Table 1.
Key Resources.
| Primary antibodies | |||
|---|---|---|---|
| Antigen | Species | Supplier | Identifier |
| ACACA/ACC | Rabbit | Cell Signaling Technology | 3662 |
| ACACA/ACC (p-S79) | Rabbit | Cell Signaling Technology | 3661 |
| PRKAA/AMPKα (p-T172) | Rabbit | Cell Signaling Technology | 2535 (D79.5E) |
| PRKAA1/AMPKα1 | Rabbit | Cell Signaling Technology | 5831A |
| PRKAA1/AMPKα1 and PRKAA2/AMPKα2 | Mouse | Abcam | ab80039 (34.2) |
| PRKAG1/AMPKγ1 | Rabbit | Abcam | ab32382 |
| PRKAG1/AMPKγ1 | Rabbit | Abcam | ab32508 |
| PRKAG1/AMPKγ1 (p-S192) | Rabbit | This paper | |
| PRKAG1/AMPKγ1 (p-T284) | Rabbit | This paper | |
| AXIN1 | Rabbit | Cell Signaling Technology | 2074 (C95H11) |
| ATG13 | Rabbit | Cell Signaling Technology | 13468 (E1Y9V) |
| ATG13 (p-Ser318) | Rabbit | Rockland | 600-401-C49 |
| ATP6V0D1 | Rabbit | Abcam | ab202899 |
| CTSB | Mouse | Sigma Aldrich | C6243 (CB59-4B) |
| CDK7 | Mouse | J. Bartek, Danish Cancer Society, Copenhagen, DK | Ref. [50] |
| PRKDC/DNA-PKcs | Rabbit | Abcam | ab230 |
| PRKDC/DNA-PKcs | Rabbit | Abcam | ab70250 |
| PRKDC/DNA-PKcs | Mouse | Abcam | ab1832 (18–2) |
| PRKDC/DNA-PKcs (p-S2056) | Rabbit | Abcam | ab24918 |
| PRKDC/DNA-PKcs (p-T2609) | Rabbit | Abcam | ab18356 (10B1) |
| FLAG® (M2 Affinity Gel) | Mouse | Sigma Aldrich | A-2220 |
| FLAG® | Rabbit | Sigma Aldrich | F7425 |
| GAPDH | Mouse | Abcam | ab9484 |
| H2AFX (p-Ser139) | Rabbit | Millipore | JBW301 |
| HA | Mouse | Sigma Aldrich | H3663 |
| HA (agarose) | Mouse | Sigma Aldrich | A2095 |
| LAMP2 antibody | Mouse | Dev. Studies Hybridoma Bank | Clone H4B4 |
| XRCC5/KU80 | Rabbit | Abcam | ab87860 |
| STK11/LKB1 | Rabbit | Cell Signaling Technology | Ca3050S (27D10) |
| STK11/LKB1 | Rabbit | Abcam | ab185734 |
| MCM7 | Mouse | J. Bartek, Danish Cancer Society, Copenhagen, DK | Ref. [51] |
| RPS6KB1 | Rabbit | Cell Signaling Technology | 2708 (49D7) |
| RPS6KB1 (p-Thr389) | Rabbit | Cell Signaling Technology | 9234 (108D2) |
| SESN2 | Rabbit | Cell Signaling Technology | 8487 (D1B6) |
| SQSTM1 | Rabbit | Enzo Life Sciences | BLM-PW9860 |
| TP53 | Rabbit | J. Bartek, Danish Cancer Society, Copenhagen, DK | Ref. [52] |
| TP53 (p-S15) | Rabbit | Cell Signaling Technology | 9284 |
| ULK1 | Rabbit | Cell Signaling Technology | 4765S |
| ULK1 (p-S555) | Rabbit | Cell Signaling Technology | mAb #586 (D1H4) |
