KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
|
| ||
| Antibodies | ||
|
| ||
| TRPM2 | Novus | NB110–81601; RRID: AB_1147355 |
| GluN2A | Cell Signaling Technology | 4205S; RRID: AB_2112295 |
| GluN2B | Cell Signaling Technology | 4207S; RRID: AB_1264223 |
| PKC-γ | Cell Signaling Technology | 59090S; RRID: AB_2799557 |
| Pan-cadherin | Cell Signaling Technology | 4068S; RRID: AB_2158565 |
| GAPDH | Cell Signaling Technology | 5174S; RRID: AB_10622025 |
| GST-Tag | Cell Signaling Technology | 2622S; RRID: AB_331670 |
| His-Tag | Cell Signaling Technology | 12698S; RRID: AB_2744546 |
| Anti-mouse IgG, HRP-linked Antibody | Cell Signaling Technology | 7076; RRID: AB_330924 |
| Anti-rabbit IgG, HRP-linked Antibody | Cell Signaling Technology | 7074; RRID: AB_2099233 |
|
| ||
| Bacterial and virus strains | ||
|
| ||
| Escherichia coli BL21(DE3) cells | NEB (New England Biolabs) | C2527 |
|
| ||
| Chemicals, peptides, and recombinant proteins | ||
|
| ||
| Tetrazolium chloride | Sigma-Aldrich | T-8877 |
| NMDA | Tocris | 0114 |
| AP5 | Cayman Chemical | 14539 |
| MK-801 | Sigma-Aldrich | M107 |
| PMA | Sigma-Aldrich | 524400 |
| H2O2 | Thermal Fisher Scientific | 200745 |
| EGTA | Cayman Chemical | 11706 |
| BAPTA-AM | Cayman Chemical | 15551 |
| NP40 | Thermal Fisher Scientific | 28324 |
| Triton™ X-100 | Thermal Fisher Scientific | T-9284 |
| Bovine Serum Albumin | Sigma-Aldrich | 9048–46-8 |
| isopropyl-D-1-thiogalactopyranoside (IPTG) |
Sigma-Aldrich | I5502–10G |
| Neurobasal® Medium | Thermal Fisher Scientific | 21103–049 |
| Pluronic™ F-127 | Thermal Fisher Scientific | P3000MP |
| Fura2-AM | Thermal Fisher Scientific | F1221 |
| DAF-FM | Thermal Fisher Scientific | D23844 |
| MitoSOX-Red | Thermal Fisher Scientific | M36008 |
| Rhodamine-123 | Thermal Fisher Scientific | R302 |
| Proteinase inhibitors | Sigma-Aldrich | 539131–10VL |
| Phosphatase inhibitors | Thermal Fisher Scientific | 78428 |
| Protein A/G PLUS-Agarose | Santa Cruz Biotechnology | sc-2003 |
| 2x Laemmli Sample Buffer | BIO-RAD | 1610737 |
| TAT-SC (sequence: YGRKKRRQRRR VILLKDHTLEYPVF) | GenScript Biotech | N/A |
| TAT-M2PBM (sequence: YGRKKRRQRRR WGLDVPNLLISVTGGA) | GenScript Biotech | N/A |
| DpnI | BioLabs | R0176L |
|
| ||
| Critical commercial assays | ||
|
| ||
| PKCγ Kinase Enzyme System | Promega | V3391 |
| PfuUltra HF | Agilent | 600380–51 |
| Pierce™ Rapid Gold BCA Protein Assay Kit | Thermal Fisher Scientific | A53225 |
| Pierce® Cell Surface Protein Isolation Kit | Thermal Fisher Scientific | 89881 |
| ProteoExtract™ Native Membrane Protein | Calbiochem | 444810 |
| Extraction Kit | ||
|
| ||
| Experimental models: Cell lines | ||
|
| ||
| HEK293T cells | ATCC | CRL-3216 |
|
| ||
| Experimental models: Organisms/strains | ||
|
| ||
| C57BJ6 mice | JAX | 000664 |
| Global TRPM2 knockout mice | Yasuo Mori’s lab at Kyoto University | N/A |
|
| ||
| Oligonucleotides | ||
|
| ||
| Primers for mutagenesis and subcloning; see Table S1 |
This study | N/A |
|
| ||
| Recombinant DNA | ||
|
| ||
| GluN1A | Luo et al.40 | Addgene, 17928 |
| GluN2A | Luo et al.40 | Addgene, 17924 |
| GluN2B | Luo et al.40 | Addgene, 17925 |
| PKC-γ | Oancea et al.41 | Addgene, 112266 |
| PKC-γ-DN | Soh et al.42 | Addgene, 21239 |
| CKAR | Violin et al.17 | Addgene, 14860 |
| pET15b vector containing a removable TEV protease recognition site |
Zong et al.6 MilliporeSigma |
69661–3 |
| pGEX-4T3 vector containing a removable | Zong et al.6 | V010916 |
| tobacco etch virus (TEV) protease recognition site |
NovoPro | |
| pcDNA4/TO-FLAG-hTRPM2 | Sharenberg AM at University of Washington | N/A |
|
| ||
| Software and algorithms | ||
|
| ||
| Prism 9 | Graphpad | https://www.graphpad.com/ |
| Photoshop 2020 | Adobe | https://www.adobe.com/ |
| Illustrator 2020 | Adobe | https://www.adobe.com/ |
| NIS-Elements | Nikon | N/A |
| ImageJ | Schneider et al.43 | https://imagej.net/ij/ |
|
| ||
| Other | ||
|
| ||
| Rotarod | Maze | https://maze.conductscience.com |
| Avestin EmulsiFlex C3 | ATA Scientific Instruments | https://www.atascientific.com.au/products/avestin-emulsiflex-c3 |
| Glutathione Sepharose 4B column | GE Healthcare | Discontinued |
| Ni2+-nitrilotriacetic acid (NTA) column | GE Healthcare | Discontinued |
| Amicon stirred ultrafiltration cell unit | EMD Millipore | UFSC40001 |
| CoolSNAP HQ2 | Teledyne Photometrics | https://www.photometrics.com |
| Ultrasonic cleaner | Thermal Fisher Scientific | CPX952136R |
| Axopatch 200B amplifier | Molecular Devices | https://www.moleculardevices.com/products/axon-patch-clamp-system/amplifiers |