KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Anti-Ly6c antibody [ER-MP20] | Abcam | ab15627;RRID:AB_302004 |
| InVivoMAb anti-mouse Ly6G/Ly6C (Gr-1), 1mg | BioXCell | BE0075;RRID:AB_10312146 |
| Anti-Rabbit p-β-Catenin (Ser552) Monoclonal Antibody | Zymed custom antibody | Affinity Pure Rb Ab, 0.5mg/ml |
| Anti-Rabbit β-actin polyclonal Antibody | Abcam | Cat# ab8227 RRID: AB_2305186 |
| Anti-Mouse MHC Class II Monoclonal Antibody (AF6-120.1), APC | Thermo Fisher | Cat# 17-5320-82;RRID:AB_2573212 |
| SP-1 Chromogranin A (Porcine) Antibody | Immunostar | 20086;RRID:AB_572226 |
| FcR Blocking Reagent, mouse | Miltenyi Biotec | 130-092-575;RRID:AB_2892833 |
| Ly-6G (Gr-1) Monoclonal Antibody (1A8-Ly6g), PE-eFluor 610, eBioscience | ThermoFisher | 61-9668-82;RRID:AB_2574679 |
| Ly-6C Monoclonal Antibody (HK1.4), Alexa Fluor 488, eBioscience | ThermoFisher | 53-5932-82;RRID:AB_2574427 |
| CD11b Monoclonal Antibody (M1/70), APC-eFluor 780, eBioscience | ThermoFisher | 47-0112-82;RRID:AB_1603193 |
| Lysozyme EC 3.2.1.17 | Agilent | A009902-2;RRID:AB_578661 |
| Prostaglandin E Receptor EP4 Antibody | VWR/Bioss | bs-8538R |
| Anti-F4/80 antibody [SP115] (ab111101), Rabbit | Abcam | ab111101;RRID:AB_10859466 |
| Anti-CD68 antibody (ab125212), Rabbit | Abcam | ab125212:RRID:AB_10975465 |
| Anti-CD11b antibody [EPR1344] (ab133357), rabbit | Abcam | ab133357;RRID:AB_2650514 |
| CD44-Biotin, mouse | Miltenyi Biotec | 130-110-082;RRID:AB_2661032 |
| CD24-Biotin, mouse | Miltenyi Biotec | 130-102-128;RRID:AB_2656584 |
| Anti-CD68 antibody [C68/684] | Abcam | ab201340;RRID:AB_2747829 |
| Anti-CD11b antibody [M1/70] | Abcam | ab8878;RRID:AB_306831 |
| Anti-CD11b antibody [EPR1344] (ab133357), rabbit | Abcam | ab133357;RRID:AB_2650514 |
| Anti-Rabbit Krt15 Polyclonal Antibody | Novus | Cat# NBP1-85602 RRID: AB_11037933 |
| Anti-Mouse Ascl2 Monoclonal Antibody | LSBio | Cat# LS-C126891;RRID:AB_10831114 |
| CD9 | Abcam | ab92726;RRID:AB_10561589 |
| Bmi1 antibody | Abcam | ab14389;RRID:AB_2065390 |
| Annexin A2 | Abcam | ab178677;RRID:AB_1140698 |
| Annexin A1 | Abcam | ab214486;RRID:AB_722804 |
| Ascl2 antibody | Biorbyt | orb155740;RRID:AB_10865613 |
| E-Cadherin (24E10) Rabbit mAb | Cell Signaling Technology | 3195;RRID:AB_2291471 |
| PD-1 Monoclonal Antibody (7A11B1) | ThermoFisher | MA5-15780;RRID:AB_11152225 |
| Beta-Actin (8H10D10) Mouse mAb | Cell Signaling Technology | 3700S;RRID:AB_2242334 |
| Mouse TrueBlot ULTRA:Anti-Mouse Ig HRP | Rockland | 18-8817-33;RRID:AB_2610851 |
| CST alpha-tubulin Antibody | Cell Signaling Technology | 2144;RRID:AB_2210548 |
