrane p24 protein 9 Transformer-2 beta homolog U2 snRNP-associated SURP vesicle amine transport 1 valosin containing protein Vimentin vaccinia related kinase 1 WD repeat domain 5 Y-box binding protein 1 yippee-like 5
Molecular function protein binding, structural molecule endopeptidase. structural molecule enzyme regulator enzyme regulator phosphorylase GTPase, myosinV binding GTPase actin binding nuclease, hydrolase RNA binding protein binding protein binding protein binding ubiquitin ligase binding RNA & protein binding transporter hydrolase, protein binding protein & RNA binding transcription factor, signaling protein kinase peptidase, chaperonin cytoskeleton organization, binding transporter, molecular function transporter, molecular function signal transducer transfer / carrier, vesicle coat nucleic acid & RNA & protein binding mRNA processing reductase, AA transporter hydrolase, protein binding structural protein protein kinase, ATP binding esterase, kinase inhibitor, splicing transcription activator metal ion binding
17
Reactome pathway multiple signaling cascades APC transport, multiple signaling pathways multiple signaling pathways multiple signaling pathways carbohydrate metabolism membrane traffic, vesicle transport signal transduction, platelet activation RAP & PKA signal., junction organiz., endosome traffic. RNA processing signal transduction protein metabolism, signal transduction DNA repair, transcription vesicle-mediated transport splicing energy metabolism epigenetic gene regulation splicing transcriptional regulation Apoptosis, programmed cell death chaperonin & protein folding, protein metabolism integrin signaling
transmembrane transport transport & vesicle-med. transport splicing splicing transmembrane transporter APC & small molecule transporter, stress response apoptotic cleavage, programmed cell death cell cycle HOX gene activation transcription, splicing neutrophil degranulation
Table S4C Higher recovery in Exosomes from ex vivo MDSC Synonym Adh5 Alad Apoe Aprt Arf4 Arpc2 C1qc Ca2 CD147 CD177 CD41 Cfp Ctsg Hgd Hspa1a Hspa1l Ilk Itih2 Itm2b Jup Myh1 Myh4 Ngp Pkm Prdx2 Prdx6 Psap Rab5a Rpl7 Rps17 Tfrc Ube2n Vcl Ywhaq
Exo - in vitro Prot.M. Pept.M. . . 10 2 . . . 3 1 2 1 . 3 2 2 1 . 1 1 . . . . . . . . . 1 1 . 1 1 . 1 1 . 7 1 . . . . .
Exo - ex vivo Prot.M. Pept.M. 5 3 13 7 22 8 8 3 22 4 24 12 5 4 8 4 4 3 14 9 6 5 143 7 13 4 3 3 38 6 41 5 7 6 100 7 5 4 12 7 54 13 63 15 19 4 186 24 9 4 10 6 8 6 17 5 18 5 8 4 20 5 4 3 5 3 18 3
Protein Description Alcohol dehydrogenase class-3 aminolevulinate dehydratase Apolipoprotein E Adenine phosphoribosyltransferase ADP-ribosylation factor 4 Actin-related protein 2/3 subunit 2 complement C1q C chain Carbonic anhydrase 2 Basigin CD177 antigen Cd177 Integrin alpha-IIb complement factor properdin Cathepsin G Homogentisate 1,2-dioxygenase Heat shock 70 kDa protein 1A Heat shock 70 kDa protein 1-like Integrin-linked kinase Inter-alpha-trypsin inhibitor H2 Integral membrane protein 2B Junction plakoglobin myosin heavy chain 1 myosin heavy chain 4 Neutrophilic granule protein Pyruvate kinase PKM Peroxiredoxin-2 Peroxiredoxin-6 Prosaposin RAB5A, RAS oncogene family ribosomal protein L7 ribosomal protein S17 Transferrin receptor Ubiquitin-conjugating enzyme E2 N Vinculin 14-3-3 protein theta
Molecular function oxidoreductase, ion & fatty acid binding lyase, ion & protein binding protein & lipid binding transferase, AMP binding GTP & protein binding cytoskeleton, protein complex binding endopeptidase dehydrase, lyase, metal ion binding carbohydrate binding molecular function protein binding ECM component serine protease oxidoreductase protein binding protein folding, GPCR binding non-receptor tyrosine kinase protease inhibitor protein binding protein binding G-protein modulator G protein modulator peptidase inhibitor catalytic activity, protein binding protein binding, oxidase binding, catalytic & hydrolase activity immune defense, surfactant GTPase DNA & RNA binding structural ribosome constituent receptor activity, protein binding transferase, protein binding Rho GTPase, protein & actin binding chaperone, protein binding
18
Reactome pathway metabolism, oxidation Porphyrin & heme metabolsim vesicle & lipid transport, signaling purine metabolism organelle biogenesis, trafficking & transport Rho GTPase & Ephrin signal., endocyt., ves.transport complement cascade, granuloc. & M differentiation O2/CO2 metabolism ECM organization, membrane transport fibrin clot formation integrin signaling, ECM organization complement activation ECM organization, MMP activation, IGF transport AA metabolism stress response ECM & cell junction organization hyaluronan & protein metabolism protein metabolism, amyloid precursor regulation junction organization, VEGF signaling membrane & vesicle med. trafficking defense, angigoenesis regulation carbohydrate metabolism stress response stress response TLR signaling clathrin endocyt., membr. & ves. transport, ves. biogen translation initiation translation, mRNA catabolism clathrin endocyt., membr. & ves. transport, ves. biogen. IFN-, cytokine-,TLR signaling platelet activation & signaling Rho GTPase sign., BAD activ., vesicle transport
Table S5 Differences in MDSC versus MDSC-Exo protein profile Table S5A Preferential peptide recovery in MDSC-Exo Synonym. Ap2m1 Ap2S1 Arrb2 Atp6ap2 C1qtnf3 C3 Casp6 Ccdc47 Ccl9 Cct4 CD316 CD81 Col1a1 CSN3 CSN4 CSN6 CSN8 Cyp4f3 Dek Dera Eea1 Eftud2 Eif3a Eif3b Eif3c Eif3e Eif3f Eif3g Eif3l Eif3m Emilin2 Fgl2 Fn1 H2afy
MDSC Prot.M. Pept.M. 5 2 . 2 1 . . 19 5 3 . 9 . . . . . . . 3 4 . 5 . 9 21 3 2 8 1 1 2 . . . .
2 3
2 2 4 4 4 2 2 1 1 1 1
MDSC-Exo Prot.M. Pept.M. 21 10 11 5 13 6 9 6 7 2 119 16 5 4 33 14 13 4 32 10 24 8 14 5 4 4 3 2 4 4 4 4 3 2 17 6 38 9 11 6 113 13 14 8 121 21 42 17 128 19 49 18 42 9 20 5 32 10 27 7 20 10 11 5 580 71 82 14
Protein Description AP-2 complex mu AP-2 complex sigma arrestin beta 2 ATPase H+ transporting 2 C1q and TNF related protein 3 Complement C3 Caspase-6 Coiled-coil domain- 47 C-C chemokine 9 chaperonin TCP1 subunit 4 Immunoglobulin superfamily 8 CD81 antigen collagen type I alpha 1 COP9 signalosome subunit 3 COP9 signalosome subunit 4 COP9 signalosome subunit 6 COP9 signalosome subunit 8 cytochrome P450 family 4F3 DEK proto-oncogene Deoxyribose-phosphate aldolase Early endosome antigen 1 elongation factor Tu 2 Eukaryotic initiation factor 3 A Eukaryotic initiation factor 3 B Eukaryotic initiation factor 3 C Eukaryotic initiation factor 3 E Eukaryotic initiation factor 3 F Eukaryotic factor 3 G Eukaryotic initiation factor 3 L Eukaryotic initiation factor 3 M elastin microfibril interfacer 2 fibroleukin Fibronectin 1 Core histone H2A.1
19
Function protein binding, receptor vesicle coat protein enzyme modulator enzyme binding, receptor protein binding cytokine, protease inhibitor peptidase calcium & RNA binding chemokine chaperonin protein binding adhesion, signaling protein binding, ECM molecular function protein binding, protease transcription, peptidase protein binding, signaling oxygenase DNA / RNA binding lyase ion/protein binding G protein, hydrolase translation protein/RNA binding, transcript. translation binding, translation peptidase, translation binding, translation translation translation ECM protein signaling, peptidase binding, signaling histone, binding
Reactome pathway trafficking, transmission, endocytosis, signaling lipid & vesicel transport, endocytosis, cargo recognition, signaling multiple signaling pathways (HH, WNT5A, FZD4, GPCR), metabolism (angiotensin) complement cascade GPCR signaling, complement activation apoptosis execution Ca++ homeostasis, ER organization, osteoblast differentiation GPCR signaling protein folding & metabolism proliferation, cell component movement immune regulation ECM organization endocytosis, cargo recogn., membrane traffic, vesicle transport excision repair, endocytosis, vesicle transport transcription, endocytosis, membrane traffic, vesicle transport endocytosis, trafficking, cargo recognition, vesicle transport lipid metabolism regulation of gene expressio carbohydrate metabolism TLR cascades splicing translation initiation translation, metabolism translation initiation translation initiation translation, protein metabolism translation initiation translation initiation translation, protein metabolism cell adhesion neutrophil degranulation matrix formation, multiple sign. cell cycle, rRNA regulation
