Journal: Biochemistry

Article Title: Targeted degradation of the oncogenic phosphatase SHP2

doi: 10.1021/acs.biochem.1c00377

Figure Lengend Snippet: (A) Chemical structure of RMC-4550 and X-ray crystal structure of SHP2 in complex with RMC-4550 (PDB code 7RCT). Surface representation of SHP2 in complex with RMC-4550 bound in the central tunnel formed at the interface of N-SH2 (green), C-SH2 (blue) and PTP (wheat) domains. (B) Chemical structures of RMC-4550-based PROTAC candidates, R1–1C, R1–3C and R1–5C.

Article Snippet: Antibodies used in this study were obtained commercially from the following sources: SHP2 (Bethyl, #A301–544A), Phospho-Thr202/Tyr204-Erk1/2 (CST, #9101), Cereblon (CST, #71810), b-actin (Millipore-Sigma, #A1978), b -tubulin (CST, #2146), GAPDH (CST, #5174).

Techniques:

Journal: Biochemistry

Article Title: Targeted degradation of the oncogenic phosphatase SHP2

doi: 10.1021/acs.biochem.1c00377

Figure Lengend Snippet: (A) Inhibition of SHP2-F285S- or PTP-mediated DIFMUP dephosphorylation by R1–1C, R1–3C, R1–5C and RMC-4550. MV4;11 cells were treated with increasing doses of R1–3C (B), R1–1C or R1–5C (C) for 24 h and subjected to Western blotting using SHP2, GAPDH and β-actin antibodies. Quantification of band intensities on the gels are shown below the blots.

Article Snippet: Antibodies used in this study were obtained commercially from the following sources: SHP2 (Bethyl, #A301–544A), Phospho-Thr202/Tyr204-Erk1/2 (CST, #9101), Cereblon (CST, #71810), b-actin (Millipore-Sigma, #A1978), b -tubulin (CST, #2146), GAPDH (CST, #5174).

Techniques: Inhibition, De-Phosphorylation Assay, Western Blot

Journal: Biochemistry

Article Title: Targeted degradation of the oncogenic phosphatase SHP2

doi: 10.1021/acs.biochem.1c00377

Figure Lengend Snippet: (A) Time course of SHP2 degradation by R1–5C (100 nM) in MV4;11 cells. Immunoblotting with SHP2 and β-actin antibodies. (B) CRBN−/− and parental MOLT4 cells were treated with increasing doses of R1–5C for 24 h and subjected to Western blotting using SHP2, CRBN and β-actin antibodies. Quantification of band intensities on the gels are shown below the blots.

Article Snippet: Antibodies used in this study were obtained commercially from the following sources: SHP2 (Bethyl, #A301–544A), Phospho-Thr202/Tyr204-Erk1/2 (CST, #9101), Cereblon (CST, #71810), b-actin (Millipore-Sigma, #A1978), b -tubulin (CST, #2146), GAPDH (CST, #5174).

Techniques: Western Blot

Journal: Biochemistry

Article Title: Targeted degradation of the oncogenic phosphatase SHP2

doi: 10.1021/acs.biochem.1c00377

Figure Lengend Snippet: (A-D) Scatterplots displaying relative fold-change in SHP2 abundance following treatment of MV4;11 cells with 100 nM R1–5C for 4 h (A), 8 h (B), 16 h (C) or 100 nM RMC-4550 (D). SHP2/PTPN11 is highlighted in red. Hits highlighted in blue in (C) and (D) indicate changes in abundance of proteins at 16 h time point due to secondary effects (such as transcriptional responses) of SHP2 degradation or inhibition. (E) Heatmap of the protein abundance changes in MV4;11 cells comparing treatment with 100 nM R1–1C (4 h and 16 h), 100 nM R1–3C (4 h and 16 h), 100 nM R1–5C (2 h, 4 h, 8 h and 16 h), 100 nM RMC-4550 (16 h) and 1 μM pomalidomide (5 h). The heatmap colors are scaled with red indicating a decrease in protein abundance (−2 log2 FC) and blue indicating an increase (2 log2 FC) in protein abundance.