| Bmi1 (D20B7) XP® Rabbit mAb | Cell Signaling Technology | 6964;RRID:AB_10828713 |
| Purified Mouse Anti-Human CD63 Clone H5C6 | Fisher/BD Biosciences | 556019;RRID:AB_396297 |
| Krt15 antibody (Cytokeratin 15) | Fisher/Novus Biologicals | NBP2-50461AF488; RRID:AB_1084834 |
| BrdU antibody | Fisher/Novus Biologicals | NB600-720;RRID:AB_10002707 |
| S100A10 Antibody | VWR/ProteinTech | 11250-1-AP;RRID:AB_2269906 |
| Donkey Anti-Rabbit IgG H&L (Alexa Fluor® 594) |
Abcam | ab150076;RRID:AB_2782993 |
| ASCL2 Antibody (C terminus) LS-C126891 | LifeSpan Biosciences | LS-C126891-100;RRID:AB_10831114 |
| Cytokeratin 15 Antibody | Novus Biologicals | NBP1-85602;RRID:AB_11037933 |
| MHC Class II I-Ab Monoclonal Antibody (AF6-120.1), APC, eBioscience | ThermoFisher | 17-5320-82;RRID:AB_2573212 |
| Mouse IgG2a kappa Isotype Control (eBM2a), APC, eBioscience | ThermoFisher | 17-4724-81;RRID:AB_470188 |
| CD45 Monoclonal Antibody (30-F11), PE-Cyanine5, eBioscience | ThermoFisher | 15-0451-83;RRID:AB_468753 |
| Rat IgG2b kappa Isotype Control (eB149/10H5), PE-Cyanine5, eBioscience | ThermoFisher | 15-4031-82;RRID:AB_470133 |
| CD11b Monoclonal Antibody (M1/70), PE, eBioscience | ThermoFisher | 12-0112-83;RRID:AB_2734870 |
| Rat IgG2b kappa Isotype Control (eB149/10H5), PE, eBioscience | ThermoFisher | 12-4031-82;RRID:AB_470042 |
| CD11c Monoclonal Antibody (N418), PE-Cyanine7, eBioscience | ThermoFisher | 25-0114-82;RRID:AB_469590 |
| Armenian Hamster IgG Isotype Control (eBio299Arm), PE-Cyanine7, eBioscience | ThermoFisher | 25-4888-82;RRID:AB_470204 |
| FITC anti-mouse CX3CR1 Antibody | VWR/Biolegend | 149020;RRID:AB_2565703 |
| FITC Mouse IgG2a, κ Isotype Ctrl Antibody | VWR/Biolegend | 400208;RRID:AB_2884007 |
| Annexin V-mFluor Violet 540 conjugate | AAT Bioquest | 20080 |
| EP4 antibody | Abcam | ab217966 |
| CD3 | Abcam | ab16669;RRID:AB_443425 |
| Anti-BrdU antibody [BU1/75 (ICR1)] | Abcam | ab6326;RRID:AB_305426 |
| CD68 Antibody | Abcam | ab201340;RRID:AB_991703 |
| CD19 Antibody | Abcam | ab203615;RRID:AB_1140606 |
| Ascl2 antibody, 100 μg | Biorbyt | orb155740 |
| Cytokeratin 15 Antibody | Fisher/Novus Biologicals | NBP2-53308-100ug |
| TCF21 Antibody | Fisher/Novus Biologicals | NBP1-88637;RRID:AB_11008184 |
| Rat monoclonal [YTS169.4] to CD8 | Abcam | ab22378;RRID:AB_447033 |
| Anti-Sox9 antibody | Sigma/Millipore | AB5535;RRID:AB_2239761 |
| RM-9106-S1/Ab Ki-67 0.5mL/EA | VWR | MISC-CLINICAL |
| Mouse monoclonal [2B3] to RUNX3 | Abcam | ab135248 |
| Rabbit monoclonal [EP3251] to Pax2 | Abcam | ab79389;RRID:AB_1603338 |