Table S5A cont. Synonym. Hist1h3a Hist1h3b Hist2h2aa1 Ighm Klhdc4 Lpl Mfge8 Mmp12 Mmp19 Nrdc Opn,Spp1 Paics Pdcd6ip Plscr3 Polr2c Prkcd Psmd1 Psmd13 Psmd2 Psmd3 Psmd6 Ruvbl1 Ruvbl2 Sbno1 Scimp Sdcbp Sec13 Serpinc1 Srsf1 Ssrp1 Stk24 Sub1 Supt16h Tnfsf13 Tspan14 Vps28 Wdr5
MDSC Prot.M. Pept.M. . . . . . . 19 5 1 1 . . . . 6 3 . . 6 4 1 1 1 1 7 6 5 3 1 1 2 1 4 3 . . 4 1 . . 6 2 . . 1 1 1 1 . 1 1 . 1 1
MDSC-Exo Prot.M. Pept.M. 64 5 69 5 107 7 9 5 27 11 174 12 120 18 324 22 56 14 10 7 20 4 8 5 118 27 5 3 10 6 222 25 12 7 19 7 32 19 24 13 35 16 17 8 20 10 2 2 6 2 68 6 4 2 41 6 67 8 48 10 17 6 58 8 7 5 4 2 9 5 11 5 10 5
Protein Description Histone H3.1 Histone H3.2 Histone H2A type 2A Ig mu chain C region Kelch domain-containing 4 Lipoprotein lipase Lactadherin matrix metallopeptidase 12 matrix metallopeptidase 19 nardilysin convertase Osteopontin p-ribosylamin lase/synthase Programmed cell death 6-interact. Phospholipid scramblase 3 RNA polymerase II subunit C protein kinase C delta proteasome 26S, non-ATPase 1 proteasome 26S, non-ATPase 13 proteasome 26S, non-ATPase 2 proteasome 26S, non-ATPase 3 proteasome 26S, non-ATPase 6 RuvB like AAA ATPase 1 RuvB like AAA ATPase 2 strawberry notch homolog 1 SLP adapter & CSK-interact. p. syndecan binding protein SEC13, COPII coat complex serpin family C member 1 Serine/arginine-rich splicing F1 1 structure recognition protein 1 Serine/threonine-protein kinase 24 SUB1 transcriptional regulator SPT16 chromatin remodeling TNF superfamily member 13 Tetraspanin-14 VPS28, ESCRT-I subunit WD repeat domain 5
20
Function histone histone, binding histone binding molecular function esterase, lipase binding, protease, transporter peptidase metallopeptidase metalloprotease cytokine ligase, lyase, binding binding transfer/carrier protein DNA & protein binding transferase, kinase, carrier enzyme modulator enzyme modulator binding, enzyme modulator enzyme modulator regulator binding, hydrolase protein & ATP binding molecular function protein binding binding, traffic regul. traffic. reulator protease inhibitor RNA & protein binding HMG box TF, signaling kinase, transferase transcription coactiv. RNA binding, transcript. cofactor cytokine adhesion, signaling binding esterase, protein binding
Reactome pathway multiple signaling pathways, stress response cytokine/interleukin signaling, stress response metalloprotease, cell cycle regulation BCR signaling retinoid/lipid metabolsim/transport signaling, transporter ECM organization/degradation ECM degradation migration, proliferation ECM organization, signaling metabolism TJ & MVB & Exo assembly, protein transport apoptosis, cholesterol & glucose homeostasis splicing, transcription, signaling multiple signaling pathways multiple sign.pathways multiple signaling pathways multiple signaling pathways multiple signaling pathways multiple signaling pathways, protein metabolism, ß-cat/TCF activation DNA repair, transcription activation, protein folding transcription regulation immune synapse formation Ephrin signaling Rho signaling, cell cycle, vesicle transport, membrane traffic. clotting cascade splicing, mRNA transport transcription apoptosis execution transcription regulation, SMAD signaling transcription cytokine signaling (TNFR) plasma membrane organization, NOTCH sign. neutrophil degranulation ESCRT, vesicle transport kinase inhibitor
Table S5B Higher peptide recoveryn in MDSC than MDSC-Exo Synonym. Acaa1a Acad Aco2 Acsl1 Actn1 Actn4 Adpgk Agps Ak2 Akr1a1 Aldh2 Alox5 Ankfy1 Anp32b Ap1b1 Api5 Apmap Arf4 Arg1 Arhgap1 Arl8a Arl8b Arpc2 Asah1 Atp1a1 Atp2a2 Atp2a3 Atp5a1 Atp5b Atp5c1 Atp5f1 Atp5h Atp5i Atp5o Atp6v1a Atp6v1b2 B4galnt1 Bax Bcap31 Blvrb Bzw1 Ca2 Calr Canx Capn1 Cat CD166 CD177 CD180 CD206 CD29 CD41 CD61 CD74 Celf2 Chil3 Ckap4 Cmpk2 Cndp2
Protein Description acetyl-CoA acyltransferase 1A Acyl-CoA dehydrogenase Aconitate hydratase, mitochondrial Acyl-CoA synthetase long-chain family member 1 actinin alpha 1 actinin alpha 4 ADP-dependent glucokinase alkylglycerone phosphate synthase Adenylate kinase 2 aldo-keto reductase family 1 member A1 aldehyde dehydrogenase 2 family Arachidonate 5-lipoxygenase Ankyrin repeat and FYVE domain containing 1 Acidic nuclear phosphoprotein 32 B AP-1 complex subunit beta-1 Apoptosis inhibitor 5 Adipocyte plasma membrane-associated protein ADP-ribosylation factor 4 Arginase-1 Rho GTPase-activating protein 1 ADP-ribosylation factor-like GTPase 8A ADP-ribosylation factor-like GTPase 8B Actin-related protein 2/3 complex subunit 2 N-acylsphingosine amidohydrolase 1 ATPase Na+/K+ transporting subunit alpha 1 ATPase Ca2+ transporting 2 ATPase Ca2+ transporting 3 ATP synthase, H+ transporting F1 alpha subunit 1 ATP synthase, H+ transporting F1 beta subunit ATP synthase, H+ transporting F1 gamma subunit ATP synthase, H+ transporting, F0 subunit B1 ATP synthase, H+ transporting, F0 subunit D ATP synthase, H+ transporting, F0 subunit E ATP synthase, H+ transporting, F1 subunit O ATPase H+ transporting V1 subunit A ATPase H+ transporting V1 subunit B2 Beta-1,4 N-acetylgalactosaminyltransferase 1 BCL2 associated X, apoptosis regulator B-cell receptor-associated protein 31 biliverdin reductase B Basic leucine zipper and W2 domain 1 Carbonic anhydrase 2 Calreticulin Calnexin Calpain-1 Catalase CD166 antigen, Alcam CD177 antigen CD180 antigen mannose receptor C-type 1 Integrin beta-1 Integrin alpha-IIb Integrin beta-3 CD74 antigen, invariant chain associated CUGBP Elav-like family member 2 Chitinase-like 3 Cytoskeleton-associated protein 4 cytidine/uridine monophosphate kinase 2 CNDP dipeptidase 2
21
MDSCinvitro Prot.M. Pept.M. 20 7 4 9 78 20 30 9 141 35 128 36 36 11 23 5 30 9 43 11 262 29 49 13 8 5 30 14 33 12 8 5 8 6 38 8 35 10 12 5 23 8 37 11 83 15 12 6 49 11 72 15 29 6 331 25 296 25 46 6 30 11 21 8 12 5 42 9 52 16 20 5 17 12 14 6 10 5 10 5 14 8 21 7 159 22 87 19 29 18 30 15 24 12 39 13 15 8 48 27 27 9 23 12 14 6 23 5 10 5 63 13 54 20 37 12 77 16
MDSC in vitro-Exo Prot.M. Pept.M.
8
4
7
5
5
3
4
3
3
2
8
4
1 1
1 1
2
1
1
1
2 3
1 2
1 6 2
1 3 1
3
1
5
2
6
3
Table S5B cont. Synonym. Colgalt1 Copz1 Cox4i1 Cox5a Cox6c Cs Ctsa Ctsc Ctsg Ctsz Cyb5r3 Cybb Cycs Dars Ddost Ddx39a Ddx39b Ddx3x Ddx5 Dhx9 Diaph1 Dlat Dld Dlst Dnajb11 Dnajc3 Dock8 Dpysl2 Dstn Ear2 Echs1 Eef1b Eef1d Eef1g Eef2 Ehd1 Ehd4 Eif1 Eif4a3 Eif5a Elane Elavl1 Emb Entpd1 Erap1 Ero1a Erp29 Erp44 Esyt1 Etfa Etfb Fabp5 Fam129b Fam162a Fam49b Fh Fis1 Fkbp2 Fkbp3
Protein Description collagen beta(1-O)galactosyltransferase 1 Coatomer protein complex subunit zeta 1 cytochrome c oxidase subunit IV isoform 1 cytochrome c oxidase subunit 5A Cytochrome c oxidase subunit 6C Citrate synthase cathepsin A cathepsin C Cathepsin G Cathepsin Z cytochrome b5 reductase 3 Cytochrome b-245 beta chain Cytochrome c, somatic Aspartate--tRNA synthase dolichyl-diphosphooligosacch glycosyltransfer. DExD-box helicase 39A DExD-box helicase 39B DEAD-box helicase 3, X-linked DEAD-box helicase 5 DEAH-box helicase 9 diaphanous related formin 1 dihydrolipoamide S-acetyltransferase dihydrolipoamide dehydrogenase dihydrolipoamide S-succinyltransferase DnaJ heat shock protein family B11 DnaJ heat shock protein family C3 Dedicator of cytokinesis 8 Dihydropyrimidinase like 2 Destrin eosinophil-associated, ribonuclease A2 Enoyl-CoA hydratase, short chain 1 Elongation factor 1-beta Elongation factor 1-delta Elongation factor 1-gamma Elongation factor 2 EH domain-containing 1 EH domain-containing 4 Eukaryotic translation initiation factor 1 Eukaryotic initiation factor 4A3 Eukaryotic translation initiation factor 5A elastase, neutrophil expressed ELAV like RNA binding protein 1 Embigin Ectonucleos. triphosphate diphosphohydrolase 1 Endoplasmic reticulum aminopeptidase 1 endoplasmic reticulum oxidoreductase 1 alpha Endoplasmic reticulum protein 29 Endoplasmic reticulum protein 44 Extended synaptotagmin-1 Electron transfer flavoprotein alpha subunit Electron transfer flavoprotein beta subunit Fatty acid-binding protein 5 family with sequence similarity 129 member B family with sequence similarity 162 member A family with sequence similarity 49 member B Fumarate hydratase fission, mitochondrial 1 FK506 binding protein 2 FK506 binding protein 3
22
MDSCinvitro Prot.M. Pept.M. 6 5 11 7 34 9 12 5 11 5 47 11 15 7 20 6 23 8 32 7 33 11 50 13 19 7 14 8 53 11 80 14 64 6 17 7 50 12 30 11 16 8 10 5 27 10 27 6 12 6 9 6 7 5 18 11 46 6 22 7 14 8 301 15 19 6 152 12 269 35 29 13 45 12 10 6 27 6 51 14 26 5 13 6 22 6 7 5 32 5 79 12 37 9 23 8 217 37 30 8 19 8 75 15 29 9 13 5 23 9 16 10 14 5 11 5 8 5
MDSC in vitro-Exo Prot.M. Pept.M.