Article Snippet: Antibodies used in this study were obtained commercially from the following sources: SHP2 (Bethyl, #A301–544A), Phospho-Thr202/Tyr204-Erk1/2 (CST, #9101), Cereblon (CST, #71810), b-actin (Millipore-Sigma, #A1978), b -tubulin (CST, #2146), GAPDH (CST, #5174).

Techniques: Inhibition

Journal: Cell reports

Article Title: Bach2 repression of CD36 regulates lipid-metabolism-linked effector functions in follicular B cells.

doi: 10.1016/j.celrep.2024.114878

Figure Lengend Snippet: Figure 6. CD36 inhibition reduces IL-6 production by B cells (A) WT and Bach2/ B cells were cultured with LPS in the presence or absence of 100 mM SSO for 24 h and assayed by intracellular FACS to detect IL-6- expressing cells. Representative FACS profiles gated on live cells and bar graphs displayed as means ± SEM are shown. (B) IL-6 concentrations in culture supernatants cultured as in (A) were determined by ELISA. Cell viability was determined by FACS using a Live/Dead kit.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Total IgM ELISA KIT Bethyl Laboratories, Inc. Cat#E99-101 Experimental models: Organisms/strains Mouse: Bach2 Knock-Out Tohoku University N/A Mouse: CD36 fl/fl Jackson Laboratory Cat#032276 Mouse: CD19-cre Jackson Laboratory Cat#006785 Experimental models: Cell line Jurkat cells ATCC Cat# TIB-152 Oligonucleotides CpG oligonucleotide BD Biosciences Cat#554714 Deoxyribonucleic acid Sigma Cat#D1501 b-actin-FWD: 50-GACGGCCAGGTCATCACTATTG -30 This paper N/A b-actin-REV: 50-AGGAAGGCTGGAAAAGAGCC-30 This paper N/A Bach2-FWD: 50-CTGTAGCCTTCTCATCTCTTCC-30 This paper N/A Bach2-REV: 50-TCCTTCTTCCGTTGGTCATTG-30 This paper N/A Blimp1-FWD: 50-ACCGTCTTGAGGACATGGAG-30 This paper N/A Blimp1-REV: GTTGCTGTGAGGCAACTTCA-30 This paper N/A Cd36-FWD: 50-GCATGGCAGCTTTGGGCAGG-30 This paper N/A Cd36-REV: 50-TGCACAGGAGAGGCGGGCAT-30 This paper N/A Uchl1-FWD: 50-GAAGCAGACCATCGGAAACTCC-30 This paper N/A Uchl1-REV: 50-GGACAGCTT CTCCGTTTCAGAC-30 This paper N/A Lxr-b-FWD: 50-CGCTACAACCACGAGACAGA-30 This paper N/A Lxr-b-REV: 50-TGTTGATGGCGATAAGCAAG-30 This paper N/A Abca1-FWD: 50-CTCAGTTAAGGCTGCTGCTG- 30 This paper N/A Abca1-REV: 50-TCAGGCGTACAGAGATCAGG-3’.

Techniques: Inhibition, Cell Culture, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Cell reports

Article Title: Bach2 repression of CD36 regulates lipid-metabolism-linked effector functions in follicular B cells.

