KEY RESOURCES TABLE
| REAGENT or SOURCES | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| 6xHis tag | Cell Signaling | 2365 |
| 6PGD | Cell Signaling | 13389S |
| Biotin goat MS IgG (IHC) | BD Pharmingen | 553999 |
| CD31 | Novartis | NBP2–44342 |
| Anti-mouse/Anti-Rabbit Envision Flex HRP labeled Polymer (IHC) | DAKO | K8000 |
| Flag tag | Cell Signaling | 14793S |
| G6PD (WB) | Cell Signaling | 12263S |
| G6PD (IF, IHC, IP) | Proteintech | 66373–1-Ig |
| GAPDH | Santa Cruz | sc-47724 |
| Goat anti-mouse IgG (IF 488) | Fisher Scientific | A32723 |
| Goat anti-rabbit (IF 594) | Fisher Scientific | A32731 |
| GST tag | Santa Cruz | Sc-138 |
| HRP-anti-mouse IgG (WB) | Cell Signaling | 7076S |
| HRP-anti-rabbit IgG (WB) | Cell Signaling | 7074S |
| Ki67 | BD Pharmingen | 550609 |
| NADK1(WB) | Cell Signaling | 55948S |
| NADK1(IP) | Santa Cruz | sc-100347 |
| NADK1(IF) | Proteintech | 15548–1-AP |
| Mouse IgG (IP) | Santa Cruz | Sc-2025 |
| p53 | Santa Cruz | Sc-126 |
| PRPS1/2 | Santa Cruz | sc-100822 |
| H-RAS | Santa Cruz | sc-35 |
| TALDO1 | Abcam | ab137629 |
| TERT | Santa Cruz | sc-377511 |
| TKT | Cell Signaling | 8616 |
| Chemicals, Recombined Proteins, and Inhibitors | ||
| (±)-6-Hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid (Trolox) | Sigma-Aldrich | 238813 |
| (−)-Riboflavin (cell culture) | Sigma-Aldrich | R9504 |
| 2′,7′-Dichlorofluorescin diacetate (DCFDA) | Sigma-Aldrich | D6883 |
| 3X FLAG® Peptide | Sigma-Aldrich | F4799 |
| 4,6-diamidino-2-phenylindole (DAPI) | Vector Laboratories | H-1200 |
| 6xHis-tagged human NADK1 protein | Sigma-Aldrich | SRP8020 |
| β-NAD | Sigma-Aldrich | N7004 |
| β-NADH phosphate disodium salt (NADP+) | Sigma-Aldrich | N5755 |
| β-NADPH | Sigma-Aldrich | N7505 |
| Adenosine triphosphate salt (ATP) | Sigma-Aldrich | A1852 |
| REAGENT or SOURCES | SOURCE | IDENTIFIER |
| ANTI-FLAG® M2 Affinity Gel | Sigma-Aldrich | A2220 |
| BamH1 (restriction enzyme) | New England Biolabs | R3136 |
| Bovine Serum Albumin (BSA) | Sigma-Aldrich | A2153 |
| Crystal violet | Sigma-Aldrich | C0775 |
| Collagenase/ Hyaluronidase (10 x) | Fisher Scientific | NC9694308 |
| D-G6P | Sigma-Aldrich | G7250 |
| Dako Dual Endogenous Enzyme Block | DAKO | S2003 |
| Dialyzed FBS | Sigma-Aldrich | F0392 |
| Disuccinimidyl suberate (DSS) | Sigma-Aldrich | S1885 |
| Dulbecco’s MEM (DMEM) Base (w/o L-Glutamine, L-Cystine, Glucose, Phenol Red, Sodium Pyruvate) | United States Biological | D9815 |
| Dpn1 (restriction enzyme) | New England Biolabs | R0176 |
| FBS | Sigma-Aldrich | TMS-013 |
| FK 866 hydrochloride | Tocris | 4808 |
| Folic acid (cell culture) | MP Biomedicals | 0219466505 |
| Glucose Solution (cell culture) | Fisher Scientific | A2494001 |
| HBSS (1x) | Fisher Scientific | 14025092 |
