KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
|
| ||
| Antibodies | ||
|
| ||
| RPPA antibodies | MD-Anderson Cancer Center | https://www.mdanderson.org/research/research-resources/core-facilities/functional-proteomics-rppa-core/antibody-information-and-protocols.html |
| Anti-FLAG | Sigma Aldrich | Cat# F3165; RRID: AB_259529 |
| Anti-AR | Abcam | Cat# Ab133273; RRID: AB_11156085 |
| Anti-AR | Millipore Sigma | Cat# 06-680; RRID: AB_310214 |
| Anti-VAMP2 | Synaptic Systems | Cat# 104 211; RRID: AB_887811 |
| Anti-NTPDase3 | Ectonucleotidases-ab | Cat# hN3-B3S and hN3-H10S |
| Anti-STX4 | Custom Designed | N/A |
| Ant-rat-Insulin | DSHB | Cat# GN-ID4; RRID: AB_2255626 |
| Anti-Glucagon | Cell signaling | Cat# 2760; RRID: AB_659831 |
| Anti-mouse Alexa Fluor 647 | Thermofischer Scientific | Cat# A-31571; RRID: AB_162542 |
| Anti-rat Alexa Fluor 555 | Cell signaling | Cat# 4417; RRID: AB_10696896 |
| Anti-rabbit Alexa Fluor 488 | Cell signaling | Cat# 4412; RRID: AB_1904025 |
|
| ||
| Biological samples | ||
|
| ||
| De-identified human pancreatic islets | PRODO Laboratories Inc | https://prodolabs.com/ |
| De-identified human pancreatic islets | Integrated Islet Distribution Program (IIDP) | https://iidp.coh.org/ |
|
| ||
| Chemicals, peptides, and recombinant proteins | ||
|
| ||
| 17β-Estradiol (E2) | Steraloids | Cat# E0950-000 |
| GLP-1 | Richard D. DiMarchi | N/A |
| Exendin 9–39 | Sigma Aldrich | Cat# E7269 |
| Forskolin | Sigma Aldrich | Cat# F3917 |
| GIP | Tocris | Cat# 2084 |
| Glucagon | Sigma Aldrich | Cat# G2044 |
| DHT | Steraloids | Cat# A2570-000 |
| H89 | CST | Cat# 9844 |
| ESI-09 | Sigma Aldrich | Cat# SML0814 |
| LRE1 | Sigma Aldrich | Cat# SML1857 |
| KH7 | Sigma Aldrich | Cat# K3394 |
| TDI-10229 | Cornell | N/A |
| Acetazolamide (AZZ) | MedChemExpress | Cat# HY-B0782 |
| CFTRinh-172 (CFTRi) | Selleckchem | Cat# S7139 |
| S0859 | MedChemExpress | Cat# HY-15529 |
| 2′,5′-Dideoxyadenosine | Sigma Aldrich | Cat# D7408 |
| PF431396 | Selleckchem | Cat# S7644 |
| PF562271 | Selleckchem | Cat# S2890 |
| PF573228 | MedChemExpress | Cat# HY-10461 |
| PF4618433 | MedChemExpress | Cat# HY-18312 |
| PP2 | Sigma Aldrich | Cat# P0042 |
| BKM120 | Selleckchem | Cat# S2247 |
| GDC-0941 | Sigma Aldrich | Cat# 5.09226 |
| Rapamycin | Sigma Aldrich | Cat# 37094 |
| JR-AB2-011 | TargetMol | Cat# T11728 |
| Linagliptin | Boehringer Ingelheim | N/A |
| Finasteride | Sigma Aldrich | Cat# F1293 |
| Dutasteride | Sigma Aldrich | Cat# SML1221 |
| SNAP-Surface 549 | NEB | Cat# S9112S |
| Exendin4 | Sigma Aldrich | Cat# E7144 |
| Exendin-Phe1 | Imperial College London | N/A |
| Streptozotocin | Sigma Aldrich | Cat# 18883-66-4 |
| D-(+)-Glucose | Sigma Aldrich | Cat# G7528 |
| PBS | Gibco | Cat# 10010023 |