2
2
4 1 2
2 1 1
6 1
3 1
4 4
2 3
3
2
2 4
2 2
1 2 8 29 6 8
1 2 4 8 2 4
4
1
1
1
3
2
22 2
9 1
1
1
3
2
Table S5B cont. Synonym. Flna G3bp1 Ganab Gba Glb1 Glg1 Glo1 Glud1 Gmfg Gnai2 Gnb1 Gnb2 Got2 Gpd2 Gpi Gpx1 Grb2 Gsr Gstm1 Gsto1 H2-D1 H2-Eb1 Hadha Hadhb Hars Hdac1 Hexa Hexb Hk1 Hk2 Hk3 Hmox1 Hmox2 Hnrnpa1 Hnrnpa2b1 Hnrnpa3 Hnrnpab Hnrnpd Hnrnpf Hnrnph1 Hnrnpk Hnrnpl Hnrnpm Hpcal1 Hprt1 Hsd11b1 Hsd17b11 Hsd17b12 Hsd17b4 Hsp90aa1 Hsp90b1 Hspa1a Hspa5 Hspd1 Hspe1 Hyou1 Idh2 Idh3a Ifi44
Protein Description Filamin-A GTPase activating protein binding protein 1 glucosidase II alpha subunit Glucosylceramidase beta galactosidase, beta 1 Golgi apparatus protein 1 glyoxalase I Glutamate dehydrogenase 1 Glia maturation factor gamma guanine nucleotide binding protein alpha inhibit 2 guanine nucleotide binding protein beta 1 guanine nucleotide binding protein beta 2 Aspartate aminotransferase, mitochondrial Glycerol-3-phosphate dehydrogenase 2 Glucose-6-phosphate isomerase Glutathione peroxidase 1 Growth factor receptor-bound protein 2 Glutathione disulfide reductase Glutathione S-transferase Mu 1 Glutathione S-transferase omega-1 histocompatibility 2, D region locus 1 histocompatibility 2, class II antigen E beta hydroxyacyl-CoA dehydrogenase alpha subunit hydroxyacyl-CoA dehydrogenase beta subunit histidyl-tRNA synthetase Histone deacetylase 1 Hexosaminidase subunit alpha Hexosaminidase subunit beta Hexokinase-1 Hexokinase-2 Hexokinase-3 Heme oxygenase 1 Heme oxygenase 2 Heterogeneous ribonucleoprotein A1 Heterogeneous ribonucleoproteins A2/B1 Heterogeneous ribonucleoprotein A3 Heterogeneous ribonucleoprotein A/B Heterogeneous ribonucleoprotein D0 Heterogeneous ribonucleoprotein F Heterogeneous ribonucleoprotein H1 Heterogeneous ribonucleoprotein K Heterogeneous ribonucleoprotein L Heterogeneous ribonucleoprotein M Hippocalcin-like 1 Hypoxanthine phosphoribosyltransferase 1 hydroxysteroid 11-beta dehydrogenase 1 hydroxysteroid 17-beta dehydrogenase 11 hydroxysteroid 17-beta dehydrogenase 12 hydroxysteroid 17-beta dehydrogenase 4 Heat shock protein HSP 90-alpha heat shock protein 90, beta 1 Heat shock 70 kDa protein 1A heat shock protein 5 heat shock protein family D (Hsp60) member 1 heat shock protein family E (Hsp10) member 1 Hypoxia up-regulated protein 1 Isocitrate dehydrogenase [NADP] 2 Isocitrate dehydrogenase [NAD] alpha Interferon-induced protein 44
23
MDSCinvitro Prot.M. Pept.M. 391 71 12 5 113 26 10 6 8 6 32 10 12 6 77 21 13 6 140 15 28 8 52 12 93 19 30 9 113 21 9 5 13 7 16 5 26 12 13 6 132 11 18 6 33 12 25 9 10 6 9 6 14 8 18 11 74 15 53 12 45 13 26 7 27 10 95 15 82 17 94 11 33 6 34 8 23 6 67 13 105 12 31 11 24 6 9 5 9 5 24 5 28 6 31 6 9 5 255 27 457 35 31 7 544 38 69 24 16 6 169 30 63 14 41 9 27 9
MDSC in vitro-Exo Prot.M. Pept.M. 27 16
5
3
19
5
3
2
5
3
2
2
1
1
7
4
2 2
1 1
1 2 4
1 1 2
14 1
3 1
2
1
20
4
43
11
Table S5B cont. Synonym. Ifit1 Ifit2 Immt Kctd12 Kpnb1 Lap3 Lbr Lcp1 Lgals1 Lgals9 Lmnb1 Lnpep Lpcat2 Lrpap1 Lrpprc Lrrc59 Lta4h Man2b1 Mapre1 Mcm2 Mdh1 Mdh2 Me2 Mogs Mta2 Mthfd1l Mybbp1a Naga Naa50 Napa Nceh1 Ncf1 Ncf2 Ncf4 Ncstn Ndufa13 Ndufa8 Ndufb10 Ndufs3 Nme1 NOMO1 Nucb1 Nudt21 Oas1a Oat Ogdh Osbpl8 Ostf1 Oxct1 P4hb Pai2 Pcbp2 Pdcd10 Pdia3 Pdia4 Pdia6 Phb Phb2 Pitpna
Protein Description Interferon-induced protein 1 Interferon-induced protein 2 inner membrane mitochondrial protein potassium channel 12 karyopherin subunit beta 1 leucine aminopeptidase 3 Lamin-B receptor lymphocyte cytosolic protein 1 Galectin-1 Galectin-9 Lamin-B1 Lmnb1 Leucyl-cystinyl aminopeptidase Lysophosphatidylcholine acyltransferase 2 LDL receptor related protein associated protein 1 leucine rich pentatricopeptide repeat containing Leucine-rich repeat-containing protein 59 Leukotriene A-4 hydrolase mannosidase alpha class 2B member 1 Microtubule-associated protein RP/EB member 1 minichromosome maintenance complex 2 Malate dehydrogenase 1 Malate dehydrogenase 2 Malic enzyme 2 Mannosyl-oligosaccharide glucosidase metastasis associated 1 family member 2 methylenetetrahydrofolate dehydrogenase 1-like Myb-binding protein 1A Alpha-N-acetylgalactosaminidase N-alpha-acetyltransferase 50 NSF attachment protein alpha Neutral cholesterol ester hydrolase 1 Neutrophil cytosol factor 1 Neutrophil cytosol factor 2 Neutrophil cytosol factor 4 Nicastrin NADH:ubiquinone oxidoreductase A13 NADH:ubiquinone oxidoreductase A8 NADH:ubiquinone oxidoreductase B10 NADH:ubiquinone oxidoreductase S3 NME/NM23 nucleoside diphosphate kinase 1 Nodal modulator 1 Nucleobindin-1 nudix hydrolase 21 2'-5'-oligoadenylate synthase 1A Ornithine aminotransferase oxoglutarate dehydrogenase Oxysterol-binding protein like 8 Osteoclast-stimulating factor 1 3-oxoacid CoA-transferase 1 prolyl 4-hydroxylase subunit beta Plasminogen activator inhibitor 2 Poly(rC)-binding protein 2 Programmed cell death 10 Protein disulfide-isomerase A3 Protein disulfide-isomerase A4 Protein disulfide-isomerase A6 Prohibitin Prohibitin-2 Phosphatidylinositol transfer protein alpha
24
MDSCinvitro Prot.M. Pept.M. 40 12 19 6 7 5 28 7 55 13 19 12 38 8 486 38 30 8 14 5 21 10 37 15 17 5 26 10 11 5 43 9 64 13 23 10 11 5 7 5 16 9 114 16 17 8 20 6 10 6 14 6 29 12 9 6 16 8 16 8 17 6 41 10 24 10 18 9 17 7 17 5 13 6 10 7 7 5 49 9 14 7 10 7 13 7 47 11 16 9 38 15 20 7 9 6 19 6 286 31 19 10 52 6 9 5 263 37 182 32 113 15 36 8 34 13 9 7
MDSC in vitro-Exo Prot.M. Pept.M.