doi: 10.1016/j.celrep.2024.114878

Figure Lengend Snippet: Figure 7. The effect of metabolic inhibitors on plasma cell differentiation WT and Bach2/ B cells were cultured with LPS plus IL-4 in the presence or absence of 100 mM etomoxir (A), 0.5 mM 2-DG (B), and 20 mM SB203580 (C). After 48 h, CD138-expressing B cells were analyzed by flow cytometry. IgM con- centrations in the culture supernatants were determined by ELISA. Cell viability was determined by FACS using a Live/Dead kit. Data are pooled from three independent experiments. *p < 0.05 and **p < 0.01 by two-tailed paired Student’s t tests.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Total IgM ELISA KIT Bethyl Laboratories, Inc. Cat#E99-101 Experimental models: Organisms/strains Mouse: Bach2 Knock-Out Tohoku University N/A Mouse: CD36 fl/fl Jackson Laboratory Cat#032276 Mouse: CD19-cre Jackson Laboratory Cat#006785 Experimental models: Cell line Jurkat cells ATCC Cat# TIB-152 Oligonucleotides CpG oligonucleotide BD Biosciences Cat#554714 Deoxyribonucleic acid Sigma Cat#D1501 b-actin-FWD: 50-GACGGCCAGGTCATCACTATTG -30 This paper N/A b-actin-REV: 50-AGGAAGGCTGGAAAAGAGCC-30 This paper N/A Bach2-FWD: 50-CTGTAGCCTTCTCATCTCTTCC-30 This paper N/A Bach2-REV: 50-TCCTTCTTCCGTTGGTCATTG-30 This paper N/A Blimp1-FWD: 50-ACCGTCTTGAGGACATGGAG-30 This paper N/A Blimp1-REV: GTTGCTGTGAGGCAACTTCA-30 This paper N/A Cd36-FWD: 50-GCATGGCAGCTTTGGGCAGG-30 This paper N/A Cd36-REV: 50-TGCACAGGAGAGGCGGGCAT-30 This paper N/A Uchl1-FWD: 50-GAAGCAGACCATCGGAAACTCC-30 This paper N/A Uchl1-REV: 50-GGACAGCTT CTCCGTTTCAGAC-30 This paper N/A Lxr-b-FWD: 50-CGCTACAACCACGAGACAGA-30 This paper N/A Lxr-b-REV: 50-TGTTGATGGCGATAAGCAAG-30 This paper N/A Abca1-FWD: 50-CTCAGTTAAGGCTGCTGCTG- 30 This paper N/A Abca1-REV: 50-TCAGGCGTACAGAGATCAGG-3’.

Techniques: Clinical Proteomics, Cell Differentiation, Cell Culture, Expressing, Cytometry, Enzyme-linked Immunosorbent Assay, Two Tailed Test

Journal: Cell reports

Article Title: UTP18-mediated p21 mRNA instability drives adenoma-carcinoma progression in colorectal cancer.

doi: 10.1016/j.celrep.2023.112423

Figure Lengend Snippet: Figure 3. UTP18 enhances the cell viability of adenoma organoids and promotes tumorigenesis of adenoma organoids in nude mice (A) Tissue morphology of No. 51 and No. 52 patient-derived adenoma organoids under a white-light microscope after 6, 9, and 12 days of culture. (B) UTP18 knockdown verification in No. 51 patient-derived adenoma organoids using qPCR (left) and ATP activity quantification (right). *p < 0.05, **p < 0.01, ***p < 0.001.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies UTP18 (Western Blot and IP) Bethyl Cat# A301-551A; RRID: AB_1039989 UTP18 (IHC) Sigma Cat# HPA052378 UTP14A Proteintech Cat# 11474-1-AP; RRID: AB_2272800 GTPBP4 Proteintech Cat# 13897-1-AP; RRID: AB_2279486 GNL3 Sigma Cat# SAB1407312; RRID: AB_10760256 ACTB GeneTex Cat# GTX124213; RRID: AB_11171102 p21 Cell Signaling Technology Cat# 2947; RRID: AB_823586 p27 Cell Signaling Technology Cat# 3686; RRID: AB_2077850 RB Cell Signaling Technology Cat# 9309; RRID: AB_823629 RB-P Cell Signaling Technology Cat# 8180; RRID: AB_10950972 GAPDH MBL Cat# M171-3; RRID: AB_10597731 SUMO1 Santa Cruz Cat# sc-5308; RRID: AB_628300 SUMO2/3 Abcam Cat# ab81371; RRID: AB_1658424 Lamin A/C Abcam Cat# ab108922; RRID: AB_10860619 Flag Sigma Cat# F1804; RRID: AB_262044 Ki67 Abcam Cat# ab15580; RRID: AB_443209 PCNA AmyJet Scientific Cat# ABM40031 Mouse (G3A1) mAb IgG1 Isotype Control Cell Signaling Technology Cat# 5415; RRID: AB_10829607 Normal Rabbit IgG Cell Signaling Technology Cat# 2729; RRID: AB_1031062 Biological samples Tissue of colorectal cancer samples Peking Union Medical College Hospital N/A Tissue of colorectal adenoma samples Peking Union Medical College Hospital N/A Tissue of gastric cancer samples Peking Union Medical College Hospital N/A Tissue of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of gastric cancer samples Peking Union Medical College Hospital N/A Serum of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of healthy samples Peking Union Medical College Hospital N/A Chemicals, peptides, and recombinant proteins Iodoacetamide GE Healthcare RPN6302V Dithiothreitol PlusOne 17-1318-01 Urea Sigma U5128 Trypsin Roche 11418033001 Trypsin/Lys-C Protease Mix Promega V5073 Lys-C Roche 11047825001 Triethylammonium bicarbonate buffer Sigma T7408 Trifluoroacetic acid Thermo Scientific 85183 L-Glutamine Thermo Fisher Scientific 25030149 N2 Supplement Thermo Fisher Scientific 17502048 B27 Gibco 12587-010 N-acetylcysteine Sigma A9165 ActD Merck SBR00013 (Continued on next page) Cell Reports 42, 112423, May 30, 2023 19

