KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Mouse monoclonal anti-Actin | Aves Labs | ACT-1010 |
| Mouse monoclonal anti-HA | BioLegend | 901502 |
| Chicken polyclonal anti-HA | AVES Labs | ET-HA100 |
| Mouse monoclonal anti-FLAG | Sigma | F3165 |
| Mouse monoclonal anti-Rhodopsin (1D4) | ThermoFisher | MA1–722 |
| Rabbit polyclonal anti-SEMA6A | ThermoFisher | PA5–81009 |
| Purified Rat Anti-Mouse CD31 | BD Pharmingen | 553370 |
| Chemicals, Peptides, and Recombinant Proteins | ||
| All the other chemicals | Sigma | N/A |
| PolyJet | SignaGen | SL100688 |
| PEIMax | Polysicences | 24765–1 |
| Lipofectamine RNAiMAX | ThermoFisher | 13778150 |
| Polybrene | Santa Cruz | sc-134220 |
| Dulbecco’s Modified Eagle Medium | Life technologies | Cat#11995–065 |
| Fetal bovine serum | Life technologies | Cat#26140–079 |
| Penicillin/streptomycin | Life technologies | Cat#15140–122 |
| F-12K Medium | ATCC | 30–2004 |
| Heparin | Sigma | H3393 |
| Endothelial cell growth supplement | BD Biosciences | # 354006 |
| Puromycin | ThermoFisher | A1113830 |
| Blasticidin S | RPI | B12150.01 |
| Hygromycin B | EMD Millipore | 400051 |
| Protease Inhibitor Cocktail | Roche | 4693159001 |
| Nitrocellulose membrane | GE Healthcare | 10600002 |
| DAPI-containing mounting medium | SouthernBiotech | 0100–20 |
| Endo H | NEB | P0702 |
| Recombinant human Fc-tagged SEMA6A | R&D Systems | 1146-S6 |
| Recombinant human Fc-tagged SEMA6B | R&D Systems | 2264-S6 |
| Recombinant human Fc-tagged SEMA6C | R&D Systems | 2219-S6 |
| Recombinant human Fc-tagged SEMA6D | R&D Systems | 2095-S6 |
| Recombinant human Fc-tagged SEMA4D | R&D Systems | 5235-S4 |
| Recombinant human Fc-tagged SEMA5A | R&D Systems | 6584-S5 |
| Recombinant human Fc-tagged SEMA7A | R&D Systems | 1835-S3 |
| Recombinant human Fc-tagged CRD2 | R&D Systems | 1307-FZ |
| Recombinant human Fc-tagged IgG | R&D Systems | 110-HG |
| TcsL | This paper | N/A |
| TcsL1285–1804 | This paper | N/A |
| TcsL1285–1804-HA | This paper | N/A |
| TcsL1865–2364 | This paper | N/A |
| TcdB-FBD | Rongsheng Jin | Chen et al., 2018 |
| TcsL(TcdB1431–1602) | This paper | N/A |
| TcdB(TcsL1431–1601) | This paper | N/A |
| mSEMA6A-ECD | This paper | N/A |
| mSEMA6A-His | This paper | N/A |
| mSEMA6C-ECD | This paper | N/A |
| Critical Commercial Assays | ||
| Gibson Assembly | NEB | E2621 |
| Bacillus Expression Systems | MoBiTec GmbH | BMEG04 |
| BCA assay kit | ThermoFisher | 23225 |
| Genomic DNA extraction kit | Qiagen | 13323 |
| Enhanced chemiluminescence kit | ThermoFisher | 34080 |
| Experimental Models: Cell Lines | ||
| HeLa | ATCC | CCL-2 |
| A549 | ATCC | CRM-CCL-185 |
| 5637 | ATCC | HTB-9 |
| ACHN | ATCC | CRL-1611 |
| Caco-2 | ATCC | HTB-37 |
| U2OS | ATCC | HTB-96 |
| Huh7 | Y. Matsuura | N/A |
| HEK293T | ATCC | CRL-3216 |
| HUVECs | Lonza | 00191027 |
| Expi293F | ThermoFisher | A14527 |
| A549-SEMA6A-KO-Mix-I | This paper | N/A |
| A549-SEMA6A-KO-Mix-II | This paper | N/A |
| A549-SEMA6B-KO-Mix-I | This paper | N/A |
| A549-SEMA6B-KO-Mix-II | This paper | N/A |
| A549-SEMA6C-KO-Mix | This paper | N/A |
| A549-SEMA6D-KO-Mix-I | This paper | N/A |
| A549-SEMA6D-KO-Mix-II | This paper | N/A |
| A549-EMB-KO-Mix | This paper | N/A |
| A549-RTN4RL2-KO-Mix | This paper | N/A |
| A549-GPR87-KO-Mix | This paper | N/A |
| A549-SLC27A5-KO-Mix | This paper | N/A |
| A549-PCDHGB6-KO-Mix | This paper | N/A |
| A549-TBC1D3-KO-Mix | This paper | N/A |
| A549-ATPV0B-KO-Mix | This paper | N/A |
| A549-SLC25A31-KO-Mix | This paper | N/A |
| A549-PPAT-KO-Mix | This paper | N/A |
| A549-ACTG2-KO-Mix | This paper | N/A |
| A549-SEMA6A/6B-KO-Mix-I | This paper | N/A |
