KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| ATP2A2 | Thermo Fisher Scientific | Cat# MA3-919; RRID:AB_325502 |
| Calreticulin | Cell Signaling | Cat# 12238; RRID:AB_2688013 |
| FLAG | Sigma-Aldrich | Cat# F1804; RRID: AB_262044 |
| β-actin | Cell Signaling | Cat# 8457; RRID:AB_10950489 |
| C4ORF3 | This paper | N/A |
| Anti-DYKDDDDK magnetic beads | Sigma-Aldrich | Cat# M8823; RRID:AB_2637089 |
| Streptavidin magnetic beads | Thermo Fisher Scientific | Cat# 65001 |
| Total oxphos cocktail | Abcam | Cat# ab110413; RRID: AB_2629281 |
| TNAP | R&D Systems | Cat# AF2910-SP |
| COX IV | Cell Signaling | Cat# 38563; RRID:AB_2799136 |
| UCP1 | Abcam | Cat# ab10983; RRID:AB_2241462 |
| Goat anti-mouse IgG (H&L) secondary antibody, HRP | Thermo Fisher Scientific | Cat# 31430; RRID: AB_10960845 |
| Goat anti-rabbit IgG (H&L) secondary antibody, HRP | Abcam | Cat# ab6721; RRID: AB_955447 |
| Biological samples | ||
| Human adipose tissue | This paper | N/A |
| Chemicals, peptides, and recombinant proteins | ||
| Indomethacin | Sigma-Aldrich | Cat# I7378 |
| Insulin | Sigma-Aldrich | Cat# I6634 |
| Adenylyl imidodiphosphate | MedChemExpress | Cat# HY-130777 |
| Isobutylmethylxanthine (IBMX) | Sigma-Aldrich | Cat# I5879 |
| Dexamethasone | Sigma-Aldrich | Cat# D4902 |
| 3,3′,5-triiodo-L-thyronine (T3) | Sigma-Aldrich | Cat# T2877 |
| L-(−)-Norepinephrine(+)-bitartrate salt monohydrate | Sigma-Aldrich | Cat# A9512 |
| DMEM | Gibco | Cat# 11965092 |
| Fetal Bovine Serum | ATLANTA biologicals | Cat# S11550 |
| Penicillin-Streptomycin | Gibco | Cat# 15140 |
| Basticidin S HCl (10 mg/mL) | Gibco | Cat# A1113903 |
| 0.05% Trypsin | Corning | Cat# MT25052CI |
| cOmplete, EDTA-free Protease Inhibitor Cocktail | Roche | Cat# 11873580001 |
| Phosphatase inhibitor cocktail 2 | Sigma-Aldrich | Cat# P5726 |
| Phosphatase inhibitor cocktail 3 | Sigma-Aldrich | Cat# P0044 |
| Dextrose | Sigma-Aldrich | Cat# D9434 |
| Palmitoyl-L-carnitine | Sigma-Aldrich | Cat# 61251 |
| Bovine Serum Albumin | Sigma-Aldrich | Cat# A1595 |
| Malate | Sigma-Aldrich | Cat# M1000 |
| Succinate | Sigma-Aldrich | Cat# 224731 |
| Glutamate | Sigma-Aldrich | Cat# G1501 |
| Rotenone | Sigma-Aldrich | Cat# R8875 |
| N,N.N’,N’-Tetramethyl-p-phenylenediamine | Sigma-Aldrich | Cat# T7394 |
| L-ascorbate | Sigma-Aldrich | Cat# A4034 |
| Thapsigargin | Sigma-Aldrich | Cat# T9033 |
| 45CaCl2 | PerkinElmer | Cat# NEZ013001MC |
| CaCl2 | Sigma-Aldrich | Cat# C3306 |
| QIAzol lysis reagent | Qiagen | Cat# 79306 |
| TRIzol reagent | Thermo Fisher Scientific | Cat# 15596026 |
| Adenosine 5’-diphosphate | Sigma-Aldrich | Cat# A5285 |
| MgCl2 | Sigma-Aldrich | Cat# M2393 |
| NaCl | Sigma-Aldrich | Cat# S7653 |
| KCl | Sigma-Aldrich | Cat# P9541 |
| Na2HPO4 | Fluka | Cat# 71639 |
| NaH2PO4 | Sigma-Aldrich | Cat# 71505 |
| Adenosine 5’-triphosphate | Sigma-Aldrich | Cat# A2383 |
| Sucrose | Sigma-Aldrich | Cat# S0389 |
| D-mannitol | Sigma-Aldrich | Cat# M4125 |
| Scintillation fluid | PerkinElmer | Cat# 6013326 |