| HBSS | Gibco | Cat# 14025092 |
| Phenol red free RPMI-1640 medium | Gibco | Cat# 11835030 |
| DMEN medium | Gibco | Cat# 11965126 |
| Phenol red free DMEN medium | Gibco | Cat# 21063029 |
| HEPES | Gibco | Cat# 15630080 |
| Sodium Pyruvate | Gibco | Cat# 11360070 |
| GlutaMAX | Gibco | Cat# 35050061 |
| Penicillin/streptomycin | Gibco | Cat# 15140063 |
| Collagenase | Sigma Aldrich | Cat# C9263 |
| Charcoal Stripped FBS | GemCell™ | Cat# 100–119 |
| FBS | GemCell™ | Cat# 100–500 |
| Glutaraldehyde | Ted Pella Inc. | Cat# 18436 |
| Sodium cacodylate buffer | Ted Pella Inc. | Cat# 18851 |
| Osmium tetroxide | Polysciences Inc. | Cat# 20816-12-0 |
| Uranyl acetate | Ted Pella Inc. | Cat# 19481 |
| Eponate 12 resin | Ted Pella Inc. | Cat# 18005 |
| Lead citrate | Sigma Aldrich | Cat# 15326 |
| Ammonium acetate | Sigma Aldrich | Cat# 73594 |
| Tributylamine | Sigma Aldrich | Cat# 8.08358 |
| Acetic acid | Sigma Aldrich | Cat# 27225 |
| Acetonitrile | Sigma Aldrich | Cat# 34851 |
| NanoLuc substrate | Promega | Cat# N1110 |
| Lipofectamine | Thermofischer Scientific | Cat# L3000001 |
| Tamoxifen | Sigma Aldrich | Cat# T5648 |
| TrypLE | Thermofischer Scientific | Cat# 12604013 |
| Ethanol | Sigma Aldrich | Cat# E7023 |
| Versene | ThermoFisher | Cat# 15040066 |
|
| ||
| Critical commercial assays | ||
|
| ||
| Seahorse XF Cell Mito Stress Test Kit | Agilent | Cat# 103010-100 |
| Human Insulin Elisa | Mercodia Inc. | Cat# 10-1113-01 |
| Rat/Mouse Insulin ELISA kit | Millipore | Cat# EZRMI-13K |
| Testosterone Elisa | IBL America | Cat# IB79106 |
| Luteinizing hormone Elisa | LSBio | Cat# LS-F22503-1 |
| 17β-estradiol Elisa | Calbiotech | Cat# ES380S |
| 10x Single Cell 3′ v3.1 RNAseq kit | 10X Genomics | Cat# PN-1000121 |
| Illumina NextSeq 2000 P3 100 kit | Illumina Biosciences | Cat# 20040559 |
| Illumina NextSeq 2000 P3 100 kit | Illumina Biosciences | Cat# 20040559 |
| CELLection Pan Mouse IgG kit | Thermofischer Scientific | Cat# 11531D |
|
| ||
| Deposited data | ||
|
| ||
| Single Cell RNAseq Data | This study. Deposited location: Gene Expression Omnibus - NCBI | GEO: GSE201256 |
| Proteomics Data | This study. Deposited location: Proteomics Identification Database - EMBL-EBI | PRIDE: PXD041592 |
| Metabolomics Data | This study. Deposited location: UCSD Metabolomics Workbench/National Metabolomics Data Repository | NMDR: PR001654 |
|
| ||
| Experimental models: Cell lines | ||
|
| ||
| INS1 832/13 | Millipore Sigma | Cat# SCC207; RRID: CVCL_7226 |
| INS1 832/3 | Millipore Sigma | Cat# SCC208; RRID: CVCL_ZL55 |
| MIN6B | University of Geneva | N/A |
| MIN6B1-SNAP-GLP-1R | Imperial College London | N/A |
|
| ||
| Experimental models: Organisms/strains | ||
|
| ||
| Mouse: C57BL/6J | The Jackson Laboratory | Strain#000664 |