Techniques: Derivative Assay, Light Microscopy, Knockdown, Activity Assay

Journal: Cell reports

Article Title: UTP18-mediated p21 mRNA instability drives adenoma-carcinoma progression in colorectal cancer.

doi: 10.1016/j.celrep.2023.112423

Figure Lengend Snippet: Figure 4. UTP18 knockdown inhibits the malignancy of CRC cells (A) UTP18 knockdown verification by western blotting in CRC cell lines. (B) UTP18 knockdown inhibits cell viability in SW480 and HCT116 cell lines. ***p < 0.001. (C) UTP18 knockdown impairs colony formation ability in SW480 and HCT116 cell lines. Data are presented as the mean ± SEM, ***p < 0.001. (D) UTP18 knockdown impedes the migration and invasion capability of SW480 and HCT116 cell lines. **p < 0.01, ***p < 0.001. (E) UTP18 knockdown slows down tumor cells proliferation and inhibits tumor growth in a xenograft mouse model (n = 6). **p < 0.01. (F) Representative IHC images of UTP18 and Ki67 expression in xenograft tumors. *p < 0.05, **p < 0.01. Data are presented as the mean ± SD (n = 3), unless otherwise specified. p values were calculated using two-tailed Student’s t test. See also Figure S2.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies UTP18 (Western Blot and IP) Bethyl Cat# A301-551A; RRID: AB_1039989 UTP18 (IHC) Sigma Cat# HPA052378 UTP14A Proteintech Cat# 11474-1-AP; RRID: AB_2272800 GTPBP4 Proteintech Cat# 13897-1-AP; RRID: AB_2279486 GNL3 Sigma Cat# SAB1407312; RRID: AB_10760256 ACTB GeneTex Cat# GTX124213; RRID: AB_11171102 p21 Cell Signaling Technology Cat# 2947; RRID: AB_823586 p27 Cell Signaling Technology Cat# 3686; RRID: AB_2077850 RB Cell Signaling Technology Cat# 9309; RRID: AB_823629 RB-P Cell Signaling Technology Cat# 8180; RRID: AB_10950972 GAPDH MBL Cat# M171-3; RRID: AB_10597731 SUMO1 Santa Cruz Cat# sc-5308; RRID: AB_628300 SUMO2/3 Abcam Cat# ab81371; RRID: AB_1658424 Lamin A/C Abcam Cat# ab108922; RRID: AB_10860619 Flag Sigma Cat# F1804; RRID: AB_262044 Ki67 Abcam Cat# ab15580; RRID: AB_443209 PCNA AmyJet Scientific Cat# ABM40031 Mouse (G3A1) mAb IgG1 Isotype Control Cell Signaling Technology Cat# 5415; RRID: AB_10829607 Normal Rabbit IgG Cell Signaling Technology Cat# 2729; RRID: AB_1031062 Biological samples Tissue of colorectal cancer samples Peking Union Medical College Hospital N/A Tissue of colorectal adenoma samples Peking Union Medical College Hospital N/A Tissue of gastric cancer samples Peking Union Medical College Hospital N/A Tissue of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of gastric cancer samples Peking Union Medical College Hospital N/A Serum of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of healthy samples Peking Union Medical College Hospital N/A Chemicals, peptides, and recombinant proteins Iodoacetamide GE Healthcare RPN6302V Dithiothreitol PlusOne 17-1318-01 Urea Sigma U5128 Trypsin Roche 11418033001 Trypsin/Lys-C Protease Mix Promega V5073 Lys-C Roche 11047825001 Triethylammonium bicarbonate buffer Sigma T7408 Trifluoroacetic acid Thermo Scientific 85183 L-Glutamine Thermo Fisher Scientific 25030149 N2 Supplement Thermo Fisher Scientific 17502048 B27 Gibco 12587-010 N-acetylcysteine Sigma A9165 ActD Merck SBR00013 (Continued on next page) Cell Reports 42, 112423, May 30, 2023 19

