| Antibodies |
| Rabbit polyclonal anti-S6K1 |
Cell Signaling Technology |
Cat#2708; RRID: AB_390722 |
| Rabbit monoclonal anti-pS6 (S240/S244) |
Cell Signaling Technology |
Cat#5364; RRID: AB_10694233 |
| Rabbit Polyclonal anti-SLC4A7 |
Invitrogen (Thermo Fisher) |
Cat#PA5-75807; RRID:AB_2719535 |
| Band 3/ AE1 Polyclonal antibody (SLC4A1) |
ProteinTech Group, Inc |
Cat#28131-1-AP |
| AE2 Polyclonal antibody (SLC4A2) |
ProteinTech Group, Inc |
Cat#26332-1-AP |
| SLC4A3 Polyclonal antibody |
ProteinTech Group, Inc |
Cat#26883-1-AP |
| SLC4A4 Polyclonal antibody |
ProteinTech Group, Inc |
Cat#11885-1-AP |
| Rabbit Polyclonal anti-SLC4A5 Antibody |
Novus Biologicals |
Cat#NBP1-59528 |
| Rabbit Polyclonal anti-SLC4A8 |
ProteinTech Group, Inc |
Cat#12531-1-AP |
| Rabbit AE4 Polyclonal Antibody (SLC4A9) |
Mybiosci |
Cat#MBS821050 |
| SLC4A10 Rabbit pAb |
Abclonal Technology |
Cat#A13810 |
| Rabbit Polyclonal anti-SLC4A11 |
BiorByt |
Cat#orb215316 |
| Phospho-p70 S6 Kinase (Thr389) (108D2) Rabbit mAb |
Cell Signaling Technology |
Cat# 9234S; RRID:AB_2269803 |
| Rabbit Anti-p70 S6 Kinase Monoclonal Antibody |
Cell Signaling Technology |
Cat# 2708S; RRID:AB_390722 |
| Phospho-S6 Ribosomal Protein (Ser240/244) Antibody |
Cell Signaling Technology |
Cat# 2215S; RRID:AB_331682 |
| S6 Ribosomal Protein (5G10) Rabbit mAb |
Cell Signaling Technology |
Cat# 2217S; RRID:AB_331355 |
| Phospho-Akt (Ser473) (D9E) Rabbit mAb |
Cell Signaling Technology |
Cat# 11962; RRID:AB_2797780 |
| Akt (pan) (C67E7) Rabbit mAb |
Cell Signaling Technology |
Cat# 4691, RRID:AB_915783 |
| 4E-BP1 Antibody |
Cell Signaling Technology |
Cat# 9452S; RRID:AB_331692 |
| Tuberin/TSC2 (D93F12) XP_ Rabbit mAb |
Cell Signaling Technology |
Cat# 4308S; RRID:AB_10547134 |
| Anti-mouse IgG, HRP-linked Antibody |
Cell Signaling Technology |
Cat#7076; RRID:AB_330924 |
| Anti-rabbit IgG, HRP-linked Antibody |
Cell Signaling Technology |
Cat#7074; RRID:AB_2099233 |
| β-actin Antibody |
Sigma-Aldrich |
Cat#A5316; RRID: AB_476743 |
| Phospho-CAD (Ser1859) Antibody |
Cell Signaling Technology |
Cat#12662; RRID:AB_2750934 |
| CAD Antibody |
Cell Signaling Technology |
Cat#11933; RRID:AB_2797772 |
| S6K2 Polyclonal antibody |
ProteinTech Group, Inc |
Cat#15268-1-AP; RRID:AB_2182250 |
| Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (E10) Mouse mAb antibody |
Cell Signaling Technology |
Cat# 9106S; RRID: AB_331768 |
| p44/42 MAPK (Erk1/2) Antibody |
Cell Signaling Technology |
Cat# 9102S; RRID: AB_330744 |
| eIF4B Antibody |
Cell Signaling Technology |
Cat# 3592; AB_2293388 |
| eIF4A (C32B4) Rabbit mAb |
Cell Signaling Technology |
Cat#2013; RRID:AB_2097363 |
| DYKDDDDK Tag (D6W5B) Rabbit mAb |
Cell Signaling Technology |
Cat#14793; RRID:AB_2572291 |
| HA-Tag (C29F4) Rabbit mAb |
Cell Signaling Technology |
Cat#3724; RRID:AB_10693385 |
| Phospho-eIF4B (S422) Antibody |