4 1
2 1
20 1
9 1
1
1
5 11
3 5
10
3
3
2
3
2
3
2
6
2
3
2
3
2
Table S5B cont. Synonym. Pkm Plbd1 Pld4 Plec Plek Pnkp Pnp Por Ppa1 Ppif Ppp1ca Ppp1cb Ppp1cc Ppp2ca Ppp2r1a Prdx2 Prdx3 Prdx4 Prdx5 Prdx6 Prtn3 Psma4 Psme1 Ptbp3 Ptgr1 Ptgs1 Ptpn1 Ptprc Pycard Pygl Rab18 Rab21 Rab27a Rab2a Rab31 Rab5a Ralb Rap2c Rbbp4 Rbbp7 Rbmxl1 Rnf213 Rnh1 Rpl13 Rpl13a Rpl15 Rpl17 Rpl18a Rpl4 Rpl7 Rpn1 Rpn2 Rps14 Rps16 Rps17 Rps9 Rrbp1 Samhd1 Sars
Protein Description Pyruvate kinase PKM phospholipase B domain containing 1 Phospholipase D4 Plectin Pleckstrin polynucleotide kinase 3'-phosphatase Purine nucleoside phosphorylase cytochrome P450 oxidoreductase pyrophosphatase 1 Peptidyl-prolylisomerase F protein phosphatase 1 catalytic subunit alpha protein phosphatase 1 catalytic subunit beta protein phosphatase 1 catalytic subunit gamma protein phosphatase 2 alpha Protein phosphatase 2A subunit A Peroxiredoxin-2 Peroxiredoxin-3 Peroxiredoxin-4 Peroxiredoxin-5 Peroxiredoxin-6 proteinase 3 Proteasome subunit alpha 4 proteasome activator subunit 1 Polypyrimidine tract-binding protein 3 Prostaglandin reductase 1 prostaglandin-endoperoxide synthase 1 protein tyrosine phosphatase, non-receptor type 1 protein tyrosine phosphatase, receptor type C PYD and CARD domain containing Glycogen phosphorylase L RAB18, member RAS oncogene family RAB21, member RAS oncogene family RAB27A, member RAS oncogene family RAB2A, member RAS oncogene family RAB31, member RAS oncogene family RAB5A, member RAS oncogene family RAS like proto-oncogene B RAP2C, member of RAS oncogene family RB binding protein 4 RB binding protein 7 RNA binding motif protein, X-linked-like-1 ring finger protein 213 ribonuclease/angiogenin inhibitor 1 ribosomal protein L13 ribosomal protein L13a ribosomal protein L15 ribosomal protein L17 ribosomal protein L18a ribosomal protein L4 ribosomal protein L7 ribophorin I ribophorin II ribosomal protein S14 ribosomal protein S16 ribosomal protein S17 ribosomal protein S9 Ribosome-binding protein 1 SAM and HD domain cont. triphosphohydrolase 1 seryl-tRNA synthetase
25
MDSCinvitro Prot.M. Pept.M. 403 31 15 7 19 7 33 9 14 5 8 5 65 11 67 14 8 5 8 5 36 13 19 9 25 11 12 7 15 7 15 5 45 5 50 5 67 12 30 13 24 6 18 6 38 11 19 8 7 5 22 8 48 9 85 17 8 5 106 22 13 6 21 8 10 5 39 9 11 5 20 6 11 5 13 6 16 5 23 5 22 5 8 6 52 14 22 8 18 5 12 5 24 5 30 6 79 12 31 13 169 21 95 15 33 7 50 8 21 7 38 12 46 19 49 16 10 5
MDSC in vitro-Exo Prot.M. Pept.M. 2
1
1
1
2
2
5
2
1
1
4
3
2 1
1 1
14
7
5
2
1 2
1 1
6 8
4 2
2 2
1 1
5 7
2 1
14
2
1
1
Table S5B cont. Synonym. Scp2 Sdf2l1 Sdha Sec11c Sec22b Sec61a1 Sept11 Serpinb1a Sfpq Sfxn1 Sgpl1 Sh3bgrl Siglec1 Slc25a11 Slc25a12 Slc25a3 Slc25a4 Slc25a5 Slc3a2 Snd1 Snx2 Sod2 Spcs2 Sptan1 Ssr4 Stat1 Stfa1 Stfa3 Stip1 Stt3a Stx7 Sun2 Surf4 Syncrip Tagln2 Taldo1 Tap1 Tap2 Tapbp Tfrc Tgm2 Tkt Tln1 Tln2 Tmem43 Tmpo Tor1aip1 Tpi1 Tpr Trap1 Trex1 Tubb2a Txn Txndc5 U2af2 Ube2n Ube2v1 Uggt1 Ugt1a7c
Protein Description sterol carrier protein 2 Stromal cell-derived factor 2-like 1 succinate dehydrogenase complex flavoprotein A SEC11 homolog C, signal peptidase complex SEC22 homolog B, vesicle trafficking protein Sec61 translocon alpha 1 Septin-11 Leukocyte elastase inhibitor A Splicing factor, proline- and glutamine-rich Sideroflexin-1 Sphingosine-1-phosphate lyase 1 SH3 domain binding glutamate rich protein like sialic acid binding Ig-like lectin 1, sialoadhesin solute carrier family 25 member 11 solute carrier family 25 member 12 solute carrier family 25 member 3 solute carrier family 25 member 4 solute carrier family 25 member 5 solute carrier family 3 member 2 Staphylococcal nuclease & tudor domain 1 Sorting nexin-2 Superoxide dismutase 2 Signal peptidase complex subunit 2 Spectrin alpha, non-erythrocytic 1 signal sequence receptor subunit 4 Signal transducer and activator of transcription 1 Stefin A1 Stefin A3 Stress-induced-phosphoprotein 1 STT3A, oligosaccharyltransferase complex Syntaxin-7 SUN domain-containing protein 2 Surfeit 4 synaptotagmin binding protein Transgelin-2 Transaldolase 1 transporter 1, ATP binding cassette subfamily B transporter 2, ATP binding cassette subfamily B TAP binding protein Transferrin receptor transglutaminase 2 Transketolase Talin-1 Talin-2 Transmembrane protein 43 thymopoietin Torsin-1A-interacting protein 1 Triosephosphate isomerase translocated promoter region TNF receptor associated protein 1 Three-prime repair exonuclease 1 Tubulin beta-2A Thioredoxin Thioredoxin domain containing 5 U2 small nuclear RNA auxiliary factor 2 Ubiquitin-conjugating enzyme E2 N Ubiquitin-conjugating enzyme E2 variant 1 UDP-glucose glycoprotein glucosyltransferase 1 UDP glucuronosyltransferase 1 polypeptide A7C
26
MDSCinvitro Prot.M. Pept.M. 20 10 20 6 19 6 9 5 12 5 12 5 23 8 156 19 19 7 15 6 17 6 9 5 6 5 33 6 40 10 93 11 109 13 204 15 91 16 51 21 12 6 11 5 16 5 7 5 29 5 27 7 17 6 13 6 13 6 18 6 17 6 125 20 24 5 19 9 66 12 52 15 16 6 21 8 66 11 30 9 21 6 163 20 297 57 26 6 15 9 12 6 23 9 43 10 14 9 20 5 14 5 166 15 8 5 11 6 9 5 15 5 8 5 116 39 35 9
MDSC in vitro-Exo Prot.M. Pept.M. 1
1
1
1
3
2
2
1
9 11 1
4 5 1
2
1
1
1
4
2
4 8
2 3
4
1
1 24
1 14
1
1
6
3
1
1
Table S5B cont. Synonym. Uqcrc2 Uso1 Vamp8 Vapa Vars Vcl Vdac1 Vdac2 Vdac3 Vim Vps35 Vwa5a Wdr1 Xpo1 Ywhaq
Protein Description ubiquinol-cytochrome c reductase core protein II USO1 vesicle transport factor Vesicle-associated membrane protein 8 VAMP associated protein A valyl-tRNA synthetase Vinculin Voltage-dependent anion channel 1 Voltage-dependent anion channel 2 Voltage-dependent anion channel 3 Vimentin vacuolar protein sorting 35 homolog von Willebrand factor A domain 5A WD repeat domain 1 Exportin-1 14-3-3 protein theta
27
MDSCinvitro Prot.M. Pept.M. 22 11 17 6 12 5 21 5 15 7 27 14 64 15 63 11 40 7 371 42 49 13 24 12 57 14 11 5 48 6
MDSC in vitro-Exo Prot.M. Pept.M.
2
1
2
2
20 6
7 2
1
1
Table S6 a MDSC-Exo peptides recovered in pulldown from Sepharose-coupled lymph node cell membranes MDSC-Exo over LNC membrane Prot.M.
cellular localization
Synonym
Protein Description
Acaa2
acetyl-CoA acyltransferase 2
14
9
mitochondrion
Acadvl
acyl-CoA dehydrogenase, very long chain
21
12
mitochondrial inner membr.
Aco2
Aconitate hydratase, mitochondrial
8
5
mitochondrion, myelin sheet
Acta2
actin, alpha 2, smooth muscle, aorta
68
20
cytoskeleton, cytoplasm
Aldh2
aldehyde dehydrogenase 2 family
15
9
mitochondrion
Ank1
Ankyrin-1
6
5
cytoskel., plasma membrane
Anxa11
Annexin A11
9
5
cytoskel., membrane
Anxa4
Annexin A4
8
5
plasma membrane
Arhgdib
Rho GDP-dissociation inhibitor beta
7
6
cytoplasm, membrane
Arhgef1
Rho guanine nucleotide exchange factor 1
6
5
cytoplasm, plasma membrane
Atp1a1
ATPase Na+/K+ transporting subunit alpha 1
22
11
ER & plasma membrane, raft
Atp2a1
ATPase ER Ca2+ transporting 1
27
13
ER membrane
Atp2a3
ATPase ER Ca2+ transporting 3
14
8
ER membrane
Cap1
CAP, adenylate cyclase-associated protein 1
17
8
cytoskeleton, raft
Capg
capping actin protein, gelsolin like
9
5
nucleus, cytoplasm
CD11b
Integrin alpha-M
7
5
plasma membrane
Copg1
coatomer protein complex subunit gamma 1
6
5
cytoplasmic vesicle coat
Cyc1
Cytochrome c1
10
5
nucleus, mitoch. inner membr.