| ASCL2 Antibody (C terminus) LS-C126891 | LifeSpan Biosciences | LS-C126891-100;RRID:AB_10831114 |
| Cytokeratin 15 Antibody | Novus Biologicals | NBP1-85602;RRID:AB_11037933 |
| Cox2 (D5H4) XP Rabbit mAb | Cell Signaling Technology | 12282S;RRID:AB_2571729 |
| Purified Mouse Anti-E-Cadherin | Fisher/BD Biosciences | 3700S |
| PTGER4/EP4 Antibody | Novus Biologicals | NBP1-84833;RRID:AB_11019807 |
| p23/PTGES3 Antibody (JJ3) | Novus Biologicals | NB300-576;RRID:AB_10000921 |
| HSP90 beta Antibody | Novus Biologicals | NBP1-77563;RRID:AB_11011699 |
| Anti-HSP90aa1 | Novus Biologicals | NB120-2928;RRID:AB_790272 |
| B7-H1/PD-L1/CD274 Antibody (M - 0.1 MG) | Fisher/Novus Biologicals | NBP1-432620 |
| Anti-Prostaglandin E2 antibody | Abcam | ab2318;RRID:AB_302974 |
| Cleaved Caspace-3 (Asp175) Antibody, 100 μl (10 western blots) | Cell Signaling Technology | 9661S;RRID:AB_2341188 |
| BrdU/CldU antibody | Fisher/Novus Biologicals | NB500-169;RRID:AB_10002608 |
| Cox2 Antibody | Cell Signaling Technology | 4842;RRID:AB_2084968 |
| ANTI-CD44 ANTIGEN ISOFORM 4 | Fisher | PIPA521419 |
| CD44v6 Monoclonal Antibody (9A4) | Fisher | MA1-81995;RRID:AB_928369 |
| Phospho-Akt (Ser473) (D9E) XP® | Cell Signaling Technology | 4060S;RRID:AB_2315049 |
| Thermo Scientific Lab Vision Ki-67, Rabbit Monoclonal Antibody | ThermoFisher | RM-9106-S1;RRID:AB_149792 |
| Anti-RIP3 antibody | Abcam | ab56164;RRID:AB_2178667 |
| Anti-CD8 antibody [YTS169.4] | Abcam | ab22378;RRID:AB_447033 |
| Annexin V-mFluor Violet 540 conjugate | AAT Bioquest | 20082 |
| CD24 Monoclonal Antibody (M1/69), eFluor 450, eBioscience | ThermoFisher/eBioscience | 48-0242-82;RRID:AB_1311169 |
| MHC Class II I-Ab Monoclonal Antibody (AF6-120.1), APC, eBioscience | ThermoFisher/eBioscience | 17-5320-82;RRID:AB_2573212 |
| Anti-CD44 antibody [T2–F4] | Abcam | ab40983;RRID:AB_726520 |
| Thermo Scientific Lab Vision Ki-67, Rabbit Monoclonal Antibody | ThermoFisher | RM-9106-S1;RRID:AB_149792 |
| CldU antibody | Novus Biologicals | NB500-169;RRID:AB_10002608 |
| Anti-Mouse CD24 FITC 50ug | eBioscience | 11-0241-81;RRID:AB_464984 |
| Lysozyme EC 3.2.1.17 | Dako | A009902-2:RRID:AB_2341231 |
| Anti-GRP78/BiP (ET-21) antibody produced in rabbit | Sigma | G9043-200UL; RRID:AB_2279879 |
| Mouse ALCAM Phycoerythrin Affinity Purified PAb, Goat IgG (CD166 PE) | R&D Systems | FAB1172P;RRID:AB_2242595 |
| CD44 P7 | BioLegend | 103030;RRID:AB_830787 |
| Anti-Mouse CD24 eFluor® 450 (Pacific Blue® replacement) 100 ug (CD24 eF450) | eBioscience | 48-0242-82;RRID:AB_1311169 |
| goat anti-rabbit IgG, F(ab’)2-APC-Cy7 | Santa Cruz Biotechnology | sc-3847; RRID:AB_649105 |
