Supplementary figures and tables

E g r 1 F g f2 F g f7

1 0 * 5 1 0

* *

e e

e *

g g

g *

n n

n *

a a

a 8 4 * 8

h h

h *

c c c

d d

d *

l l l

o o

o *

f f f

*

n n n

o o

o 6 3 6

i i i

s s s

s s s

e e e

r r r

p p p

x x x

e e e

4 2 4

e e e

n n n

e e e

g g g

e e e

v v v

i i i

t t

t 2 1 2

a a a

l l l

e e e

R R R

0 0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

J a k 2 N o tc h 2 H if1 

* 3 4 6

* * *

e e

e

g g

g

n n

n

a a

* * a * h

h *

h

c c

c

3 *

d d

* d

l l

l *

o o

o

f f

2 f

4

n n

n

o o

o

i i

i

s s

s

s s

s

e e

e r

r 2

r

p p

p

x x

x

e e

e

e e

e

n n

n e e 1

e 2

g g

g

e e

1 e

v v

v

i i

i

t t

t

a a

a

l l

l

e e

e

R R R

0 0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

Z e b 2 C d h 1 S n a i1 * * 7 1 .5 4 *

*

e e e

g g

g 6

n n

n *

a a

a *

h h h

c c c

3 *

d d d

l l

l 5

o o o

f f

f 1 .0 *

n n

n *

o o o

i i

i *

s s

s 4

s s s

e e e

r r

r 2

p p p

x x

x 3

e e e

e e e

n n n

e e

e 0 .5

g g g

2

e e

e 1

v v v

i i i

t t t

a a

a *

l l l

e e

e 1 *

R R R

0 0 .0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

M m p 9 L o x V im

2 0 0 2 0 8 *

* *

e e

e *

g g

g 1 5 0 *

n n

n *

a a

a *

h h

h *

c c c

1 5 * 6

d d d

l l

l 1 0 0

o o o

f f f

n n n

o o o

i i

i 5 0

s s s

s s

s *

e e e

r r r 1 0 4

3 0

p p

p *

x x x

e e e

*

e e e

n n n

e e

e 2 0

g g g

e e

e 5 2

v v v

i i i

t t t

a a a

l l

l 1 0

e e e

R R R

0 0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

Supplementary Figure 1. Validation of RNA-Seq results by the comparison to our previous QPCR results related to epithelial-to-mesenchymal transition [13, 15]. The relative gene expression of Egr1, Fgf2, Fgf7, Jak2, Notch2, Hif1α, Zeb2, Cdh1, Snai1, Mmp9, Lox and Vim is shown from control (left columns), uninflamed (middle columns) and inflamed (right columns) rat colon sections. White and black columns represent RNA-Seq and previously published QPCR data, respectively. Data are presented as the mean ± SEM; *p < 0.05.

1

Supplementary table 1 List of significantly activated canonical pathways in the in vivo rat model of IBD based on p-value, calculated by Ingenuity Pathway Analysis (IPA) application. The abbreviations “UI—C,” “I—C” and “I—UI” represent the TNBS treated—uninflamed vs. control, TNBS-treated—inflamed vs. control and TNBS treated—inflamed vs. TNBS treated—uninflamed comparisons.

2

Supplementary Table 2. List of significantly activated canonical pathways in the in vivo rat model of IBD based on z-score, calculated by IPA application. Abbreviations as “UI—C,” “I—C” and “I—UI” represent the TNBS treated—uninflamed vs. control, TNBS-treated—inflamed vs. control and TNBS treated—inflamed vs. TNBS treated—uninflamed comparisons.

3

Supplementary Figure 2. Expression pattern of NOD-like receptors in experimentally induced colitis. Heat map shows gene expression alteration of NLR receptors between control, TNBS- treated uninflamed and TNBS-treated inflamed rat colon samples, where red marks elevation, while blue marks decrease by at least two-fold in expression level; false discovery rate (FDR) < 0.05. Bar plots show the expression alteration of Nod2, Nlrp3, Nlrp6 and Nlrc4 between rat sample groups; white and black columns represent RNA-Seq and QPCR data, respectively, *p < 0.05. Abbreviations as “UI—C,” “I—C” and “I—UI” represent the TNBS treated—uninflamed vs. control, TNBS-treated—inflamed vs. control and TNBS treated—inflamed vs. TNBS treated—uninflamed comparisons.