Techniques: Knockdown, Western Blot, Migration, Expressing, Two Tailed Test

Journal: Cell reports

Article Title: UTP18-mediated p21 mRNA instability drives adenoma-carcinoma progression in colorectal cancer.

doi: 10.1016/j.celrep.2023.112423

Figure Lengend Snippet: Figure 5. UTP18 regulates cell-cycle progression (A) KEGG pathway analysis of DERs. (B) KEGG pathway analysis of DEPs. (C) UTP18 knockdown induces cell-cycle arrest in the G0/G1 phase. (D) Volcano plot of the distribution of all differentially expressed mRNAs. (E) qRT-PCR validation of differentially expressed mRNAs. *p < 0.05, **p < 0.01, ***p < 0.001. (F) Western blotting validation of cell-cycle regulation proteins. Data are presented as the mean ± SD (n = 3). p values were calculated using two-tailed Student’s t test. See also Figure S2; Tables S1 and S7.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies UTP18 (Western Blot and IP) Bethyl Cat# A301-551A; RRID: AB_1039989 UTP18 (IHC) Sigma Cat# HPA052378 UTP14A Proteintech Cat# 11474-1-AP; RRID: AB_2272800 GTPBP4 Proteintech Cat# 13897-1-AP; RRID: AB_2279486 GNL3 Sigma Cat# SAB1407312; RRID: AB_10760256 ACTB GeneTex Cat# GTX124213; RRID: AB_11171102 p21 Cell Signaling Technology Cat# 2947; RRID: AB_823586 p27 Cell Signaling Technology Cat# 3686; RRID: AB_2077850 RB Cell Signaling Technology Cat# 9309; RRID: AB_823629 RB-P Cell Signaling Technology Cat# 8180; RRID: AB_10950972 GAPDH MBL Cat# M171-3; RRID: AB_10597731 SUMO1 Santa Cruz Cat# sc-5308; RRID: AB_628300 SUMO2/3 Abcam Cat# ab81371; RRID: AB_1658424 Lamin A/C Abcam Cat# ab108922; RRID: AB_10860619 Flag Sigma Cat# F1804; RRID: AB_262044 Ki67 Abcam Cat# ab15580; RRID: AB_443209 PCNA AmyJet Scientific Cat# ABM40031 Mouse (G3A1) mAb IgG1 Isotype Control Cell Signaling Technology Cat# 5415; RRID: AB_10829607 Normal Rabbit IgG Cell Signaling Technology Cat# 2729; RRID: AB_1031062 Biological samples Tissue of colorectal cancer samples Peking Union Medical College Hospital N/A Tissue of colorectal adenoma samples Peking Union Medical College Hospital N/A Tissue of gastric cancer samples Peking Union Medical College Hospital N/A Tissue of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of gastric cancer samples Peking Union Medical College Hospital N/A Serum of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of healthy samples Peking Union Medical College Hospital N/A Chemicals, peptides, and recombinant proteins Iodoacetamide GE Healthcare RPN6302V Dithiothreitol PlusOne 17-1318-01 Urea Sigma U5128 Trypsin Roche 11418033001 Trypsin/Lys-C Protease Mix Promega V5073 Lys-C Roche 11047825001 Triethylammonium bicarbonate buffer Sigma T7408 Trifluoroacetic acid Thermo Scientific 85183 L-Glutamine Thermo Fisher Scientific 25030149 N2 Supplement Thermo Fisher Scientific 17502048 B27 Gibco 12587-010 N-acetylcysteine Sigma A9165 ActD Merck SBR00013 (Continued on next page) Cell Reports 42, 112423, May 30, 2023 19