Ddx39b
DExD-box helicase 39B
9
5
nucleus, cytopl.,spliceosome
Ddx5
DEAD-box helicase 5
11
7
nucleus, cytopl.,spliceosome
Dync1h1
dynein cytoplasmic 1 heavy chain 1
8
6
cytopl., cytoskel. (complex)
Eftud2
elongation factor Tu GTP binding domain containing 2
28
15
nucleus, spliceosome compl.
Eif3b
eukaryotic translation initiation factor 3 subunit B
9
5
cytoplasm (complex)
Eif4a2
Eukaryotic initiation factor 4A-II Eif4a2
13
6
nucleolus, cytoplasm
Eno1
enolase 1
17
8
raft, synaptic membrane
Etfa
electron transfer flavoprotein alpha subunit
10
6
mitochondrion (complex)
Etfdh
electron transfer flavoprotein dehydrogenase
8
5
mitochondrial membrane
Fgg
Fibrinogen gamma chain
7
5
cytopl,plasm.membr. (extern.)
Flna
Filamin-A
17
9
nucleus, cytosol, pl.membrane
Gimap4
GTPase IMAP family member 4
9
6
cytoplasm
Gpd2
glycerol phosphate dehydrogenase 2, mitochondrial
20
11
membrane (mitochondr.)
H2afx
H2A histone family member X
20
6
nucleus
H2afy
H2A histone family member Y
15
7
nucleus
H2-D1
histocompatibility 2, D region locus 1
18
8
ER, plasma membr., raft
Hadha
hydroxyacyl-CoA dehydrogenase, alpha subunit
43
20
mitochondrion (complex)
Hadhb
hydroxyacyl-CoA dehydrogenase, beta subunit
12
7
mitochondrial membrane
Hist1h1c
histone cluster 1 H1 family member c
13
6
nucleus
Hist1h1e
histone cluster 1 H1 family member e
11
5
nucleosome
Hist1h2bb
histone cluster 1 H2B family member b
84
5
nucleus, cytoplasm
Hnrnpk
heterogeneous nuclear ribonucleoprotein K
11
5
nucleus, spliceosome compl.
Hnrnpu
heterogeneous nuclear ribonucleoprotein U
15
7
mitotic spindle, spliceosome
Hsp90ab1
heat shock protein 90 alpha, class B member 1
30
16
plasma & vesicle membrane
Idh3a
isocitrate dehydrogenase 3 (NAD+) alpha
17
7
mitochondrion, myelin sheet
Krt8
Keratin 8
10
6
cytoskel., plasma membrane
Kpnb1
karyopherin (importin) beta 1
11
6
nuclear membr., ER network
Lman1
lectin, mannose binding 1
8
5
organelles (membrane bound)
Lmnb1
Lamin-B1 Lmnb1
10
6
nucl. membr., intermed.filam.
Msn
Moesin
7
5
plasma membrane (cytopl)
Mtco2
Cytochrome c oxidase subunit 2 Mtco2
16
4
mitochondrial inner membr.
Myh14
Myosin-14
14
7
stress fibers (complex)
Myh4
Myosin-4
156
72
cytoplasm, myosin filament
28
Pept.M
Table S6 cont. MDSC-Exo over LNC membrane
cellular localization
Synonym
Protein Description
Prot.M.
Pept.M
Myh9
Myosin-9
161
70
pl.membr.,signalos.,imm.synapse
Myl1
Myosin light chain 1/3, skeletal muscle isoform
21
8
myosin complex (fibres)
Mylpf
Myosin regulatory light chain 2, skeletal muscle isoform
10
6
lysosome complex
Ndufs1
NADH-ubiquinone oxidoreductase 75 kDa subunit
8
5
mitoch.inner membrane
Ogdh
oxoglutarate dehydrogenase
8
5
mitochondrion (complex)
Pcbp1
Poly(rC)-binding protein 1
14
7
nucleus, ribosome complex
Pcbp2
Poly(rC)-binding protein 2
9
5
nucleus, cytopl., membrane
Pdia6
protein disulfide isomerase family A member 6
6
5
ER (complex), pl.membr.
Pfn1
Profilin-1
5
3
cytoskeleton, membrane
Phgdh
phosphoglycerate dehydrogenase
7
5
myelin sheath
Pkm
Pyruvate kinase, muscle
19
10
cytopl.,mitochondr. (complex)
Pml
promyelocytic leukemia
8
5
nucleus, cytosol, ER membr.
Psmd1
proteasome 26S subunit, non-ATPase 1
9
6
proteasome complex
Psmd13
proteasome 26S subunit, non-ATPase 13
14
7
proteasome complex
Psmd2
proteasome 26S subunit, non-ATPase 2
9
6
proteasome complex
Psmd6
proteasome 26S subunit, non-ATPase 6
17
9
proteasome complex
Psme1
Proteasome activator subunit 1
9
5
proteasome complex
Ptprc
protein tyrosine phosphatase, receptor type C
6
5
plasma membr. (outer side)
Rack1
receptor for activated C kinase 1
14
7
cytopl., ribosome, pl.membr.
Rpl12
ribosomal protein L12
9
5
ribosome complex
Rpn1
ribophorin I
9
6
ER membrane
Sdha
succinate dehydrogenase complex flavoprotein subunit A
16
8
mitoch.inner membr.(complex)
Sf3b3
Splicing factor 3B subunit 3
15
8
spliceosome complex
Slc25a11
solute carrier family 25 member 11
4
3
mitochondrial inner membrane
Slc25a12
solute carrier family 25 member 12
18
11
mitochondr.inner membrane
Slc25a4
solute carrier family 25 member 4
15
8
mitoch.membr., cytoskel., raft
Sptb
spectrin beta, erythrocytic
71
32
cytoskel.,pl.membr., (complex)
Stat1
Signal transducer and activator of transcription 1
12
7
nucleus, cytoplasm (complex)
Tap2
transporter 2, ATP binding cassette subfamily B member
7
5
ER membrane (complex)
Tfrc
Transferrin receptor
15
9
endosome & plasma membr.
Tgtp1
T cell specific GTPase 1
10
4
ER membrane
Tln1
Talin-1
38
19
cytoskel.,plasma membrane
Tmed10
transmembrane p24 trafficking protein 10
9
5
organelle (membr., compl.)
Ucp1
uncoupling protein 1 (mitochondrial, proton carrier)
10
7
mitochondr.inner membrane
Vcp
valosin containing protein
13
9
proteas.compl., ER membr.
Vdac1
voltage-dependent anion channel 1
20
9
mitochonr. & plasma membrane
Vdac2
voltage-dependent anion channel 2
12
7
mitochonr. & plasma membr., raft
Vdac3
voltage-dependent anion channel 3
11
5
mitochondrial membrane
Vim
Vimentin
12
6
cytoskel., plasma membrane
Xpo1
Exportin-1
10
5
cytopl., organelle membr.
a
proteins with ≥3 peptide hits are shown
29
Table S7 In vivo impact of MDSC-Exo on AA spleen cells Table S7A Increased mRNA recovery in spleen cells of MDSC-Exo treated AA mice AA ExoSynonym SC treated Protein name Abca1 1595 10791 ATP-binding cassette sub-family A member 1 Abcg1 1370 5488 ATP-binding cassette sub-family G member 1 Adgre1 1664 7722 Adhesion G protein-coupled receptor E1 Agap1 316 1005 Arf-GAP with GTPase,ANK repeat & PH domain prot.1 Aldh1l2 238 1042 Mitochondrial 10-formyltetrahydrofolate dehydrogenase Als2cl 3427 12879 ALS2 C-terminal-like protein Angptl2 301 1478 Angiopoietin-related protein 2 Ankrd55 270 1993 Ankyrin repeat domain-containing protein 55 Arg1 14325 57553 Arginase-1 Atf3 1102 3326 Cyclic AMP-dependent transcription factor ATF-3 Atf5 2046 6599 Cyclic AMP-dendent transcription factor ATF-5 Atn1 3185 9859 Atrophin-1 Atp6v0d2 1684 6401 V-type proton ATPase subunit d2 C3ar1 1476 8124 C3a anaphylatoxin chemotactic receptor Cacna1d 329 1108 Voltage-dependent L-type calcium channel alpha-1D Cacnb1 284 1131 Voltage-dependent L-type calcium channel beta-1 Ccl2 1445 8996 C-C motif chemokine 2 Ccl7 262 2216 C-C motif chemokine 7 Ccnd1 299 1254 G1/S-specific cyclin D1 Ccr9 420 1790 C-C chemokine receptor type 9 Cd200 820 3180 OX-2 membrane protein Cd302 473 1423 CD302 antigen Cd36 2234 11144 Platelet glycoprotein 4 Cd83 2025 11032 CD83 antigen Cd86 656 2229 T-lymphocyte activation antigen Cdk18 422 1483 Cyclin-dependent kinase 18 Cdkn2a 279 1003 Tumor suppressor ARF Ch25h 123 1868 Cholesterol 25-hydroxylase Ckb 295 1416 Creatine kinase B-type Clec4a1 239 1181 C-type lectin domain family4 member a1 Clec4a2 361 1233 C-type lectin domain family4 member a2 Clec4d 1784 6243 C-type lectin domain family4 member d Clmp 308 1901 CXADR-like membrane protein Colec12 213 1060 Collectin-12 Csf1r 3625 10806 Macrophage colony stimulating factor 1 receptor Ctsk 509 8274 Catheppsin K Ctsl 29630 225943 Catheppsin L1 Cxcl1 265 1346 Growth-regulated alpha protein Cxcl16 1035 3221 C-X-C motif chemokine 16 Cxcl2 1539 5103 C-X-C motif chemokine 2
function transport, lipoprotein biosynthesis, LPS response transport, lipoprotein biosynthesis signal transduction, GPCR signaling ARP3 complex regulation oxidation-reduction GTPase endothelial growth factor protein-protein interaction hydrolase, ion binding transcription, gluconeogenesis chromatin binding, transcription factor transcription corepressor ion transport C3a binding, GPCR activ. ion chanel, ankyrin binding calcium transport cytokine activity, CCR2 binding, chemotaxis cytokine activity, CCR2 binding, chemotaxis transcription chemotaxis, chemokine / GPCR signaling protein binding, adhesion C-type lectin transmembrane receptor cytokine secretion humoral IR TLR & PI3K & BCR & FGFR signaling transferase cell cycle regulation oxidation-reduction kinase activity, IL6, IL12, TNF production , dendritic cell inhibitory receptor, inflamm. monocytes carbohydrate binding Ig receptor binding cell-cell adhesion pattern recognition receptor RTK, cytokine binding, migration peptidase, proteolysis peptidase, histone binding chemokine activ, CXCR binding chemokine & cytokine activity, CXCR binding chemokine & cytokine activity, CXCR binding
30