4

F u t1 F u t2

* 1 .5 4 *

* e

e *

g

g

n

n a

a *

h

h

c

c

3

d

d

l

l

o

o f

1 .0 f

n

n

o

o

i

i

s

s

s

s

e

e r

r 2

p

p

x

x

e

e

e

e

n

n e

0 .5 e

g

g

e

e 1

v

v

i

i

t

t

a

a l

* l e

e * R * R

0 .0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

F u t4 F u t9 * *

3 * 2 5 * e

e * g

g *

n

n a

a 2 0

h

h

c

c

d

d l

l *

o

o

f f

2

n

n 1 5

o

o

i

i

s

s

s

s

e

e r

r 3

p

p

x

x

e

e

e

e *

n

n e

e 1 2

g

g

e

e

v

v

i

i

t

t

a

a l

l 1

e

e

R R

0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

Supplementary Figure 3. Diverse expression pattern of FUTs at the site of colon inflammation in rat experimental colitis. The relative gene expression of Fut1, Fut2, Fut4 and Fut9 is shown from control (left columns), uninflamed (middle columns) and inflamed (right columns) rat colon sections. White and black columns represent RNA-Seq and QPCR data, respectively. Data are presented as the mean ± SEM; *p < 0.05.

5

Il1  Il6 Il1 0

4 0 0 2 0 1 2 0 *

3 5 0

e e

e *

g g

g *

n n

n 1 0 0

a a

a 3 0 0 h h *

h *

c c

c

1 5

d d

d l

l 2 5 0

l

o o

o

f f

f 8 0

n n

n 2 0 0

o o

o

i i

i

s s

s

s s

s e

e 1 5 0

e r r 1 0

r 6 0 3 0 *

p p

p *

x x

x

e e

e

e e

e * *

n n

n

e e

e 4 0 2 0

g g

g

*

e e

e 5

v v

v

i i

i

t t

t

a a

a

l l

l 2 0 1 0

e e

e

R R R

0 0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

C x c l1 T n f T n fa ip 6

5 0 8 * 1 0 * *

*

e e

e *

g g

g *

n n

n *

a a

a 4 0 8

h h

h *

c c

c *

6

d d d

l l

l *

o o o

f f f

n n n

o o

o 3 0 6 *

i i i

s s s

s s s

e e

e *

r r

r 4

p p p

x x x

e e e

2 0 4

e e e

n n n

e e e

g g g

e e

e 2

v v v

i i i

t t

t 1 0 2

a a a

l l l

e e e

R R R

0 0 0

c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d c o n tro l u n in fla m e d in fla m e d

Supplementary Figure 4. Synchronous upregulation of inflammatory cytokines in the involved colon tissues. The relative gene expression of Il1β, Il6, Il10, Cxcl1, Tnf and Tnfaip6 is shown from control (left columns), uninflamed (middle columns) and inflamed (right columns) rat colon sections. White and black columns represent RNA-Seq and QPCR data, respectively. Data are presented as the mean ± SEM; *p < 0.05.

6

Supplementary Figure 5. Canonical pathway of Agranulocyte adhesion and diapedesis in the TNBS induced rat model of IBD. A. Heat map representing expression changes of genes belonging to the “Agranulocyte adhesion and diapedesis” pathway between control, TNBS treated uninflamed and TNBS treated inflamed rat colon samples, where red marks elevation, while blue marks decrease by at least a two-fold in expression level; FDR p < 0.05. Grey background refers no significant alterations. B. Visualizing “Agranulocyte adhesion and diapedesis” pathway in the comparison of TNBS treated inflamed to control samples. Red signs show upregulated genes while green signs represent downregulated genes. Genes with no significant changes labelled with grey background, while white signs missing genes from the RNA-Seq analysis. Abbreviations as “UI—C,” “I—C” and “I—UI”

7

represents the TNBS treated—uninflamed vs. control, TNBS-treated—inflamed vs. control and TNBS treated—inflamed vs. TNBS treated—uninflamed comparisons.