| Tris | Sigma-Aldrich | Cat# 11814273001 |
| EGTA | Sigma-Aldrich | Cat# E4378 |
| n-Dodecyl β-D-maltoside | Sigma-Aldrich | Cat# D4641 |
| DTT | Sigma-Aldrich | Cat# 43816 |
| Glycerol | Sigma-Aldrich | Cat# G7793 |
| 3X-FLAG peptide | Sigma-Aldrich | Cat# F4799 |
| Paraformaldehyde | Santa Cruz Biotechnology | Cat# SC281692 |
| PBS | Gibco | Cat# 10010023 |
| NaN3 | Sigma-Aldrich | Cat# S2002 |
| MES potassium salt | Santa Cruz | Cat# sc-286156 |
| Chloroform | Sigma-Aldrich | Cat# 650498 |
| Acetonitrile | Thermo Fisher Scientific | Cat# A955 |
| Methanol | Thermo Fisher Scientific | Cat# A456 |
| Rosiglitazone | Enzo Life Sciences | Cat# ALX-350-125 |
| Hygromycin B | Thermo Fisher Scientific | Cat# 10687010 |
| 2,2,2-Tribromoethanol | Sigma-Aldrich | Cat# T48402 |
| KH2PO4 | Sigma-Aldrich | Cat# P5655 |
| GlutaMAX supplement | Thermo Fisher Scientific | Cat# 30055061 |
| Antimycin A | Sigma-Aldrich | Cat# A8674 |
| Water LC/MS | Thermo Fisher Scientific | Cat# W64 |
| Formic acid | Thermo Fisher Scientific | Cat# A117 |
| Guanosine 5’-diphosphate | Sigma-Aldrich | Cat# G7127 |
| SDS | Thermo Fisher Scientific | Cat# BP166 |
| HEPES | Sigma-Aldrich | Cat# H3375 |
| LiCl | Sigma-Aldrich | Cat# L4408 |
| EDTA | Sigma-Aldrich | Cat# E5134 |
| NP-40 | Boston Bioproducts | Cat# P-877 |
| Sodium deoxycholate | Sigma-Aldrich | Cat# D6750 |
| Triton-X 100 | Thermo Fisher Scientific | Cat# BP151 |
| Tween 20 | Thermo Fisher Scientific | Cat# BP337 |
| KOH | Boston Bioproducts | Cat# BZ-8038 |
| Rac-Glycerol 1-Phosphate | Santa Cruz | Cat# sc-215789 |
| Pyruvate | Sigma-Aldrich | Cat# P4562 |
| HBSS | Thermo Fisher Scientific | Cat# 88284 |
| XF calibrant solution | Agilent | Cat# 100840-000 |
| SBI-425 | MedChemExpress | Cat# HY-124756 |
| Creatine | Sigma-Aldrich | Cat# C3630 |
| Laemmli sample buffer | Bio-rad | Cat# 1610747 |
| β-mercaptoethanol | Sigma-Aldrich | Cat# M3148 |
| Potassium oxalate | Sigma-Aldrich | Cat# P0963 |
| MOPS | Sigma-Aldrich | Cat# M3183 |
| Lauryl maltose neopentyl glycol | Thermo Fisher Scientific | Cat# A50940 |
| Biotin | Sigma-Aldrich | Cat# B4639 |
| Urea | Sigma-Aldrich | Cat# 51456 |
| Bismaleimidohexane | Thermo Fisher Scientific | Cat# 22330 |
| Polybrene transfection reagent | Sigma-Aldrich | Cat# TR-1003-G |
| KF | Sigma-Aldrich | Cat# 449148 |
| AlCl3 | Sigma-Aldrich | Cat# 237051 |
| CL 316,243 | Sigma-Aldrich | Cat# C5976 |
| Phenylmethanesulfonyl fluoride | Sigma-Aldrich | Cat# 93482 |
| DNAse | Qiagen | Cat# 79254 |
| ERCC RNA spike-in mix | Thermo Fisher Scientific | Cat# 4456740 |
| Critical commercial assays | ||
| RNeasy lipid tissue mini kit | Qiagen | Cat# 74804 |
| Fluo-8 calcium flux assay kit | Abcam | Cat# ab112129 |
| EnzChek phosphate assay kit | Thermo Fisher Scientific | Cat# E6646 |
| Triglyceride quantification kit | Abcam | Cat# ab65336 |
| iScript Reverse Transcription Supermix for rt-qPCR | Bio-rad | Cat# 1708841 |
| iTaq Universal SYBR Green Supermix | Bio-rad | Cat# 1725125 |
| Biorad gels 4–20% 15 well | Bio-rad | Cat# 4561096 |