biological processesa lipoprotein metab, endosomal transport, phagocytosis lipoprotein metabolism, ABC family transport IR regulation membrane traffic, cytoskeleton organization vitamin metabolism endosome organization organism development suggested involvement in autoimmune diseases AA metabolism, T proliferation, IFN signaling TRAIL-activation, apoptotic signaling cell cycle regulation, proliferation, apoptosis CNS development, neuron apoptosis ROS/RNS production, endocytosis, transport chemotaxis, inflamm.IR, C' cascade, GPCR signaling transmembrane transport transmembrane transport, synaptic transmission IL1, IFN, TNF, TGFß, GPCR signaling GPCR signaling, IL1, IFN, TNF response cell cycle regul., Wnt signaling, DNA damage response IR regulation IR (lymphoid-nonlymphoid) regulation myeloid DC migration, phagocytosis antigen cross-present.,vesicl.-med.transport, TLR sign. CD4 different., IL2 & IL10 production CD28-dependent T cell stimulation phosphorylation cell growth (T & B cells) , NFB signaling lipid & lipoprotein metabolism, B cell chemotaxis innate IR, Myd88/TLR, NFB, IL1 signaling innate & adaptive IR FcR signaling, IR regulation phagocytosis, TLR signaling RTK, STAT, ERK1/2 signaling, innate IR, hemopoiesis TLR cascades ECM organiz, MHCII antig.present., hair follicle morph.. chemotaxis regulation, inflamm. IR, GPCR signaling T cell chemotaxis, TNF, IFN, GPCR signaling chemotaxis, inflammatory IR, GPCR signaling
Table S7A cont. AA ExoSynonym SC treated Protein name Cxcl3 1929 10842 C-X-C motif chemokine 3 Cxcl5 247 2461 C-X-C motif chemokine 5 Dab2 1152 3841 Disabled homolog 2 Dtx1 476 1729 E3 ubiquitin-prootein ligase DTX1 Dusp10 1832 8026 Dual specificity protein phosphatase 10 Ebf1 342 1128 Transcription factor COE1 Ephx1 783 2807 Epoxide hydrolase 1 Fam19a3 1563 5337 Protein FAM19A3 Fam20c 943 3954 Extracellular serine/threonine protein kinase FAM20C Flrt2 617 3655 Leucine-rich repeat transmembrane protein FLRT2 Foxp3 544 2328 Forkhead box protein P3 Fpr1 809 2453 fMet-Leu-Phe receptor Gja1 307 1080 Gap junction alpha-1 protein Gm15056 80 1496 Protein Gm15056 Gpnmb 22678 105624 Transmembrane glycoprotein nMB Hdac9 209 1557 Histone deacetylase 9 Hmox1 9634 49122 Heme oxygenase 1 Ifngr2 1310 4129 Ifngr2 protein Igf1 194 4505 Insulin-like growth factor 1 Igsf23 332 1321 Igsf23 protein Il1r1 301 1675 Interleukin-1 receptor type 1 Il6st 3286 10126 Interleukin6 receptor subunit beta Inadl 1109 4672 InaD-like protein Inhba 814 2779 Inhibin beta A chain Irak3 1023 3230 Interleukin-1 receptor-associated kinase 3 Jag1 289 1143 Protein jagged-1 Kcna2 544 2961 Potassium voltage-gated channel subfamily A 2 Lgmn 3430 10461 Legumain Lhfpl2 1198 4169 Lipoma HMGIC fusion partner-like 2 protein Lpl 1797 11860 Lipoprotein lipase Lrp1 2537 10001 Prolow-density lipoprotein receptor-related protein 1 Lrpap1 5607 18250 Alpha-2-macroglobulin receptor-assoc. protein Lrrc25 505 1584 Leucine-ricj repeat-cont. protein 25 Maf 902 2842 Transcription factor Maf Marcks 726 2765 Myristoylated alanine-rich C-kinase Marco 66 1218 Macrophage receptor Marco Mfge8 2382 22112 Lactadherin Mmp12 33388 179224 Matrix metalloproteinase 12 Mmp13 2876 32434 Matrix metalloproteinase 13 Mmp14 982 3534 Matrix metalloproteinase 14 Mmp9 1637 13706 Matrix metalloproteinase 9 Mrc1 930 5855 Macrophage mannose receptor 1 Msr1 3838 16063 Macrophage scavenger receptor types i and II
function chemokine & cytokine activity, CXCR binding chemokine activity, CXCR binding, chemotaxis AP2 complex binding, clathrin adaptive activity ion binding, transferase MAPK, JNK cascade transcription hydrolase, catalytic activity C-C chemokine family protein serine/threonine kinase protein kinase inhibitor transcription factor, IL2, IL5, IL10, IFN production chemotaxis, leukocyte migration channel activity, signaling beta defensin gene family receptor-ligand activity, TNF production transcription factor binding oxidation-reduction IL10 receptor activity receptor binding & signaling IR gene family IL-1 signaling signaling protein binding cytokine activ, receptor binding, cell cycle kinase activity, neg.regul.IL6, IL12, TNF product.,. regulation of proliferation ion channel activity peptidase, proteolysis tetraspanin family, platelet degranulation lipid metabolism, chemokine secretion endocytosis, transcytosis lipase binding binding, monocytes and plamacytoid dendtritc cells transcription regulation actin filament assembly endocytosis, apoptotic clearance adhesion, phagocytosis binding, proteolysis, virus IR regulation proteolysis peptidase proteolysis, leukocyte migration,collagen metabolism endocytosis, LPS response endocytosis, phagocytosis, cholesterol transport
31
biological processes chemotaxis, inflammatory IR, GPCR signaling cytokine production, inflamm.IR, GPCR, JAK/Stat sign. myeloid cell diff., endocytosis, vesicle-med. transport T cell differentiation, NOTCH signaling Treg differentiation, innate IR regulation adipocyte differentiation response to toxic substances microglia cell activation muscle development adhesion, migration, FGFR1 signaling T cell prolif.,chronic inflammation, Treg differentiation GPCR signaling Junction assembly, membr.trafficking, vesicle- transport T cell activation , ERK1/2 signaling transcript.regulation, inflamm. IR, B cell activation hypoxia resp., intrinsic apoptotic sign., IB-NFB sign. IR regulation, IFN signaling regulation IGF1R signaling, hemostasis, development IR regulation IL6, oncostatinM, NOTCH, IL27 signaling, T cell prolif. multimeric complex organization, TJ, signaling extrinsic apoptosis, SMAD phosphoryl., hair follicle innate IR, Myd88/TLR, NFB sign. , IL1 signaling NOTCH signaling ion transmembrane transport TLR cascade, MHCII antigen processing hemostasis lipid & lipoprotein metabolism actin organiz., GPCR signaling, lysosomal catabolism regulation endocytosis & cell death cell development, cytokine production energy metabolisms GPCR signaling, vesicle-mediated transport angiogenesis, phagocytosis, apoptotic cell clearance ECM organization, regulation IFN/ß signaling ECM disassembly regulation cell migration, proteolysis, ECM organization axon guidance, EPH-Ephrin signaling IL4 & IFN response Ligand uptake, vesicle-mediated transport
Table S7A cont. AA ExoSynonym SC treated Protein name Nrp2 2457 8104 Neuropilin Pacsin1 563 2113 PKC and casein kinase substrate in neurons protein1 Pcdh7 325 1321 Prootein Pcdh7 Pdlim4 156 1026 PDZ and LIM domain protein 4 Pdpn 1397 6282 Podoplanin Pid1 295 2137 PTB-containing, cubilin and LRP1-interacting protein Plau 1864 6029 Urokinase-type plasminogen activator Pltp 650 2297 Phospholipid transfer protein Pmepa1 746 2509 Protein TMEPAI Pou2f2 834 2612 POU domain, class 2, transcription factor 2 Ppap2b 536 1947 Phospholipid phosphatase 3 Pros1 1325 4996 Vitamin K-dependent protein S Ptges 330 1688 Prostagalandin E synthase Qsox1 5343 21041 Sulfhydryl oxidase 1 Rgl1 987 3411 Ral guanine nucleotide dissociation stimulator-like 1 Rin2 462 1406 Ras and Rab interactor 2 Rragd 208 1273 Ras-related GTP-binding protein D Rxra 657 2037 Retinoic acid receptor RXR-alpha Saa3 2181 13528 Serum amyloid A-3 protein Sash1 434 1884 SAM and SH3 domain containing protein 1 Sbf2 473 1538 Protein Sbf2 Sdc4 1157 5059 Syndecan 4 Serpinb8 241 1042 Serpin B8 Siglec1 184 1303 Sialoadhesin Slc15a3 1830 5825 Solute carroer family 15 member 3 Slc6a8 370 1314 Sodium- nd chloride-dependent creatine transporter 1 Slc7a2 6214 29655 Cationic amino acid transporter 2 Slfn5 2137 7068 Schlafen family member 5 Snx29 272 1148 Sortin nexin-29 Spib 1227 4146 Transcription factor Spi-B Spsb1 269 1736 SPRY domain-containing SOCS box Stab1 387 1270 Stabilin-1 Stac2 323 1025 SH3 and cystein-rich domain-containing protein2 Tgfbi 2818 12397 Transforming growth factor-beta-induced protein ig-h3 Timp2 638 2240 Metalloproteinase inhibitor 2 Tlr8 294 1240 Toll-like recptor 8 Tmie 278 1580 Transmembrane inner ear expressed proteii Tnfrsf4 889 2817 Tumor necrosis factor receptor superfamily member 4 Tnfsf8 870 3307 Tumor necrosis factor ligand superfamily member 8 Trat1 238 2001 T-cell receptor-associated transmembrane adaptor 1 Trem2 602 3204 Triggering receptor expressed on myeloid cells 2 Vcan 1087 5151 Versican core protein Wfdc17 1545 5877 Activated macrophage/microglia WAP domain protein
function protein binding, migration, chemotaxis phospholipid & cytoskeleton binding protein binding actin cytoskeleton organization binding response to IL6 & TNF peptidase lipid transporter R-SMAD binding regulation of transcription cell-cell adhesion, regul.phosphorylation phagocytosis, negative regulation coagulation lipid metabolism oxidation-reduction GTPase signal transduction GTPase activator, GTPase regulation of transcription chemotaxis, response to IL-1 protein complex scaffold activity phosphatase regulator fibronectin binding, thrombospondin receptor activity peptidase inhibitor endocytosis, extrinsic apoptosis signaling peptide, protein transport ion transport AA transport, NO production, Mf activation ATP binding phosphatidylinositol binding transcription ubiquitin transferase scavenger receptor ion binding, intracellular signaling adhesion cytokine response, proteolysis IL1ß, IL6, IL8, IFN/ß regulation/secretion, inflamm.IR regulation IR & AICD CD8 T cell differentiation, response to bacterium calcium-mediated signaling transmembrane signaling receptor, adhesion endopeptidase inhibitor