8

Supplementary Table 3 Summary of genes used for the determination of immune and pathway signatures of in vivo model of IBD by ImSig.

B cells Interferon Macrophages Monocytes Neutrophils NK cells Plasma cells Proliferation T cells Translation Total genes in ImSig 37 66 78 37 47 20 14 99 85 86 Total genes in ImSig (with Rat homologs) 33 59 73 35 43 10 4 98 78 84 No. of ImSig genes in user dataset 31 54 70 30 38 9 4 90 70 67 No. of ImSig genes after feature selection 26 48 65 26 36 0 2 84 67 61 Median correlation of ImSig genes 0,93 0,34 0,76 0,87 0,78 0,13 0,56 0,75 0,76 0,51 Median correlation of feature selected ImSig genes 0,94 0,4 0,8 0,89 0,79 NA 0,86 0,77 0,79 0,56 feature selection at r = 0.7 Supplementary Table 4. List of human and rat gene IDs used for gene signature analysis by ImSig (Part 1).

B cells Monocytes Neutrophils NK cells Plasma cells human ID rat ID human ID rat ID human ID rat ID human ID rat ID human ID rat ID BANK1 Bank1 PILRA LOC100910669 THBD Thbd KIR3DL2 Zbtb21 TXNDC5 Txndc5 HLA-DOB RT1-DOb PSAP Psap FPR1 Fpr1 KIR3DL1 Mroh1 TNFRSF17 Tnfrsf17 CD72 Cd72 LILRB2 BCL6 Bcl6 KIR2DS2 MZB1 Mzb1 TLR10 Tlr10 HCK Hck TLR2 Tlr2 KIR2DS1 IGLV1-44 CD19 Cd19 THEMIS2 Themis2 TNFRSF1A Tnfrsf1a KIR2DS3 IGH TCL1A Tcl1a EMILIN2 Emilin2 RNF149 Rnf149 KIR2DL2 IGJ Jchain MS4A1 Ms4a1 LILRB3 Lilrb3b PLXNC1 Plxnc1 KIR2DL5A IGKC STAP1 Stap1 FCN1 Fcnb CFLAR Cflar KIR2DS5 IGHG3 BTLA Btla KIAA0930 RGD1304694 PREX1 Prex1 TBX21 Tbx21 GUSBP11 CR2 Cr2 TPP1 Tpp1 DYSF Dysf KIR2DL1 IGKV1D-13 FCRL2 Fcrl2 LST1 Lst1 S100A9 S100a9 KIR2DL4 IGLV@ CD180 Cd180 AIF1 Aif1 S100A8 S100a8 KIR3DL3 Kir3dl1 IGLJ3 P2RX5 P2rx5 LILRA6 LOC690948 CSF3R Csf3r KIR2DL3 IGLC1 VPREB3 Vpreb3 CD93 Cd93 RALB Ralb PRF1 Prf1 IGLL3P FCRL1 Fcrl1 RHOG Rhog KIAA0247 Susd6 KLRC3 Vom1r81 FCRL3 Fcrl5 CD300LF Cd300lf STAT3 Stat3 SH2D1B Sh2d1b FCRLA Fcrla FGR Fgr PHC2 Phc2 KLRC2 Klrc2 CD79B Cd79b CD14 Cd14 IL17RA Il17ra SAMD3 Samd3 SNX22 Snx22 TIMP2 Timp2 GPR97 Adgrg3 KLRC4 IGHV5-78 PLXDC2 Plxdc2 MOB3A Mob3a KLRD1 Klrd1 FAM129C Niban3 CTSD Ctsd CEP19 Cep19 FCRL5 Fcrl5 HMOX1 Hmox1 SLC25A37 Slc25a37 LOC100507616 C10orf54 Vsir LILRA2 CD79A Cd79a SLC7A7 Slc7a7 PHF21A Phf21a CCR6 Ccr6 FES Fes NAMPT Nampt LY9 Ly9 GNS LOC100909505 SNORD89 LINC00926 VCAN Vcan NCF4 Ncf4 CD37 Cd37 TGFBI Tgfbi TLR4 Tlr4 KIAA0125 NFAM1 Nfam1 GLT1D1 Glt1d1 PNOC Pnoc PRAM1 Pram1 DENND5A Dennd5a CD22 Cd22 LRRC25 Lrrc25 ACSL1 Acsl1 PAX5 Pax5 GRN Grn BASP1 LOC100910172 AFF3 Aff3 SERPINA1 Serpina1 PADI2 Padi2 POU2F2 Pou2f2 PYCARD Pycard LIMK2 Limk2 S1PR4 S1pr4 CD33 KCNJ2 Kcnj2 BLK Blk AGTRAP Agtrap ALPK1 Alpk1 EBF1 Ebf1 NOTCH2 Notch2 CD97 Adgre5 SSH2 Ssh2 MGAM LOC679818 STAT5B Stat5b IFITM2 Ifitm2 CXCR2 Cxcr2 FAM65B Ripor2 LINC01002 AQP9 Aqp9 FCGR2C TMEM154 Tmem154

