KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Goat anti-PEPCK-M | Abcam | 40843 |
| PKL pS113 (A/G purified) | Covance-Custom Antibody | N/A |
| PKL pS12 (A/G purified) | Covance-Custom Antibody | N/A |
| PKL | Protein tech | AB_10918271 |
| Phospho-AKT (ser 473 ) | Cell signaling | #9271 |
| Total AKT | Cell signaling | #9272 |
| Phospho-insulin receptor beta | Cell signaling | #3023S |
| Insulin receptor beta | Cell signaling | #3020S |
| Rabbit anti-beta-actin | Cell signaling | 4970 |
| Bacterial and Virus Strains | ||
| Biological Samples | ||
| Human islets | University of Alberta Diabetes Institute | NA |
| Human islets | University of Chicago Diabetes Research and Training Center | NA |
| Human islets | Prodo Laboratories Inc | NA |
| Chemicals, Peptides, and Recombinant Proteins | ||
| DMEM-Base Cell culture media | Sigma Aldrich | D5030 |
| RPMI 1640 Cell culture media | Sigma Aldrich | R8758 |
| Quantitect reverse transcriptase | Qiagen | 205311 |
| SYBR PCR reagent | BioRad | 1708880 |
| CMRL culture media | GIBCO | NA |
| Complete mini EDTA-free protease inhibitor cocktail (Roche) | Sigma Aldrich | 11836170001 |
| PKa2 | Agios pharmaceutical INC | AGI-17540 |
| PKa (TEPP-46) | National Institute of Health | NA |
| GKa2 | MK-0941 | |
| GKa1 | PF-04967319 | |
| Glycogen | Sigma | 10901393001 |
| Glucosidase | Sigma | G5003 |
| High Fat Diet | Dyets | #112245 |
| Critical Commercial Assays | ||
| Micro BCA protein assay kit | Thermo Fisher Scientific | 23235 |
| Rat High Range ELISA | ALPCO | 80-INSRTH-E10 |
| Mouse Ultrasensitive ELISA | ALPCO | 80-INSMSU-E01 |
| RNeasy Mini Kit | Qiagen | 74106 |
| DNeasy Blood & Tissue Kit | Qiagen | 69504 |
| PEPCK Activity Assay | Biovision | K359 |
| HR Series NEFA-HR(2) Color Reagent A | Wako | 999-34691 |
| HR series NEFA-HR(2) solvent A | Wako | 995-34791 |
| HR Series NEFA-HR(2) Color Reagent B | Wako | 991-34891 |
| HR series NEFA-HR(2) solvent B | WAKO | 993-35191 |
| NEFA standard solution | WAKO | 276-76491 |
| TRIGLYCERIDE-SL | SEKISUI | 236-99 |
| GlUCOSE-SL | SEKISUI | 235-60 |
| Human Insulin ELISA | ALPCO | 80-INSHU-E10 |
| Deposited Data | ||
| Experimental Models: Cell Lines | ||
| Clonal INS-1 832/13 cell line over-expressing the human insulin gene (INS-1) | C.B. Newgard (Duke University School of Medicine) | N/A |
| Rat insulinoma cells (INS-1) | NA | NA |
| Experimental Models: Organisms/Strains | ||
| Denoted C57BL/6NTac-Pck2tm1 (KOMP)Vlcg, project ID VG13655 | University of California Davis Knockout Mouse Project repository | NA |
| ZDF (obese) | Charles river | 370 |
| ZDF (lean) | Charles river | 371 |
| Oligonucleotides | ||