Techniques: Knockdown, Quantitative RT-PCR, Biomarker Discovery, Western Blot, Two Tailed Test

Journal: Cell reports

Article Title: UTP18-mediated p21 mRNA instability drives adenoma-carcinoma progression in colorectal cancer.

doi: 10.1016/j.celrep.2023.112423

Figure Lengend Snippet: Figure 7. UTP18 decreases the stability of p21 mRNA and screening of small-molecule inhibitors (A) Negative correlation between p21 and UTP18 mRNA expression in the GEO database. **p < 0.01. (B) Protein lysates of HCT116 cells were subjected to RNA IP followed by qRT-PCR analysis to measure the relative quantities of p21 mRNA in UTP18 IP compared with control IgG IP. *p < 0.05, **p < 0.01. (C) HCT116 cells were treated with ActD, and p21 mRNA expression levels after treatment were examined at the indicated time points. *p < 0.05. (D) RNA IP followed by qRT-PCR analysis was performed to measure the relative quantities of p21 mRNA in NCM460 and HCT116 cells. **p < 0.01. (E) RNA IP followed by qRT-PCR analysis was performed to measure the relative quantities of p21 mRNA in UTP18 SUMO-deficient mutant HCT116 cells. **p < 0.01, ***p < 0.001.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies UTP18 (Western Blot and IP) Bethyl Cat# A301-551A; RRID: AB_1039989 UTP18 (IHC) Sigma Cat# HPA052378 UTP14A Proteintech Cat# 11474-1-AP; RRID: AB_2272800 GTPBP4 Proteintech Cat# 13897-1-AP; RRID: AB_2279486 GNL3 Sigma Cat# SAB1407312; RRID: AB_10760256 ACTB GeneTex Cat# GTX124213; RRID: AB_11171102 p21 Cell Signaling Technology Cat# 2947; RRID: AB_823586 p27 Cell Signaling Technology Cat# 3686; RRID: AB_2077850 RB Cell Signaling Technology Cat# 9309; RRID: AB_823629 RB-P Cell Signaling Technology Cat# 8180; RRID: AB_10950972 GAPDH MBL Cat# M171-3; RRID: AB_10597731 SUMO1 Santa Cruz Cat# sc-5308; RRID: AB_628300 SUMO2/3 Abcam Cat# ab81371; RRID: AB_1658424 Lamin A/C Abcam Cat# ab108922; RRID: AB_10860619 Flag Sigma Cat# F1804; RRID: AB_262044 Ki67 Abcam Cat# ab15580; RRID: AB_443209 PCNA AmyJet Scientific Cat# ABM40031 Mouse (G3A1) mAb IgG1 Isotype Control Cell Signaling Technology Cat# 5415; RRID: AB_10829607 Normal Rabbit IgG Cell Signaling Technology Cat# 2729; RRID: AB_1031062 Biological samples Tissue of colorectal cancer samples Peking Union Medical College Hospital N/A Tissue of colorectal adenoma samples Peking Union Medical College Hospital N/A Tissue of gastric cancer samples Peking Union Medical College Hospital N/A Tissue of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of colorectal cancer samples Peking Union Medical College Hospital N/A Serum of gastric cancer samples Peking Union Medical College Hospital N/A Serum of ulcerative colitis samples Peking Union Medical College Hospital N/A Serum of healthy samples Peking Union Medical College Hospital N/A Chemicals, peptides, and recombinant proteins Iodoacetamide GE Healthcare RPN6302V Dithiothreitol PlusOne 17-1318-01 Urea Sigma U5128 Trypsin Roche 11418033001 Trypsin/Lys-C Protease Mix Promega V5073 Lys-C Roche 11047825001 Triethylammonium bicarbonate buffer Sigma T7408 Trifluoroacetic acid Thermo Scientific 85183 L-Glutamine Thermo Fisher Scientific 25030149 N2 Supplement Thermo Fisher Scientific 17502048 B27 Gibco 12587-010 N-acetylcysteine Sigma A9165 ActD Merck SBR00013 (Continued on next page) Cell Reports 42, 112423, May 30, 2023 19

Techniques: Expressing, Quantitative RT-PCR, Control, Mutagenesis