| Mouse: MIP-CreERT | The Jackson Laboratory | Strain#:024709 |
| Mouse: ARfl/fl | Catholic University of Leuven | N/A |
| Mouse: Ins2-creTg(Ins2–cre)25Mgn/J | The Jackson Laboratory | Strain# 003573 |
| Mouse: Glp1rf/f | Dr. David D’Alessio | N/A |
| Mouse: Raptorfl/fl | The Jackson Laboratory | Strain# 013188 |
| Mouse: Rictortm1.1Mnh | The Jackson Laboratory | Strain# 020649 |
| Mouse: Ins2-creTg(Ins2–cre)23Herr | MMRC | Strain# 011155-MU |
| Mouse: KD-mTOR | Dr. Druie Cavender | N/A |
|
| ||
| Recombinant DNA | ||
|
| ||
| pTEpacVV | The Netherlands Cancer Institute) | N/A |
| pEndo-TEpacVV | University of Birmingham | N/A |
| pbPAC-Endo | UC San Francisco | N/A |
| pbPAC-PM | UC San Francisco | N/A |
| pPM-TEpacVV | University of Birmingham | N/A |
| pGLP-1R-GFP | Imperial College London | N/A |
| pFLAG-AR | Imperial College London | N/A |
| pEpac2-camps | University of Cambridge | N/A |
| LgBiT-GLP-1R | University of Birmingham | N/A |
| SmBiT-AR | University of Birmingham | N/A |
| SmBiT-Empty | University of Birmingham | N/A |
| SmBiT-GLP1R | University of Birmingham | N/A |
| LgBiT-Gαs | University of Birmingham | N/A |
|
| ||
| Software and algorithms | ||
|
| ||
| Single cell analysis architecture (Github + Zenodo) | FMJ Lab | https://zenodo.org/record/7814544 https://github.com/FMJLabTulane/AR-DHT |
| Gen5 imager software | Agilent | https://www.biotek.com/products/software-robotics-software/gen5-microplate-reader-and-imager-software/ |
| Elements Advanced Research software | Nikon | https://www.microscope.healthcare.nikon.com/products/software/nis-elements/nis-elements-advanced-research |
| Zeiss Zen Software | Zeiss | https://www.zeiss.com/microscopy/en/products/software/zeiss-zen.html |
| Cellranger | 10X Genomics | https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger |
| Seurat | The Satija Lab | https://satijalab.org/seurat/ |
| R | The R Consortium | https://www.r-project.org/ |
| RStudio v1.2.1335 (64x bit, for Windows) | The R Consortium | https://www.rstudio.com/products/ |
| Ingenuity Pathway Analysis (IPA) | Qiagen | https://digitalinsights.qiagen.com/products-overview/discovery-insights-portfolio/analysis-and-visualization/qiagen-ipa/?cmpid=QDI_GA_IPA&gclid=Cj0KCQrKQBhCNARIsACUEW_beTxh0IgfRAtfWcvbawY1cFpVLowb2G6pe8Qf7jWZtJY1s6KfuAGIaAuxEEALw_wcB |
| Proteome Discoverer 2.4 | ThermoFisher | https://www.thermofisher.com/us/en/home/industrial/mass-spectrometry/liquid-chromatography-mass-spectrometry-lc-ms/lc-ms-software/multi-omics-data-analysis/proteome-discoverer-software.html |
| GraphPad Prism v8 | GraphPad | https://www.graphpad.com/ |
| STRING | v11.5 | https://string-db.org/ |
| EnrichR | Developed in the Ma’ayan Lab | https://maayanlab.cloud/Enrichr/ |