| HIS-Select® Nickel Affinity Gel | Sigma-Aldrich | P6611 |
| Human G6PD protein (From E. coli) | Sigma-Aldrich | SRP6505 |
| lipofectamine 2000 | Fisher Scientific | 11668019 |
| lipofectamine RNAimax | Fisher Scientific | 13778075 |
| Low temperature melting agarose | Lonza | 50101 |
| MEGM™ BulletKit™ Growth Media | Lonza | CC-3162 |
| MgCl2 | Sigma-Aldrich | 1374248 |
| Mouse diet | LabDiet | 5010 |
| Nicotinamide-2,6,7-13C3-(pyridyl-15N) | Sigma-Aldrich | 809799 |
| Nucleosides | ||
| Deoxy Adenosine monohydrate | DN-1001 | |
| Deoxy Cytidine | DN-1002 | |
| Deoxy Guanosine | DN-1003 | |
| Thymidine | ChemGenes Corp | DN-1004 |
| Ribo Adenosine | RP-1183 | |
| Ribo Cytidine | RP-1184 | |
| Ribo Guanosine | RP-1185 | |
| Ribo Uridine | RP-1186 | |
| Novocastra Epitope Retrieval Solutions | Leica | AR9640 |
| N-Acetyl Cysteine (NAC) | Sigma-Aldrich | A9165 |
| Paraformaldehyde (PFA) | Sigma-Aldrich | P6148 |
| Pierce™ Glutathione Agarose | Fisher Scientific | 16100 |
| Poly (2-hydroxyethyl methacrylate) (ploy-HEMA) | Sigma-Aldrich | P3932 |
| Propidium Iodide (PI) | Sigma-Aldrich | P4864 |
| Protease inhibitor cocktail | Sigma-Aldrich | P8340 |
| REAGENT or SOURCES | SOURCE | IDENTIFIER |
| Protein A/G agarose beads | Santa Cruz | sc-2003 |
| SalI (restriction Enzyme) | New England Biolabs | R0138 |
| Sodium pyruvate solution (cell culture) | Sigma-Aldrich | S8636 |
| Streptavidin-Horseradish-Peroxidase | BD Pharmingen | 554066 |
| T4 ligase | Fisher Scientific | EL0014 |
| Thiamine pyrophosphate (cell culture) | Sigma-Aldrich | C8754 |
| Tris-HCl | Sigma-Aldrich | 93363 |
| Trypan blue solution | Fisher Scientific | 15–250-061 |
| Critical Commercial Assays | ||
| Glucose 6 Phosphate Dehydrogenase Activity Assay Kit (Fluorometric) | Abcam | ab176722 |
| Bio-Rad protein assay kit I | Bio-Rad | 5000001 |
| NADP/NADPH Quantitation Colorimetric Kit | BioVision | K347 |
| NADP/NADPH-GloTM assays kit | Promega | G9081 |
| ATP Assay Kit (Colorimetric/Fluorometric) |
Abcam | ab83355 |
| FITC Annexin V Apoptosis Detection Kit II | BD Pharmingen | 556570 |
| Cell Counting Kit-8 | Sigma | 96992 |
| TRIzol | Fisher Scientific | 15596 |
| High-Capacity cDNA Reverse Transcription Kit | Fisher Scientific | 43688 |
| 2X GoTaq® Master Mixes | Promega | M7122 |
| BrdU Cell Proliferation Assay Kit | BioVision | K306 |
| Enhanced chemiluminescence system | Amersham Biosciences | RPN2232 |
| Oligonucleotides | ||
| siRNA for G6PD (h) | Santa Cruz | sc-60667 |
| siRNA for PRPS1/2 (m) | Santa Cruz | sc-62895 |
| siRNA for PRPS1/2 (h) | Santa Cruz | sc-62894 |
| shRNA for G6PD (m) | UPENN HTS core | TRCN0000041446 |
| shRNA for G6PD (h) | Open Biosystem | RHS3979–9595543 |
| shRNA-1 for PRPS1 (m) | UPENN HTS core | TRCN0000024886 |