Supplementary Table 5. List of human and rat gene IDs used for gene signature analysis by ImSig (Part 2).

9

Macrophages T cells Proliferation Interferon Translation human ID rat ID human ID rat ID human ID rat ID human ID rat ID human ID rat ID CECR1 GIMAP4 Gimap4 GINS2 Gins2 STAT2 Stat2 RPS3A LOC100365839 CTSB Ctsb CD2 Cd2 UHRF1 Uhrf1 GBP4 Gbp4 FAU LOC687780 HLA-DRB6 ARHGAP9 Arhgap9 ZWILCH Zwilch IRF7 Irf7 RPS23 Rps23 FCGR2A LOC498276 IL23A Il23a FANCI Fanci IFI44L Ifi44l RPL18A Rpl18a TNFSF13B Tnfsf13b CD48 Cd48 RAD51AP1 Rad51ap1 ISG15 Isg15 EIF3F Eif3f SLAMF8 Slamf8 RASSF5 Rassf5 SMC2 Smc2 SP110 Sp110 RPL24 Rpl24 IFI30 Ifi30 CD52 Cd52 GINS1 Gins1 IFI44 Ifi44 RPS15A Rps15a CCR1 Ccr1 ARHGAP25 Arhgap25 NCAPG2 Ncapg2 XAF1 Xaf1 SNHG8 CD163 Cd163 TBC1D10C Tbc1d10c RAD51 Rad51 STAT1 Stat1 RPL11 Rpl11 ITGB2 Itgb2 NLRC3 Nlrc3 NDC80 Ndc80 DTX3L Dtx3l RPL15 Rpl15 C1QB C1qb C1orf162 LOC100911379 MCM6 Mcm6 OAS3 Oas3 PFDN5 Pfdn5 C3AR1 C3ar1 SP140 Sp140 DTL Dtl MX1 Mx2 EIF3E Eif3el1 FCER1G Fcer1g GPR18 Gpr18 HMMR Hmmr SAMD9L Samd9l RPS25 Rps25 TYROBP Tyrobp HCST Hcst EZH2 Ezh2 IFIH1 Ifih1 RPL31 Rpl31 TNFAIP2 Tnfaip2 RHOH Rhoh KIAA0101 Pclaf PARP9 Parp9 RPL18 Rpl18 SLC15A3 Slc15a3 GZMK Gzmk MAD2L1 Mad2l1 IFIT3 Ifit3 EEF1B2 Eef1b2 CD74 Cd74 CORO1A Coro1a PBK Pbk TRAFD1 Trafd1 RPL5 Rpl5 FCGR3B Fcgr3a ITGAL Itgal CDK1 Cdk1 C5orf56 RPS9 Rps9 CLEC7A Clec7a GIMAP7 Gimap7 TOP2A Top2a BST2 Bst2 RPL10 Rpl10 TRPV2 Trpv2 TRAC CCNB1 Ccnb1 IFITM1 Ifitm1 RPS27A Rps27a-ps1 NCKAP1L Nckap1l IL16 Il16 ZWINT Zwint C19orf66 Shfl RPL7A Rpl7a SPI1 Spi1 TRAF3IP3 Traf3ip3 CENPE Cenpe RNF213 Rnf213 RPSA Rpsa CYBB Cybb EVI2B Evi2b ANLN Anln IFI16 Mnda RPS19 Rps19 TYMP Tymp DOCK2 Dock2 CENPF Cenpf IRF9 Irf9 RPS6 Rps6 SNX10 