Key resources table
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Anti-PGC-1α | Millipore Sigma | RRID:AB_10807905 |
| Anti-GOT1 | ProSci | RRID:AB_10912319 |
| Anti-GOT2 | Thermo Fisher Scientific | RRID:AB_2545048 |
| Anti-MDH1 | Santa Cruz Biotechnology | RRID:AB_10609257 |
| Anti-MDH2 | Thermo Fisher Scientific | RRID:AB_11156233 |
| Anti-UCP1 | Commins, S.P. et al. | N/A |
| Anti-GLUT1 | Cell Signaling Technology | RRID:AB_2687899 |
| Anti-p-AMPKα (T172) | Cell Signaling Technology | RRID:AB_330330 |
| Anti-AMPKα | Cell Signaling Technology | RRID:AB_330331 |
| Anti-p-ACC (S79) | Cell Signaling Technology | RRID:AB_330337 |
| Anti-ACC | Cell Signaling Technology | RRID:AB_2219400 |
| Anti-p-PDH E1α (S232) | Millipore Sigma | RRID:AB_10616070 |
| Anti-PDH E1α | Cell Signaling Technology | RRID:AB_2162926 |
| Anti-GAPDH | Abcam | RRID:AB_307275 |
| Anti-VDAC | Abcam | RRID:AB_2214787 |
| Anti-α-tubulin | Abcam | RRID:AB_2241126 |
| Anti-β-actin | Abcam | RRID:AB_445482 |
| Chemicals, peptides, and recombinant proteins | ||
| CL 316243 disodium salt | TOCRIS | Cat#1499 |
| D-(+)-Glucose | Sigma-Aldrich | Cat#G-6152 |
| Insulin from Bovine Pancreas | Sigma-Aldrich | Cat#I-5500 |
| T3 | Sigma-Aldrich | Cat#T-2877 |
| 3-Isobutyl-1-methylxanthine | Sigma-Aldrich | Cat#I-5879 |
| Dexamethasone | Sigma-Aldrich | Cat#D-1756 |
| Indomethacin | Sigma-Aldrich | Cat#I-7378 |
| (−)-Isoproterenol (+)-bitartrate salt | Sigma-Aldrich | Cat#I-2760 |
| DMEM, high glucose, pyruvate, no glutamine | Thermo Fisher Scientific | Cat#10-313-039 |
| DMEM, no glucose, no glutamine, no phenol red | Thermo Fisher Scientific | Cat#A1443001 |
| FBS | Cytiva | Cat#SH30396.03 |
| Penicillin-Streptomycin (10,000 U/mL) | Thermo Fisher Scientific | Cat#15-140-122 |
| L-Glutamine (200 mM) | Thermo Fisher Scientific | Cat#25-030-081 |
| Formalin solution, neutral buffered, 10% | Sigma-Aldrich | Cat#HT501128 |
| Glycine | Bio-Rad Laboratories | Cat#161-0718 |
| Trizma Base | Sigma-Aldrich | Cat#T-1503 |
| Trizma Hydrochloride | Sigma-Aldrich | Cat#T-5941 |
| Sodium Chloride | Fisher Scientific | Cat#S271-500 |
| Manganese Chloride Tetrahydrate | Sigma-Aldrich | Cat#M-9522 |
| Nonidet-P40 (NP-40) | US Biological | Cat#N-3500 |
| Protease Inhibitor tablets | Pierce | Cat#88265 |
| SDS | Bio-Rad Laboratories | Cat#161-0302 |
| EDTA Disodium Salt | Fisher Scientific | Cat#BP120-500 |
| Sodium Bicarbonate | Fisher Scientific | Cat#S233-500 |
| Albumin Bovine Fraction V Fatty Acid Free | MP Biomedicals | Cat#152401 |
| O-(Carboxymethyl)hydroxylamine hemihydrochloride (AOA) | Sigma | Cat#C13408 |
| Iodoacetate | Alfa Aesar | Cat#A12410-14 |
| Sodium Pyruvate | Sigma-Aldrich | Cat#P-2256 |
| L-(−)-Malic acid | Sigma-Aldrich | Cat#M-1000 |
| Rotenone | Sigma-Aldrich | Cat#R-8875 |
| Antimycin A | Sigma-Aldrich | Cat#A-8674 |
| Sucrose | Fisher Scientific | Cat#S2-500 |
| Dipotassium Phosphate | Fisher Scientific | Cat#S80146-3 |
| Potassium Chloride | Fisher Scientific | Cat#P333-500 |
| L-Carnitine hydrochloride | Sigma-Aldrich | Cat#C-0283 |
| DL-Dithiothreitol (DTT) | Promega | Cat#V-3151 |
| β-Nicotinamide adenine dinucleotide hydrate | Sigma-Aldrich | Cat#N-7004 |
| Adenosin 5’-triphosphate (sodium salt) | Cayman Chemical Company | Cat#14498 |