| Anti-GRP78/BiP (ET-21) antibody produced in rabbit | Sigma | G9043-200UL;RRID:AB_2279879 |
| MTA1 (D40D1) XP® Rabbit mAb 100ul (10 western blots) | Cell Signaling Technology | 5647S;RRID:AB_10705601 |
| BrdU antibody | VWR | 95019-028 |
| CD44 | Fisher | MA1-81995;RRID:AB_928369 |
| Biological samples | ||
| Human familial adenomatous polyposis (FAP) paraffin-embedded tissue sections | University of Kansas Medical Center | Pathology & Laboratory UKMC, KC, KS 66160 |
| Human colorectal cancer (CRC) paraffin-embedded tissue sections | University of Kansas Medical Center | Pathology & Laboratory UKMC, KC, KS 66160 |
| Chemicals, peptides, and recombinant proteins | ||
| EP2 antagonist | Cayman Chemical | 14050 |
| M-CSF, Macrophage-Colony Stimulating Factor Protein, Recombinant human | Millipore | GF053 |
| EP4 antagonist | Sigma | A8227-25MG |
| Human Recombinant M-CSF, Size: 100 μg | StemCell Technologies | 78057 |
| DNase I | Fisher/Worthington | LS006361 |
| Liberase TM Research Grade | Sigma | 5401119001 |
| Celecoxib | LC Laboratories | C-1502 |
| Capecitabine, 10 g | LC Laboratories | C-2799 |
| JAG-1 (188–204), Jagged-1 (188–204), Notch Ligand” | Anaspec | AS-61298 |
| Recombinant Murine Noggin, 20 ug | PeproTech | 250-38 |
| Recombinant Murine EGF, 100 ug | PeproTech | 315-09 |
| Recombinant Mouse R-Spondin 1, CF, 250 ug | R&D Systems | 3474-RS-250 |
| Recombinant Murine Wnt-3a, 2 ug | R&D Systems | 1324-WN-002 |
| Stemolecule Thiazovivin (ROCK inhibitor), 1 mg | Reprocell | 04-0017 |
| Stemolecule CHIR99021, 2 mg | Reprocell | 04-0004 |
| IntestiCult Organoid Growth Medium (Mouse) | StemCell Technologies, Inc. | 06005 |
| Corning® Matrigel® Growth Factor Reduced (GFR) Basement Membrane Matrix, Phenol Red-free, LDEV-free, 10 mL | VWR | 356231 |
| PF3644022, 10 mg | Sigma | H6278-50MG |
| 4-HYDROXYTAMOXIFEN | VWR | 103302-538 |
| Collagenase 1A, 100mg | Sigma | C2674-100MG |
| Hyaluronidase, 100mg | Sigma | H3506-100MG |
| Evans Blue, 10 g | Sigma | E2129-10G |
| MATRIGEL MATRIX NO PHENOL 10ML | VWR/Corning | 356237 |
| Advanced DMEM/F-12 | ThermoFisher | 12634-010 |
| E4 pipette | Rainin | 17014490 |
| B-27® Serum-Free Supplement (50X), Liquid, 10 ml | ThermoFisher | 17504-044 |
| N-2 Supplement (100X), Liquid, 5 ml | ThermoFisher | 17502-048 |
| N-ACETYL-L-CYSTEINE BIOREAGENT CELL CUL& | Sigma | A9165-5G |
| Y-27632 | Sigma | Y0503-1MG |
| Stemolecule Thiazovivin 1 mg | Stemgent | 04-0017 |
| Recombinant Human R-Spondin 1, CF 25 μg | R&D Systems | 4645-RS-025/CF |
| Basement Membrane Matrix, Growth Factor Reduced (GFR), Phenol Red-free, 10ml *LDEV-Free | BD Biosciences | 356231 |