Snx10 IL10RA Il10ra CCNA2 Ccna2 TRIM22 RPLP2 Rplp2 VSIG4 Vsig4 ARHGAP15 Arhgap15 KIF20A Kif20a RTP4 Rtp4 RPL13A Rpl13a HK3 Hk3 PRKCH Prkch BUB1B Bub1b BATF2 Batf2 RPS13 Rps13 IGSF6 Igsf6 LCP1 Lcp1 TTK Ttk TAP1 Tap1 RPL6 Rpl6 MSR1 Msr1 CD27 Cd27 PTTG1 Pttg1 GBP1 Gbp1 RPS18 Rps18l1 LILRB4 Lilrb4 FAM26F Calhm6 NUSAP1 Nusap1 PSMB9 Psmb9 RPS15 Rps15 TBXAS1 Tbxas1 DOCK8 Dock8 NUF2 Nuf2 MX2 RGD1308751 RPS28 Rps28 CD300A Cd300a CD3G Cd3g RACGAP1 Racgap1 ZNFX1 Znfx1 RPL9 Rpl9 TLR8 Tlr8 GIMAP2 MELK Melk LAMP3 Lamp3 RPL32 LOC680959 MNDA Mnda NCF1B RRM1 Rrm1 IFIT5 RPL23 Rpl23 FCGR1B FLI1 Fli1 TCF19 Tcf19 SAMD9 Samd9 RPS3 Rps3 FPR3 Fpr3 CXCR6 Cxcr6 CDT1 Cdt1 IFI35 Ifi35 RPS5 Rps5 FCGR1A Fcgr1a SH2D1A Sh2d1a POLE2 Pole2 EPSTI1 Epsti1 RPL17 Rpl17 NPL Npl PVRIG Pvrig TK1 Tk1 PARP12 Parp12 RPS14 Rps14 CD4 Cd4 CYTIP Cytip MCM4 Mcm4 IFIT2 Ifit2 RPL12 Rpl12 LY96 Ly96 TRAT1 Trat1 GMNN Gmnn CMPK2 Cmpk2 RPL3 Rpl3 MYO1F Myo1f CD3E Cd3e MCM2 Mcm2 UBE2L6 Ube2l6 RPL21 Rpl21 CYTH4 Cyth4 GIMAP6 Gimap6 MND1 Mnd1 OAS2 Oas2 NACA Naca CD86 Cd86 CD96 Cd96 CDC20 Cdc20 PARP14 Parp14 RPL13 Rpl13 LAIR1 Lair1 CD3D Cd3d DEPDC1B Depdc1b PHF11 Phf11 RPL34 Rpl34l1 LAPTM5 Laptm5 FYB Fyb1 PCNA Pcna SHISA5 Shisa5 RPL14 Rpl14 PLA2G7 Pla2g7 CRTAM Crtam CDC6 Cdc6 FBXO6 Fbxo6 RPL29 Rpl29 BCL2A1 Bcl2a1 CCL19 Ccl19 FOXM1 Foxm1 PARP10 Parp10 RPS8 Rps8 C2 C2 BIN2 Bin2 RRM2 Rrm2 IFI6 RPS16 Rps16 ADORA3 Adora3 PARVG Parvg TYMS Tyms APOL6 EIF3H LOC100911110 MAN2B1 Man2b1 DOCK10 Dock10 TRIP13 Trip13 USP18 Usp18 RPL35A Rpl35al1 ARRB2 Arrb2 TARP Tarp MTFR2 Mtfr2 DDX60 Ddx60 RPS10 Rps10l1 EMR2 KLRB1 Klrb1b CENPL Cenpl LAP3 Lap3 RPL19 Rpl19 DPYD Dpyd KLHL6 Klhl6 BIRC5 Birc5 HERC6 Herc6 EIF3D Eif3d ADAMDEC1 Adamdec1 CCR7 Ccr7 CKS1B Cks1b RSAD2 Rsad2 