Cell Signaling Technology |
Cat#3591; RRID:AB_2097522 |
| Phospho-eIF4B (Ser406) Antibody |
Cell Signaling Technology |
Cat#5399; RRID:AB_10695246 |
| eIF4A Antibody |
Cell Signaling Technology |
Cat#2490; AB_2097363 |
| H2AX Antibody |
Thermo Fisher (product of: Bethyl) |
Cat#A300-083A-M |
| Bacterial and virus strains |
| NEB Stable Competent E. coli |
NEB |
Cat#C3040 |
| DH5 alpha |
Fisher Scientific |
Cat#18258012 |
| Chemicals, peptides, and recombinant proteins |
| DMEM without Bicarbonate |
US Biological |
Cat#D9812-07 |
| DMEM without glucose, bicarbonate |
US Biological |
Cat#D9800-02B |
| DMEM without glycine (and serine) |
US Biological |
Cat#D9800-16 |
| Rapamycin |
Calbiochem/Sigma-Aldrich |
Cat#553210 |
| Cycloheximide |
EMD Millipore/Merck |
Cat#239764 |
| Glycine |
Sigma-Aldrich |
Cat#G5417 |
| PF-9366 |
MedChemExpress |
Cat#HY-107778 |
| Insulin human |
Alpha diagnostic |
Cat#INSL16-N-5 |
| EGF (human recombinant) |
Life Technologies/GIBCO |
Cat#PHG0311 |
| Insulin-like Growth Factor-I human |
Sigma-Aldrich |
Cat#I3769 |
| Hypoxanthine |
Sigma-Aldrich |
Cat#H9636 |
| Inosine |
Sigma-Aldrich |
Cat#I4125 |
|
13C2-15N-glycine |
Sigma-Aldrich |
Cat#489522 |
| L-glutamine-(amide-15N) |
Sigma-Aldrich |
Cat#490024 |
| 1,2-3H-2-deoxy-D-Glucose |
Perkin-Elmer |
Cat#NET328A250UC |
| U-14C-glycine |
Perkin-Elmer |
Cat#NEC276E050UC |
| U-14C-glucose |
Perkin-Elmer |
Cat#NEC042V250UC |
|
3H-hypoxanthine |
Perkin-Elmer |
Cat#NET177001MC |
|
14C-glutamine |
Perkin Elmer |
Cat#NEC451050UC |
|
14C-formate |
American Radiolabeled Chemicals |
Cat#ARC 0163A |
| 3-14C-serine |
Perkin-Elmer |
Cat#NEC853050UC |
|
3H-uridine |
Perkin Elmer |
Cat#NET367250UC |
| 4-13C aspartate |
Sigma-Aldrich |
Cat#489999 |
|
14C-aspartate |
Perkin Elmer |
Cat#NEC268E050UC |
| Sodium, 14C- bicarbonate |
American Radiolabeled Chemicals |
Cat#ARC 0138C-250 μCi |
| DMSO |
Sigma-Aldrich |
Cat#D2650-100ML |
| L-glutamine |
Corning |
Cat#MT25005Cl |
| 2-mercaptoethanol |
Sigma-Aldrich |
Cat#M6250 |
| Thiazolyl Blue Tetrazolium Bromide |
Sigma-Aldrich |
Cat#M5655 |
| Polyethylenimine (PEI) |
Polysciences |
Cat#23966-2 |
| Opti-MEM media |
Thermo Fisher |
Cat#31985062 |
| Lipofectamine 3000 Transfection reagent |
Invitrogen |
Cat#L3000008 |
| Lipofectamine RNAi max |
Invitrogen |
Cat#13778150 |
| SuperSignal West Femto Chemiluminescent |
Thermo Fisher scientific |
Cat#PI34096 |
| SuperSignal West Pico PLUS |
Thermo Fisher scientific |
Cat#PI34580 |
| Fetal bovine serum (FBS) |
Sigma-Aldrich |
Cat#F2442 |
| Fetal Bovine Serum dialyzed |
Sigma-Aldrich |
Cat#F0392 |
| Microcystin-LR |
Fisher Scientific |
Cat#ALX350012C500 |
| Protease inhibitor cocktail |
Sigma-Aldrich |
Cat#P8340 |
| Bio-Rad Braford |
Bio-Rad |
Cat#5000006 |
| Acrylamide Protogel 30% |
Fisher Scientific |
Cat#EC-890 |
| UltraPure LMP Agarose |
Invitrogen |
Cat#16520-050 |
| Uridine |