32
biological processes LIF response, VEGFR signaling endocytosis, actin filament organization platelet activation & signaling bone marrow development cell-cell adhesion, morphogenesis, platelet activation regulation of ROS response regulation proliferation, migration, hemostasis lipid metabolism & transport SMAD & TGFßR signaling B cell maturation lipid & lipoprotein metabolism, Wnt signaling clotting cascade prostaglandin synthesis hemostasis regulation of GTPase activity & lipid metabolism endocytosis, signaling PI3K & TOR signaling impact on organ development TLR signaling LPS & JUN & NFB & p38MAPK signaling membrane organization, myelination ECM organiz., focal adhesion, Exo assembly & secret. cell-cell adhesion, platelet activation T cell apoptotic process transmembrane transport amino acid metabolism transmembrane transport, inflammatory IR regulation cell differentiation M differentiation MHCI antigen process./present., adaptive IS response inflammatory IR, vesicle-mediated transport ion channel inactivation ECM organization, angiogenesis ECM organization,regul. Ras, cAMP, MAPK signaling MyD88-TLR, IB-NFB, sign., MAPK, TLR7/8,9 activ. sensory hair development inner ear cytokine signaling, TNFR2-NFB signaling IR receptor recycling, T cell receptor signaling IR (lymph.-nonlymph.), CCR7, ERK1/2, CD40 signaling GAG synthesis, ECM organization .
Table S7B Reduced mRNA recovery in spleen cells of MDSC-Exo treated AA mice AA SC Synonym Adamts14 Add2 Alox12 Angpt1 Aqp1 Arsb Atp1b2 B4galnt2 Bzrap1 Camp Capn5 Car1 Car2 Casp12 Ccnb2 Cd160 Cd177 Cd244 Cd34 Ces2g Chil1 Clca3a1 Clec12a Clnk Clstn3 Cpa3 Ctla2a Ctla2b Dixdc1 Dsc2 Elane Epx Ermap Esco2 Fads2 Fam64a Fcrl6 Fdps Gfi1b Gp1ba Gp5 Gp9
2537 2158 3067 1447 4415 5461 3492 2486 1884 1519 3836 9846 10370 2002 3305 1718 3385 3150 4550 2691 1095 1379 1465 1719 1407 43212 13192 1894 1248 2697 14228 5245 1811 1810 1463 1393 3671 1967 1565 2834 1396 1090
Exotreated 356 44 445 217 183 969 106 298 295 43 702 404 2065 204 620 240 99 515 739 327 193 226 260 321 185 6709 2223 221 205 57 162 186 49 336 136 278 45 289 251 520 253 126
Protein name Adamts14 Beta-adducin Arachidonate 12-lipoxygenase 12S Angiopoietin-1 Aquaporin-1 Arylsulfatase B Sodium/potassium-transport ATPase subunit ß2 Beta-1,4 N-acetylgalactosaminyltransferase 2 Peripheral-type benzodiazepine rec.-assoc. protein1 Cathelicidin antimicrob. Peptide Calpain 5 Carbonic anhydrase 1 Carbonic anhydrase 2 Caspase 12 G2/mitotic-specific cyclin-B2 CD160 antigen CD177 NK receptor 2B4 Hematopoieitc progenitor cell antigen CD34 Carboxylic ester hydrolase Chitinase-3-like protein-1 Calcium-sens. chloride conductane protein1 C-type lectin domain fam.12memberA Cytokine-dep. hematopoietic cell linker Calsystenin-3 Mast cell carboxypeptidase A Protein CTLA-2a CTLA-2b Dixin Desmocollin-2 Neutrophil elastase Eosinophil peroxidase Erythroid membr.-assoc. protein N-acetyltransferase Fatty acid desaturase 2 Protein FAM64A Fc receptor-like protein6 Farnesyl pyrophosphate synthase Zinc finger protein Gfi-1b Platelet glycoprotein Ib- Platelet glycoprotein V Platelet glycoprotein IX
function peptidase actin & protein kinase binding oxidoreductase activity angiogenesis transporter hydrolase ATPase activity glycosylation protein binding antimicrobial peptide protease inhibition carbon metabolism carbon metabolism protease cyclin-dependent serine/threonine kinase activity MHCI receptor activity fibrin clot formation NK & mDC activation binding oxidation glycosidase peptidase, ion channel signaling transmembrane receptor ion binding proteolysis peptidase peptidase inhibitor enzyme modulator, signaling ion binding inflammatory response, phagocytosis, proteolysis defense to nematode erythroid membrane receptor DNA replication oxidorectuctase controls metaphase to anaphase transition MHCII ligand acyltransferase regul.transcription coagulation collagen binding adhesion
33
biological processes ECM organization transmembrane transport, leukocyte migration lipid & lipoprotein metabolism multiple signaling pathways transmembrane transport autophagy ion transport negative regulation adhesion synapse formation innate IR, chemotaxis proteolysis O2/CO2 exchange (erythrocytes) chloride transport intrinsic apoptosis signaling cell cycle, T cell homeostasis IR (bacterium) clotting cascade hemostasis hemopoiesis, TGFß&IL10 production, leukoc. migr. Inflamm. IR, apoptosis, PKB & NFB&ERK1/2 activ. ion transport immune response regulation tyrosine kinase signaling, IR synapse assembly and transmission peptide hormone metabolism regulation Treg differentiation Treg conversion microtubule organization, JNK cascade cell adhesion ECM organization & degradation oxidative stress response cell cycle lipid & lipoprotein metabolism cell cycle CTL & NK receptor, upregulated in chronic imm.stim. lipid & lipoprotein metabolism hemopoiesis clotting cascade clotting cascade clotting cascade
Table S7B cont. AA SC ExoSynonym treated Protein name Gucy1a3 1838 321 Guanylate cyclase soluble subunit 3 Gucy1b3 1844 341 Guanylate cyclase soluble subunit 1 Gzma 91251 5036 Granzyme A Gzmb 149194 12697 Granzyme B Gzmc 43167 1693 Granzyme C Gzmd 75163 898 Granzyme D Gzme 28887 345 Granzyme E Gzmf 26081 359 Granzyme F Gzmg 96614 1092 Granzyme G Havcr2 8662 649 Hepatatis A virus receptor 2 Hmgcs1 10861 1759 Hydroxymeth.-CoA synthase Ifitm1 52676 10125 IFN-induced transmembrane protein Ifng 1637 255 Interferon Itga2b 23212 4422 Integrin -Ib Kalrn 1263 171 Kalirin Klrg1 1073 57 Killer cell lectin-like receptor G1 Lipg 4652 620 Endothelial lipase Ltbp1 1839 206 Latent transforming growth factor ß-binding protein 1 Ltf 4922 181 Lactotransferrin Ly6c1 6129 768 Lymphocyte antigen 6C1 Ly6c2 9867 1265 Lymphocyte antigen 6C2 Maob 1138 149 Amine oxidase B Mcpt8 18692 2308 Mast cell protease Mfsd2b 3503 511 Major facilitator superfam.dom.-contain.protein2B Mns1 1283 140 Meiosis-specific nuclear struct.protein 1 Mpl 1661 229 Thrombopoietic receptor Mpo 23160 789 Myeloperoxidase Mrvi1 5207 768 Mrvi1 Ms4a2 7186 1234 High affinity Igepsilon R subunit ß Ms4a3 2383 80 Membr.-spanning 4 domain subfamily A3 Myb 5422 809 Transcriptional activator Myb Mycn 1669 268 N-myc proto-oncogene protein Myl10 1050 180 Myosin regulatory light chain 10 Mylk 3936 642 Myosin light chain kinase Ngp 31130 394 Neutrophilic granule protein Nkg7 17727 2786 Protein NKG7 Nlrp6 5308 912 NACHT,LRR & PYD dom. containing prot.6 Nqo1 1689 276 NAD(P)H dehydrogenase 1 Nrgn 1471 206 Neurogranin Pkd2l1 1615 298 Polycystic kidney disease 2-like 1 Podxl 5715 920 Podocalyxin Prf1 39787 2235 Perforin Prg2 3593 335 Bone marrow proteoglycan
function cGMP biosynthesis cGMP biosynthesis, Hsp90 binding peptidase peptidase peptidase peptidase peptidase peptidase peptidase adaptive and innate IR metabolism response to virus regulation of cytokine production leukocyte migration, platelet aggregation guanyl exchange factor innate IR phospholipase TGFß binding protease, ROS/RNS production monocyte marker monocyte marker oxidation-reduction proteolysis enzyme activity protein binding cytokine receptor activity oxidation-reduction process regulation of smooth muscle contraction FcRI signaling CD20L,cell cycle regulation transcription factor transcription cytoskeletal protein actin binding peptidase inhibition cytokeletal protein transcription cofactor regulation of oxidate and apoptotic processes synaptic potentiol ion transport protein binding wide pore formation component of eosinophil granule
34