| Genotyping primer PCK2 FWD: | acttgctttaaaaaacctcccaca | N/A |
| Genotyping primer PCK2 REV: | gaggcggaaattggttctgcaatgg | N/A |
| Insulin2 FWD: | cagtgccaaggtctgaaggt | N/A |
| Insulin2 REV: | cagcacctttgtggttctca | N/A |
| Insulin1 FWD: | ctgcccaggcttttgtcaa | N/A |
| Insulin1 REV: | tccccacacaccaggtacaga | N/A |
| MafA FWD: | tccaggctggtgcgcgaaag | N/A |
| MafA REV: | gcaagcccactcaggagccg | N/A |
| FOXO1 FWD: | cacagtccaagcgctcaa | N/A |
| FOXO1 REV: | gtgaacaccatgcctcac | N/A |
| mtPEPCK (PCK2) FWD: | ttatgcacgatccctttgccatgc | N/A |
| mtPEPCK (PCK2) REV: | tccttcctttggtacgagcccaat | N/A |
| SCS-ATP FWD: | tgtggtacggttacaaggtaca | N/A |
| SCS-ATP REV: | acaaccattttagcagcttcat | N/A |
| PC FWD: | agatgcacttccatcccaag | N/A |
| PC REV: | ccttggtcacgtgaaccttt | N/A |
| Pdx1 FWD: | gacacatcaaaatctggttccaaa | N/A |
| Pdx1 REV: | tcccgctactacgtttcttatcttc | N/A |
| MLXIPL-CHREBP FWD: | tactgttccctgcctgctc | N/A |
| MLXIPL-CHREBP REV: | cttggaaaccttcaccagg | N/A |
| Lmyc FWD: | gaaggtgcatccgtaccact | N/A |
| Lmyc REV: | tctggggaatagccttgatg | N/A |
| SCS-GTP FWD: | gaggaagaggaaaaggtgtctt | N/A |
| SCS-GTP REV: | tctagaaatatccagggcttca | N/A |
| MafB FWD: | gaagcccgcgaggcatat | N/A |
| MafB REV: | ggccctggcactcacaaa | N/A |
| GCK FWD: | cttggtccaattgaggagga | N/A |
| GCK REV: | cagtggagcgtgaagacaaa | N/A |
| PKM1 FWD: | tggagagcatgatcaagaagc | N/A |
| PKM1 REV: | agggggtctgtggattgact | N/A |
| aChREBP FWD: | tgcatcgatcacaggtcatt | N/A |
| aChREBP REV: | aggctcaagcattcgaagag | N/A |
| PK-M2 FWD: | tggagagcatgatcaagaagc | N/A |
| PK-M2 REV: | cactgcagcacttgaaggag | N/A |
| PK-L FWD: | taagcaacgtagcagcatgg | N/A |
| PK-L REV: | cgatatccagaaggcagagg | N/A |
| NKx 6.1 FWD: | tcttctggcctggggtgatg | N/A |
| NKx 6.1 REV: | ggctgcgtgcttctttctcca | N/A |
| bChREBP FWD: | tctgcagatcgcgcggag | N/A |
| bChREBP REV: | cttgtcccggcatagcaac | N/A |
| TXNIP FWD: | cgagtcaaagccgtcaggat | N/A |
| TXNIP REV | ttcatagcgcaagtagtccaaggt | N/A |
| Recombinant DNA | ||
| Software and Algorithms | ||
| ImageJ software | (NIH; https://imagej.nih.gov/ij/) | N/A |
| El-MAVEN and Polly software | Elucidata | N/A |
| GraphPad Prism 7 software | GraphPad | N/A |
| Other | ||
| Spectromax 384 Plus Spectrophotometer | Molecular Devices | |
| Analox GL5 Analyzer | Anolox InstrumentsLTD, UK | |
| Biorep Perifusion Instrument | Biorep Technologies | |
| Multiquant v2.1 | Sciex | |
| SCIEX 5500 QTRAP equipped with a SelexION for differential mobility separation (DMS) | Sciex | |
| Hypercarb Column (3μm particle size, 4.6x100 mm) | Thermo Fisher Scientific | 35003-104630 |
| Hamamatsu ORCA-Flash4.0 V2 Digital CMOS camera | NA | NA |
| Nikon Elements and custom MATLAB software | NA | NA |