Sigma Aldrich |
Cat#U3003 |
| Glutamine-free DMEM |
GIBCO |
Cat#11960-044 |
| Glucose-free DMEM |
Invitrogen |
Cat#11966025 |
| Propidium iodide |
Sigma |
Cat#P4170 |
| Critical commercial assays |
| Fluorometric Intracellular pH Assay Kit |
Sigma-Aldrich |
Cat# MAK150 |
| Intracellular pH Calibration Buffer Kit |
Invitrogen |
Cat# P35379
|
| SYBR Green PCR Master Mix |
Life Technologies |
Cat# 4312704 |
| RNeasy Mini Kit |
QIAGEN |
Cat#74104 |
| HiSpeed Plasmid Maxi Kit (25) |
QIAGEN |
Cat#12663 |
| AllPrep DNA/RNA Mini Kit |
QIAGEN |
Cat#80204 |
| KOD Extreme Hot Start DNA polymerase kit |
Merck Millipore |
Cat#71975-3 |
| GeneJET Plasmid Miniprep Kit |
Thermo Fisher |
Cat# K0503 |
| Deposited data |
| Ali et al. unprocessed files |
Mendeley Data |
http://dx.doi.org/10.17632/9zd4vncrf4.1
|
| Experimental models: Cell lines |
| HEK293E |
John Blenis lab (Weill Cornell Medicine) |
N/A |
| ΔTSC2 HEK293E |
Villa et al. 2021
|
N/A |
| HeLa |
ATCC |
Cat#CCL-2 |
| ΔTSC2 HeLa |
Villa et al. 2021 |
N/A |
| A549 |
ATCC |
Cat#CCL-185 |
| CAL51 |
DSMZ |
Cat#ACC 302 |
| AML 621-102 (TSC2-null-Empty Vector) |
Dr. David Kwiatkowski (Brigham and Women’s Hospital) |
N/A |
| Experimental models: Organisms/strains |
| Athymic nude Foxn1nu |
The Jackson laboratory |
Cat#002019 |
| Oligonucleotides |
| See Table S4 for list of primers used in this study |
Integrated DNA Technologies |
N/A |
| siRNA targeting human SLC4A7 |
Dharmacon |
Cat#L-007586-01-0005 |
| siRNA targeting human S6K1 |
Dharmacon |
Cat#L-003616-00-0005 |
| siRNA targeting human S6K2 |
Dharmacon |
Cat#L-004671 -00-0005 |
| siRNA targeting human eIF4A |
Dharmacon |
Cat#L-020178-00-0005 |
| siRNA targeting human eIF4B |
Dharmacon |
Cat#L-020179-00-0005 |
| Non-targeting siRNA pool |
Dharmacon |
Cat#D-001810-10-20 |
| Recombinant DNA |
| SLC4A7 (NM_003615) Human Tagged ORF Clone |
OriGene |
Cat#RC220734 |
| EIF4B (NM_001417) Human Tagged ORF Clone |
OriGene |
Cat#RC209955 |
| pcDNA3 Flag HA |
Addgene |
Cat#10792 |
| pCMV6-Entry Tagged Cloning Vector |
OriGene |
Cat#PS100001 |
| PX458 CRISPR |
Addgene |
Cat#48138) |
| pRK7-HA-S6K1-WT |
Addgene |
Cat#8984 |
| pRK7/HA-S6K1-F5A-E389-R3A |
Addgene |
Cat#8991 |
| pCMV6/Flag-EIF4B-S422A |
This study |
N/A |
| pCMV6/Flag-EIF4B-S406A |
This study |
N/A |
| Software and algorithms |
| Image J |
National Institutes of Health |
RRID: SCR_003070 |
| Prism 9 |
GraphPad Software |
https://www.graphpad.com/
|
| FlowJo Software |
BD Biosciences |
https://www.flowjo.com/
|
| Other |
| Microplate Reader |
Tecan |
Infinite M1000 Pro |
| Multipurpose Scintillation Counter |
Beckman Coulter |
Model: LS 6500 |
| Nitrocellulose membrane 0.2 μm |
GE Healthcare |
Cat# 10600001 |
| Polypropylene centrifuge tubes |
Beckman Coulter |
Cat# 331372 |
| PCR plates |
Bio-Rad |
Cat#HSP9601 |
| UV transparent flat bottom 96 well plate |
Corning |
Cat#3635 |