| A549-SEMA6A/6B-KO-Mix-II | This paper | N/A |
| HeLa-SEMA6A-KO-Mix-I | This paper | N/A |
| HeLa-SEMA6A-KO-Mix-II | This paper | N/A |
| HeLa-SEMA6B-KO-Mix-I | This paper | N/A |
| HeLa-SEMA6B-KO-Mix-II | This paper | N/A |
| HeLa-SEMA6C-KO-Mix | This paper | N/A |
| HeLa-SEMA6D-KO-Mix-I | This paper | N/A |
| HeLa-SEMA6D-KO-Mix-II | This paper | N/A |
| HeLa-SEMA6A/6B-KO-Mix-I | This paper | N/A |
| HeLa-SEMA6A/6B-KO-Mix-II | This paper | N/A |
| HeLa + SEMA6A-FLAG-C | This paper | N/A |
| HeLa + SEMA6B-FLAG-C | This paper | N/A |
| HeLa + SEMA6C-FLAG-C | This paper | N/A |
| HeLa + SEMA6D-FLAG-C | This paper | N/A |
| 5637 + SEMA6A-FLAG-C | This paper | N/A |
| 5637 + SEMA6B-FLAG-C | This paper | N/A |
| 5637 + SEMA6C-FLAG-C | This paper | N/A |
| 5637 + SEMA6D-FLAG-C | This paper | N/A |
| Experimental Models: Mice | ||
| CD-1 strain | Charles River | #022 |
| Oligonucleotides | ||
| See Table S1 for oligonucleotides used in these studies | This paper | N/A |
| Recombinant DNA | ||
| SEMA6A cDNA | Sino Bio | HG11189-M |
| SEMA6B cDNA | GE Dharmacon | 40147342 |
| SEMA6C cDNA | Sino Bio | HG23441-UT |
| SEMA6D cDNA | R&D Systems | RDC2156 |
| pcDNA3.1 | ThermoFisher | V80020 |
| pET28a | Novagen | 69864 |
| pHIS1522 | MoBiTec GmbH | BMEG12 |
| pMD2.G | Addgene | #12259 |
| pSPAX2 | Addgene | #12260 |
| LentiGuide-puro | Addgene | #52963 |
| Lenti-SpCas9 blast | Addgene | #104997 |
| pLenti-Hygro | Addgene | #17484 |
| Sema6a.a-Fc-His | Addgene | #72163 |
| Sema6c-Fc-His | Addgene | #72167 |
| pRK5-mFzd2–1D4 | Addgene | 42264 |
| GeCKO-V2 sgRNA library | Addgene | #1000000049 |
| pET28a-TcdB-FBD | Liang Tao | Chen et al., 2018 |
| Lentiguide-SEMA6A-I | This paper | N/A |
| Lentiguide-SEMA6A-II | This paper | N/A |
| Lentiguide-SEMA6B-I | This paper | N/A |
| Lentiguide-SEMA6B-II | This paper | N/A |
| Lentiguide-SEMA6C | This paper | N/A |
| Lentiguide-SEMA6D-I | This paper | N/A |
| Lentiguide-SEMA6D-II | This paper | N/A |
| Lentiguide-EMB | This paper | N/A |
| Lentiguide-RTN4RL2 | This paper | N/A |
| Lentiguide-GPR87 | This paper | N/A |
| Lentiguide-SLC27A5 | This paper | N/A |
| Lentiguide-PCDHGB6 | This paper | N/A |
| Lentiguide-TBC1D3 | This paper | N/A |
| Lentiguide-ATP6V0B | This paper | N/A |
| Lentiguide-SLC25A31 | This paper | N/A |
| Lentiguide-PPAT | This paper | N/A |
| Lentiguide-ACTG2 | This paper | N/A |
| pcDNA3.1-SEMA6A-FLAG-C | This paper | N/A |
| pcDNA3.1-SEMA6A-ECD-His | This paper | N/A |
| LentiHygro-SEMA6A-FLAG-C | This paper | N/A |
| pcDNA3.1-SEMA6B-FLAG-C | This paper | N/A |
| LentiHygro-SEMA6B-FLAG-C | This paper | N/A |
| pcDNA3.1-SEMA6C-FLAG-C | This paper | N/A |
| LentiHygro-SEMA6C-FLAG-C | This paper | N/A |
| pcDNA3.1-SEMA6D-FLAG-C | This paper | N/A |
| LentiHygro-SEMA6D-FLAG-C | This paper | N/A |
| pET28a-TcsL-1285-1804 | This paper | N/A |
| pET28a-TcsL-1856-2364 | This paper | N/A |
| pET28a-TcsL-1285-1804-HA | This paper | N/A |
| pET28a-TcsL(TcdB-1431-1602) | This paper | N/A |
| pET28a-TcdB(TcsL-1431-1601) | This paper | N/A |
| Software and Algorithms | ||
| OriginPro v8.5 | OriginLab | https://www.originlab.com/ |
| Excel | Microsoft | https://products.office.com/en-us/home |
| BLItz pro. Software Version 1.1.0.29 | ForteBio | https://www.fortebio.com/products/label-free-bli-detection/personal-assay-blitz-system |
| ImageJ Version 1.52o | Image J | https://imagej.nih.gov/ij/ |
| Others | ||
| Fluorescence microscope | Olympus | IX51 |
| Spinning Disk Confocal Microscope | Olympus | DSU-IX81 |
| Fuji LAS3000 imaging system | Fuji | LAS3000 |
| Personal assay BLItz System | ForteBio | BLItz System |
| Dip and Read Anti-Human IgG Fc Capture | ForteBio | 18–5060 |