| Seahorse XF24 islet capture Fluxpak | Agilent | Cat# 101122-100 |
| Pierce BCA Protein Assay Kit | Thermo Fisher Scientific | Cat# 23225 |
| Rat/mouse insulin ELISA | Sigma-Aldrich | Cat# EZRMI |
| Seahorse XFe24 FluxPak | Agilent | Cat# 102340-100 |
| Glucometer | Abbott | Cat# Freestyle Lite |
| Glucose strips | Abbott | Cat# 70827 |
| Biorad gels 12% 10 well | Bio-rad | Cat# 4568044 |
| RNeasy mini kit | Qiagen | Cat# 74104 |
| FastSelect –rRNA HMR Kits | Qiagen | Cat# 334386 |
| NEBNext Ultra II RNA Library Prep Kit for Illumina | New England Biolabs | Cat# E7770L |
| Deposited data | ||
| RNA sequencing | This paper | GSE268770 |
| Proteomics | This paper | PXD059431 |
| Experimental models: Cell lines | ||
| ALN Mutant_A overexpressing 293T | This paper | N/A |
| ALN Mutant_B overexpressing 293T | This paper | N/A |
| ALN Mutant_C overexpressing 293T | This paper | N/A |
| ALN Mutant_TM overexpressing 293T | This paper | N/A |
| ALN wild type overexpressing 293T | This paper | N/A |
| C4orf3-TurboID immortalized inguinal SVF | This paper | N/A |
| CRISPRi-C4orf3 immortalized inguinal SVF | This paper | N/A |
| W48C ALN mutant-overexpressing inguinal SVF | This paper | N/A |
| Immortalized inguinal SVF | Kajimura Lab43 | N/A |
| SERCA2b-overxpressing inguinal adipocytes | This paper | N/A |
| Experimental models: Organisms/strains | ||
| Mouse: C57BL6J mice | Jackson Laboratory | Cat# 000664 |
| Mouse: CRISPRi-C4orf3 | This paper | N/A |
| Mouse: dCas9/KRAB | Kajimura lab51 | N/A |
| Mouse: UCP1 KO | Jackson Laboratory | Cat# 003124 |
| Mouse: UCP1 × C4orf3 double knockout | This paper | N/A |
| Oligonucleotides | ||
| A full list of qPCR primers in Table S2 | This paper | N/A |
| Recombinant DNA | ||
| AAV8-CAG-EGFP-U6-Scramble | VectorBuilder | N/A |
| AAV8-CAG-EGFP-U6-gRNA-long tracr | VectorBuilder | N/A |
| Software and algorithms | ||
| CaIR | Banks Lab | https://calrapp.org/ |
| Biorender | Biorender | https://biorender.com/ |
| Protter | Protter | https://wlab.ethz.ch/protter/start/ |
| GO Enrichment Analysis | GeneOntology | http://geneontology.org/ |
| GraphPad Prism 8 | GraphPad Software | https://www.graphpad.com/scientific-software/prism/ |
| ImageJ | ImageJ Software | https://imagej.nih.gov/ij/download.html |
| R | Open Source | https://www.r-project.org/ |
| Trimmomatic v.0.36 | Open Source | http://www.usadellab.org/cms/?page=trimmomatic |
| STAR aligner v2.5.2b | Open Source | https://github.com/alexdobin/STAR |
| Subread package v1.5.2 | Open Source | https://subread.sourceforge.net/ |
| edgeR | Open Source | https://bioconductor.org/packages/release/bioc/html/edgeR.html |
| clusterProfiler | Open Source | https://bioconductor.org/packages/release/bioc/html/clusterProfiler.html |
| Pheatmap | Open Source | https://www.rdocumentation.org/packages/pheatmap/versions/1.0.12/topics/pheatmap |
| HDOCK server | Open Source | http://hdock.phys.hust.edu.cn |
| AlphaFold server | Open Source | https://alphafoldserver.com |
| Other | ||
| Standard Diet | Lab Diet | Cat# 5008 |