| Coenzyme-A | Cayman Chemical Company | Cat#16147 |
| Palmitic Acid | Sigma-Aldrich | Cat#P-0500 |
| 70% Perchloric Acid | Sigma-Aldrich | Cat#311421 |
| Sodium Hydroxide | Fisher | Cat#S318-500 |
| Thiamine pyrophosphate (TPP) | Sigma | Cat#C-8754 |
| HEPES (1M) | Thermo Fisher Scientific | Cat#15-630-080 |
| Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid | Sigma-Aldrich | Cat#E-3889 |
| α-Cyano-4-hydroxycinnamic acid (CHC) | Sigma-Aldrich | Cat#476870 |
| 2-Deoxyglucose (2-DG) | Sigma-Aldrich | Cat#D-8375 |
| Cytochalasin B | Sigma-Aldrich | Cat#C-6762 |
| Oligomycin from Steptomyces diaststochromgenes | Sigma-Aldrich | Cat#O-4876 |
| iTaq Universal SYBR Green Supermix | Bio-Rad Laboratories | Cat#1725125 |
| D-[4-2H] glucose | Omicron Biochemicals | Cat#GLC-035 |
| 15N-valine | Sigma-Aldrich | Cat#490172 |
| 1-14C]-palmitic acid | Perkin Elmer | Cat#NEC075H |
| [2-14C]-pyruvic acid, sodium salt | Perkin Elmer | Cat#NEC256 |
| [3H]-2-deoxy-D-glucose | Perkin Elmer | Cat#NET328A |
| Critical commercial assays | ||
| EnzyChrom NAD+/NADH assay kit | BioAssay System | Cat#E2ND-100 |
| AST(GOT1) Activity Assay Kit | Sigma-Aldrich | Cat#MAK055-1KT |
| ADP/ATP Ratio Assay kit | Sigma-Aldrich | Cat#MAK135-1KT |
| Pyruvate Dehydrogenase Activity Assay Kit | Sigma-Aldrich | Cat#MAK183 |
| Experimental models: Cell lines | ||
| Pgc-1α−/− brown adipocytes | Uldry, M. et al.40 | N/A |
| FL-Pgc-1α−/− brown adipocytes | Jun, H.J. et al.41 | N/A |
| NT-Pgc-1α−/− brown adipocytes | Kim, J. et al.42 | N/A |
| Got1KO brown adipocytes | This paper | N/A |
| Got1OE brown adipocytes | This paper | N/A |
| Experimental models: Organisms/strains | ||
| C57BL/6 mice | Jackson Laboratory | RRID:IMSR_JAX:000664 |
| Ucp1-cre transgenic mice | Jackson Laboratory | RRID:IMSR_JAX:024670 |
| C57BL/6N-Got1tm1a(EUCOMM)Hmgu/H mice | EMMA | RRID:IMSR_EM:10399 |
| R26-Got1 knock-in mice | This paper | N/A |
| Oligonucleotides | ||
| Primers for qPCR from PrimerBank | https://pga.mgh.harvard.edu/primerbank/ | N/A |
| Recombinant DNA | ||
| CAG-loxP-neo-3xpA(stop)-loxP-GOT1 | This paper or Lab | N/A |
| pBABE-Puro-SV40LT | Addgene | RRID:Addgene_13970 |
| WZL-neo-cre | AddGene | RRID:Addgene_34568 |
| pcDNA3.1-Peredox-mCherry | AddGene | RRID:Addgene_32383 |
| Software and algorithms | ||
| ImageJ software | https://imagej.nih.gov/ij | RRID:SCR_002285 |
| Prism 10 software | GraphPad Software Inc. | RRID: SCR_002798 |
| Graphic abstract and schematics in figures | https://www.biorender.com | RRID:SCR_018361 |
| Integrative Genome Viewer (v2.3) | https://igv.org/ | RRID:SCR_011793 |
| Analysis instrument | ||
| Promethion | Sable Systems International | N/A |
| Next EZ glucometer | Bayer | N/A |
| Lionheart FX Automated Microscope | BioTek | RRID:SCR_019744 |
| OROBOROS Oxygraph-2k | Oroboros Instruments | N/A |
| Seahorse XF24 analyzer | Agilent Technologies | RRID:SCR_019539 |
| Hamamatsu NanoZoomer slide scanner | Hamamatsu | RRID:SCR_023762 |
| Applied Biosystems 7900 | Thermo Fisher Scientific | RRID:SCR_018060 |
| SpectraMax Plus 384 Absorbance Plate Reader | Molecular Devices | RRID:SCR_018598 |
| Tri-Carb 2910TR liquid scintillation analyzer | Perkin Elmer | N/A |