| shRNA-2 for PRPS1 (m) | UPENN HTS core | TRCN0000024888 |
| shRNA-2 for PRPS1 (h) | UPENN HTS core | TRCN0000367679 |
| shRNA-3 for PRPS1 (h) | UPENN HTS core | TRCN0000355958 |
| shRNA-2 for PRPS2 (m) | UPENN HTS core | TRCN0000025539 |
| shRNA-3 for PRPS2 (m) | UPENN HTS core | TRCN0000025542 |
| shRNA-1 for PRPS2 (h) | UPENN HTS core | TRCN0000010131 |
| shRNA-5 for PRPS2 (h) | UPENN HTS core | TRCN0000010133 |
| shRNA for TALDO1 (h) | Sigma-Aldrich | SHCLNG-NM_006755 |
| shRNA for TKT (h) | Sigma-Aldrich | SHCLNG-NM_001064 |
| Forward primer for human GAPDH | 5’ GTCTCCTCTGACTTCAACAGCG 3’ | |
| Reverse primer for human GAPDH | 5’ ACCACCCTGTTGCTGTAGCCAA 3’ | |
| REAGENT or SOURCES | SOURCE | IDENTIFIER |
| Experimental Models: Organism/Strains | ||
| Athymic nude mice | Homozygous for Foxn1nu | Jackson Laboratory |
| Cell lines | ||
| NIH3T3 (unknown) | ATCC | ATCC® CRL-1658™ |
| BJ (male) | ATCC | ATCC® CRL-2522™ |
| PHMLEB (female) | R. A. Weinberg | Ref (Elenbaas et al., 2001) |
| HEK293T (female) | ATCC | ATCC® CRL-11268™ |
| Plasmids | ||
| 6PGD | pReceiver-Lv151 | EX-A6274-Lv151 |
| Ampho | pCL (Novus Biologicals) | NBP2–29541 |
| dR8.91 | pCMV (Nova lifetech) | PVT2323 |
| Flag-control | pRK5 | Ref (Du et al., 2013) |
| Flag-G6PD | pRK5 | Ref (Du et al., 2013) |
| Flag-G6PDm1 (R365A and K366A) | pRK5 (Site-directed mutagenesis) | N/A |
| Flag-G6PDm2 (K171Q) | PRK5 (Site-directed mutagenesis) | N/A |
| Flag-NADK1 | pRK5 | N/A |
| Flag-NADK1∆N | pRK5 | N/A |
| G6PD | pReceiver-Lv105 | EX-Z2649-Lv105 |
| G6PD | pBABE-puro | Ref (Du et al., 2013) |
| G6PDm1 (R365A and K366A) | pBABE-puro (Site-directed mutagenesis) | N/A |
| G6PDm2 (K171Q) | pBABE-puro (Site-directed mutagenesis) | N/A |
| GST-control | pRK5 | Ref (Guo et al., 2014) |
| GST-NADK1 | pRK5 | N/A |
| GST-NADK1N | pRK5 | N/A |
| hRASV12 | pBABE-puro (Addgene) | #9051 |
| hTERT | pBABE-hygro (Addgene) | #1773 |
| NADK1 | pReceiver-Lv152 | EX-Z7851-Lv152 |
| T antigens | pBABE-neo (Addgene) | #10891 |
| TALDO1 | pReceiver-Lv152 | EX-C0584-Lv152 |
| TKT | pReceiver-Lv105 | EX-M0713-Lv105 |
| VSV-G | pCMV (Addgene) | #8454 |
| Software and Algorithms | ||
| MORPHEUS | Broad Institute | https://software.broadinstitute.org/morpheus/ |
| Image Pro plus7 | Media Cybernetics | N/A |
| ZEN lite | Carl Zeiss Microscopy | N/A |
| BD Accuri C6 | BD Pharmingen | N/A |
| MetaboAnalyst | Open source project | https://www.metaboanalyst.ca/ |
| Image J | NIH | https://imagej.nih.gov/ij/ |
| Fiji ImageJ | Open source project | https://imagej.net/Using_Fiji |
| JACoP plugin | JACoP_.java | https://imagej.nih.gov/ij/plugins/track/jacop2.html |
| MAVEN software35 | Maven Software | N/A |
| GraphPad Prism 8 | Graph Pad Software | N/A |