RPL38 Pramef27 GPNMB Gpnmb CD6 Cd6 MCM10 Mcm10 IFIT1 Ifit1bl RPS7 Rps7 MFSD1 Mfsd1 UBASH3A Ubash3a SPC25 Spc25 PML Pml RPS17 Rsl1 ITGAX Itgax TRGV9 E2F8 E2f8 TRIM5 Trim5 RPL27 Rpl27 CMKLR1 Cmklr1 PSTPIP1 Pstpip1 CDCA2 Cdca2 APOL1 RPL37 Rpl37 MS4A7 Ms4a7 IL7R Il7r CDKN3 Cdkn3 DHX58 Dhx58 RPL35 Rpl35 TNFRSF1B Tnfrsf1b GPR171 Gpr171 CDCA3 Cdca3 HSH2D Hsh2d RPS2 Rps2 PLEKHO2 Plekho2 EVI2A Evi2a CENPA Cenpa SIGLEC1 Siglec1 RPL36A LOC100912182 MS4A4A Larp1b APBB1IP Apbb1ip UBE2C Ube2c DDX58 Ddx58 EEF1G Eef1g CTSS Ctss AMICA1 Jaml OIP5 Oip5 HELZ2 Helz2 GNB2L1 Rack1 AOAH Aoah BTK Btk KIF11 Kif11 HERC5 EEF1D Eef1d ITGAM Itgam HMHA1 Arhgap45 CCNB2 Ccnb2 OASL Oasl RPL7 Rpl7 CSF1R Csf1r PTPRCAP Ptprcap NEK2 Nek2 TRIM21 Trim21 SNHG6 SLC31A2 Slc31a2 ITK Itk STIL Stil RPS20 Rps20 C1QA C1qa SLA Sla ECT2 Ect2 RPL30 Rpl30 SCPEP1 Scpep1 GIMAP5 Gimap5 AURKA Aurka RPL28 Rpl28 C1orf54 RCSD1 Rcsd1 KIF15 Kif15 RPS29 Rps29 TMEM140 Tmem140 SASH3 Sash3 KIF2C Kif2c RPL8 Rpl8 C5AR1 C5ar1 FYN Fyn KIF18B Kif18b RPS21 Rps21 NR1H3 Nr1h3 TNFRSF9 Tnfrsf9 KIF14 Kif14 RPL23A RGD1564606 ATP8B4 Atp8b4 CD28 Cd28 TPX2 Tpx2 RPL22 Rpl22 LIPA Lipa HVCN1 Hvcn1 FAM72C RPL27A Rpl27a CD68 Cd68 CXCL9 Cxcl9 DLGAP5 Dlgap5 RPS11 Rps11 CCRL2 Ccrl2 LY86 Ly86 ASPM Aspm RPL37A Rpl37a SLCO2B1 Slco2b1 RGS18 Rgs18 DEPDC1 Depdc1 RPL10L Rpl10l TRGC2 KIF4A Kif4a RPLP0 Rplp0 DPEP2 Dpep2 HMGB3 LOC688583 RPL4 Rpl4 SIRPG CDCA8 Cdca8 EIF3G Eif3g CD8A Cd8a PLK4 Plk4 SNRPD2 Snrpd2 ICOS Icos CCNE2 Ccne2 RPL39 Rpl39 GAB3 CDCA7 Cdca7 EEF1A1 Eef1a1 GMFG Gmfg SHCBP1 Shcbp1 EIF3K Eif3k STMN1 Stmn1 UXT Uxt DONSON Donson PARPBP Parpbp SKA1 Ska1 FBXO5 Fbxo5 MKI67 Mki67 CEP55 Cep55 NCAPG Ncapg POLQ Polq AURKB Aurkb CASC5 Knl1 HJURP Hjurp CDCA5 LOC686151 BUB1 Bub1

10