biological processes hemostasis NO-mediated signaling proteolysis, cytolysis cytolysis, T cell cytotoxicity proteolysis, cytolysis proteolysis, cytolysis proteolysis, cytolysis proteolysis, cytolysis proteolysis cytokine signaling lipid & lipoprotein metabolism immune regulation (nonlymphoid-lymphoid) cytokine signaling in the IS hemostasis, multiple signaling pathways MAPK & Rho & GPCR signaling, death signaling Immunoregulation (lymphoid-nonlymphoid) lipid & lipoprotein metabolism ECM organization, TFGßR signaling regulation cytokine production, innate IR inflammatory process orchestration inflammatory process orchestration biological oxidation inflammatory respone in the skin transport cilium organization regulation of hemopoiesis and homeostasis azurophilic granules, innate IR cGMP signaling MAPK & NFB activation, inflammatory IR IS signal transduction, neutrophil degranulation TH & B & myeloid cell differentiation regulation cell death regulation focal adhesion, leukocyte migration regul. migration & transport, RhoGTPase signaling regulation defense response adhesion, transporter IR , TLR&MAPK&IB/NFB signaling oxidation-reduction process intracellular signaling channel activity regulation leukocyte adhesion & migration channel activity, apoptosis, cytolysis IR to nematode & bacterium
Table S7B cont. AA SC Synonym Prss34 6940 Prtn3 5128 Rab27b 3767 Rab38 4129 Rhag 1136 S100a4 9322 S100a6 14291 S100a8 37682 S100a9 57146 Samd14 1645 Scin 14700 Sdpr 1753 Serpinb9b 3296 Sh2d5 1812 Slc6a4 1510 Spns2 2092 Spta1 1842 Sytl2 3530 Tfr2 1478 Timp3 6914 Tjp1 1380 Tmem119 4546 Tmem40 1096 Treml1 1496 Tubb1 1651 Vash1 3055 Vcam1 1055 Vwf 13563 Zbtb32 5045 a
Exotreated 934 168 698 787 110 692 2415 4415 4653 304 2765 268 376 123 259 317 66 462 240 995 211 467 104 255 154 345 39 2121 816
Protein name Mast cell protease 11 Myeloblastin Ras-related protein Rab-27B Ras-relat. protein Rab-38 Ammonium transporter Rh type A Protein S100-A4 Protein S100-A6 Protein S100-A8 Protein S100-A9 Sterile alpha motic dom.cont. protein 14 Adseverin Serum-depriv. response protein Serpinb9b SH2 domain-contain.protein 5 Sodium-depend. serotonin transporter Protein spinster homolog 2 Spectrin chain,erythrocytic-1 Synaptogamin-like protein2 Transferrin receptor protein2 Metalloproteinase inhibitor 3 TJ protein ZO1 Transmembrane protein 119 Transmembrane protein 40 Trem-like transcript 1 protein Tubulin 1 chain Vasohibin1 Vascular cell adhesion protein1 vonWillebrand factor Zinc finger and BTB domain-cont. protein 32
function peptide crosslinking & proteolysis protease inhibition GTPase activity GTPase activity iron homeostasis actin binding calcium and protein binding neutrophil chemotaxis, apoptosis chemotaxis, inflamm.response, apoptosis aberrant promoter methylation actin binding PKC substrate, phosphatidyl binding cytolysis protein binding transporter lipid transport actin organization membrane traffic regulator regulation iron ion homeostasis protease inhibition GAP junction formation CD11b+ cells in microglia Ig receptor GTPase protein binding integrin binding blood coagulation transcription corepressor
biological processes basophil mediated tissue damage proteolysis, phagocytosis Exo sorting & exocytosis organelle organization & transport transmembrane transport IB/NFB signaling ion transport, plasma membrane tubulation inflammatory response TLR cascade actin nucleation, apoptosis regulation plasma membrane tabulation protection from cell-mediated cytotoxicity synapse transmission B & T cell homeostasis, lymphocyte migration regul.hemopoiesis, T cell prolif., multiple signaling vesicle docking & exocytosis regulation of endocytosis regulation proteolysis, TRAIL signaling pathway membrane & vesicle-mediated transport osteoblast differentiation, spermatogenesis biological process lymphoid-nonlymph. interaction, neg.regul. signaling organelle biog., membr.traff., vesicle-med. transport angiogenesis regulation, cellular sensecence IR regulation (lymphoid-nonlymphoid) clotting cascade, beta3 signaling osteoblast different.,hematopoietic prog. proliferation
AA: amino acid, AICD: activation-induced cell death, DC: dendritic cells, ECM: extracellular matrix, IS: immune system, IR: immune response, TJ: tight junction, : reduced recovery / activity, : pronounced recovery / activity, red: engagement in hair follicle organization
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Figure S1 Proteome analysis of ex vivo and in vitro generated MDCS and MDSC-exosomes. (A) MDSC were either isolated from SC by magnetic bead sorting for CD11b+Gr1+ cells or were generated in vitro from BMC cultured in the presence of GMCSF, IL6 and PGE2. After SDS-Page and digestion, proteins were identified by mass spectrometry. Proteins preferentially recovered in ex vivo or in vitro generated MDSC were clustered using Reactome according to their engagement in immune response, cell cycle, transport and cytoskeleton organization, proteolysis / apoptosis and signaling. Proteins engaged in two clusters are identified by a distinct frame color. (B,C) Exo were collected from MDSC as described above after 2d culture in medium with 3% Exo-depleted FCS. After purification, SDS-Page and digestion, proteins were identified by mass spectrometry. (B) Comparison of abundantly recovered proteins in MDSC-Exo derived from ex vivo versus in vitro generated MDSC including proteins that are selectively recovered from the two distinct MDSC preparations and (C) MDSC-Exo preferentially recovered from ex vivo or in vitro generated MDSC were clustered according to major functional activities as described for MDSC. A proteome analysis confirmed no significant differences in ex vivo versus in vitro generated MDSC according to immune system-related functions. The abundance of distinctly in in vitro generated MDSC relying mostly on pronounced cell division and associated metabolic processes due to the culture condition. This also accounts for the differential recovery in Exo from ex vivo or in vitro MDSC, where the significant increase in proteins engaged in cell division and metabolic processes is also reflected in Exo from culture-derived MDSC.
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Figure S2 Proteome analysis comparing ex vivo and in vitro generated MDCS with MDSC-exosomes. MDSC were generated in vitro from BMC. After maturation, MDCS were cultured for 2d in medium with 3% FCS that was Exo-depleted. MDSC and MDSC-Exo proteins were identified by mass spectrometry. (A) Panther Gene Analysis of molecular functions of in vitro and ex vivo MDSC versus MDSC-Exo; (B) Panther Gene Analysis of molecular functions of proteins that are distinctly recovered in in vitro and ex vivo MDSC versus MDSC-Exo. (C) Reactome analysis of distinctly recovered proteins in BMC-derived MDSC versus MDSC-Exo, which are engaged in immune response regulation including activation of signaling cascades. Three proteins engaged in MDSC activity, Arg1, Sod2 and Bax were recovered at a lower level in MDSC-Exo. On the other hand, tetraspanins, C’ and proteasome subunits were more abundant in MDSC-Exo than MDSC. Hypothesizing that pro and contra of function-relevant proteins in MDSC versus MDCS-Exo could possibly be balanced, we proceeded comparing their effectivity in immune response regulation.
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Figure S3 MDSC-exosome membrane attached proteins. A lymphocyte membrane preparation was coupled to Sepharose. MDSC-Exo were lysed (mild conditions, Lubrol) and were passed over the lymphocyte-membrane Sepharose column. After washing, Exo lysates were eluted, dialyzed, concentrated and subjected to SDS-PAGE for proteome analysis. (A) Pulled-down MDSC-Exo proteins were clustered according to molecular functions (Panther Gene Analysis) and (B) using Reactome according to proteins engaged in Exo biogenesis, transport, cytoskeleton organization, engagement in innate and adaptive immune responses as well as lipid metabolism. Proteins engaged in signal transduction are framed in red. The pulldown of MDSC-Exo indicated that invaginated membrane domains remain intact during Exo biogenesis including attached cytoskeletal and signaling molecules. The latter may well contribute to immune target cell modulation.
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Figure S4 Examples of hair regrowth in MDSC exosome-treated mice with AA partialis and incipient. (A) Hair loss progression in untreated mice with AA partialis and partial hair growth recovery in MDSC-Exo treated mice is demonstrated after 2wk, 6wk and 10wk (3 mice / group); (B) Hair loss progression in untreated mice with AA incipient 16wk after initiating MDSC-Exo application; hair growth in MDSC-Exo treated mice is shown after 2wk, 6wk, 10wk and 16wk. MDSC-Exo treatment is curative when started at a very early stage of hair loss. At an intermediate state it prevents progression. Partial hair regrowth proceeds from remaining nonaffected areas.
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