| SOLUTION BSS W/O CA+ MG+ PH-RD 1L | VWR | 12001-980 |
| Critical commercial assays | ||
| Myeloid-Derived Suppressor Cell Isolation Kit | Miltenyi Biotec | Cat# 130-094-538 |
| MACS separation using LS columns | Miltenyi Biotec | Cat# 130-042-401 |
| 10X Chromium Single Cell 3′ Library & Gel Bead Kit (v2) | 10x Genomics | Cat# PN-120267 |
| TruSeq Stranded mRNA Prep Kit | illumine | Cat# 20020594 |
| Bioanalyzer RNA Analysis | Agilent Technologies | Cat# 5067-1511 |
| Complementary DNA synthesis; High-capacity RNA to cDNA | Life Technologies | Cat# 4387406 |
| Propidium Iodide Solution | Miltenyi Biotec | 130-093-233 |
| MACS BSA Stock Solution | Miltenyi Biotec | 130-091-376 |
| Dead Cell Removal Kit | Miltenyi Biotec | 130-090-101 |
| autoMACS Rinsing Solution | Miltenyi Biotec | 130-091-222 |
| Anti-Mouse IgG1 MicroBeads | Miltenyi Biotec | 130-047-102;RRID:AB_244355 |
| CD45 MicroBeads, mouse | Miltenyi Biotec | 130-052-301;RRID:AB_2877061 |
| CD31 MicroBeads, mouse | Miltenyi Biotec | 130-097-418;RRID:AB_2814657 |
| Anti-Ter-119 MicroBeads, mouse | Miltenyi Biotec | 130-049-901 |
| Miltenyi Biotec | Miltenyi Biotec | 130-042-201 |
| Dead Cell Removal Kit | Miltenyi Biotec | 130-090-101 |
| RBC lysis buffer | Sigma | 11814389001 |
| Annexin V-mFluor Violet 540 conjugate | AAT Bioquest | 20082 |
| CD11b MicroBeads | Miltenyi Biotec | 130-049-601 |
| CD11c MicroBeads | Miltenyi Biotec | 130-108-338 |
| Trypan blue solution 0.4%, 100ml | ThermoFisher | 15250061 |
| KnockOut Serum Replacement | ThermoFisher | 10828028 |
| Restore PLUS Western Blot Stripping Buffer | ThermoFisher | 46430 |
| Deposited data | ||
| 10X single cell (sc) RNA-seq data | Gene Expression Omnibus (GEO: GSE136256) | |
| Experimental models: Organisms/strains | ||
| C57BL/6 | Jackson Laboratories | 000664;RRID:IMSR_JAX:000664 |
| C57BL/6J-ApcMin/J (Apc-Min) | Jackson Laboratories | 002020;RRID:IMSR_JAX:002020 |
| BALB/cJ (BALB/c) | Jackson Laboratories | 000651;RRID:IMSR_JAX:000651 |
| B6.129X1-Gt(ROSA)26Sortm1(EYFP)Cos/J (R26-YFP) | Jackson Laboratories | 006148;RRID:IMSR_JAX:006148 |
| B6;129-Bmi1tm1(cre/ERT)Mrc/J (Bmi1-CreER) | Jackson Laboratories | 010531;RRID:IMSR_JAX:010531 |
| B6.129P2-Lgr5tm1(cre/ERT2)Cle/J (LGR5KI) | Jackson Laboratories | 008875;RRID:IMSR_JAX:008875 |
| Software and algorithms | ||
| FlowJo Software v 7.0 | FlowJo | https://www.flowjo.com/solutions/flowjo/downloads |
| ImageJ | OpenSource | https://imagej.nih.gov/ij/download.html |
| GraphPad Prism v 8.1.2 | Prism | https://www.graphpad.com/ |