| ULK1 (p-S757) | Rabbit | Cell Signaling Technology | 14202 (D7O6U |
| ULK1 (p-S317) | Rabbit | Cell Signaling Technology | 12753 (D2B6Y) |
| ULK1 | Rabbit | Santa Cruz Biotechnology | SC-33182 (H-240) |
| ULK1 | Mouse | Santa Cruz Biotechnology | SC-390904 (F-4) |
| WIPI2 | Mouse | Abcam | ab105459 (2A2) |
| Plasmids | |||
| Name | Supplier | Identifier | |
| pFLAG-Cherry-N1-PRKAG1 | Jay Brenman, University of North Carolina, Chapel Hill, NC |
Addgene ID: 30308 (Ref. [53]) | |
| pRluc-LC3wt | Marja Jäättelä, Danish Cancer Society, Copenhagen, Denmark |
Addgene ID: 105002 (Ref. [19]) | |
| pRluc-LC3G120A | Marja Jäättelä, Danish Cancer Society, Copenhagen, DK |
Addgene ID: 105003 (Ref. [19]) | |
| pDEST27-GST-PRKAA2 | Jon Oakhill, University of Melbourne, Victoria, Australia |
Ref. [60] | |
| pcDNA3.1-PRKAB1-FLAG | Jon Oakhill, University of Melbourne, Victoria, Australia |
Ref. [60] | |
| pMT2-HA-PRKAG1 | Jon Oakhill, University of Melbourne, Victoria, Australia |
Ref. [60] | |
| pMT2-HA-PRKAG1 (S192A) | Jon Oakhill, University of Melbourne, Victoria, Australia |
This paper | |
| pMT2-HA-PRKAG1 (T284A) | Jon Oakhill, University of Melbourne, Victoria, Australia |
This paper | |
| siRNAs | |||
| Target gene | Sequence | Supplier | |
| Non-targeting control | 5ʹ-CGA CCG AGA CAA GCG CAAG-3’ | Dharmagon | |
| BECN1 | 5ʹ-CAG UUU GGC ACA AUC AAUA-3’ | Sigma Aldrich | |
| PRKDC A | 5ʹ-GCG UUG GAG UGC UAC AACA-3’ | Sigma Aldrich | |
| PRKDC B | 5ʹ-GCG CUU UUC UGG GUG AACU-3’ | Sigma Aldrich | |
| PRKDC C | 5ʹ-CAA GCG ACU UUA UAG CCUU-3’ | Sigma Aldrich | |
| RPTOR | 5ʹ-GAU GAG GCU GAU CUU ACAG-3’ | Sigma Aldrich | |
| ULK1 | 5ʹ-CCC UUU GCG UUA UAU UGUA-3’ | Sigma Aldrich | |
| XRCC5 #1 | 5ʹ-CUC CAU UCC UGG UAU AGAA-3’ | Sigma Aldrich | |
| XRCC5 #2 | 5ʹ-CCG CUG AGG UGA CAA CAAA-3’ | Sigma Aldrich | |
| PRKAG1 mutagenesis oligos (ref. P54619, UniProtKB) | |||
| Construct | Primer name | Sequence | |
| S192A mutant | t574g_antisense | 5ʹ-ctgtagctcttccagagccttggacatgaactctg-3’ | |
| t574g_ | 5ʹ-cagagttcatgtccaaggctctggaagagctacag-3’ | ||
| T284A mutant | a850g_antisense | 5ʹ-atgatggtctccagagcctcatgcaggtagcac-3’ | |
| a850g_ | 5ʹ-gtgctacctgcatgaggctctggagaccatcat-3 | ||
| shRNA resistant | g195c_t198g_c201t_antisense | 5´-ggcagctcgtacaccattcgtgaccaaagcaaaaaaagctttcttca-3´ | |
| g195c_t198g_c201t_ | 5´-tgaagaaagctttttttgctttggtcacgaatggtgtacgagctgcc-3´ | ||