| Antibodies |
| Rabbit anti-ATG13 |
Cell Signaling Technology |
#13468 |
| Rabbit anti-TBK1 |
Cell Signaling Technology |
#3504 |
| Rabbit anti-ULK1 |
Cell Signaling Technology |
#6439 |
| Rabbit anti- TBK1PS172
|
Cell Signaling Technology |
#5483 |
| Rabbit anti-GABARAP |
Cell Signaling Technology |
#13733 |
| Rabbit anti-GABARAPL2 |
Cell Signaling Technology |
#14256 |
| Rabbit anti-LC3B |
Cell Signaling Technology |
#3868 |
| Rabbit anti-Stx17pS202
|
This work |
Rabbit No: BL31269 and prep PP1 |
| Rabbit anti- Atg1/ULK1 |
Sigma Aldrich |
A7481 |
| Rabbit anti-Stx17 |
Sigma Aldrich |
HPA001204 |
| Rabbit anti-GFP |
Abcam |
ab290 |
| Rabbit anti-GFP |
Abcam |
ab38689 |
| Rabbit anti-LC3 |
MBL International |
PM036 |
| Rabbit anti-FIP200 |
Proteintech |
17250–1-AP |
| Mouse anti-GM130 |
BD Biosciences |
RUO-610822 |
| Mouse anti-ATG13 |
Millipore |
MABC46 |
| Mouse anti-FLAG |
Sigma Aldrich |
F1804 |
| Mouse anti-ERGIC53 |
Santa Cruz Biotechnology |
sc-365158 |
| Mouse anti- β actin |
Santa Cruz Biotechnology |
sc-47778 |
| Mouse anti-Myc |
Santa Cruz Biotechnology |
sc-40 |
| Goat anti mouse IRDye 680 |
LI-COR |
925–68020 |
| Goat a Rabbit IRDye 800 |
LI-COR |
926–32211 |
| Trueblot anti-mouse DyLight 680 |
Rockland |
18-4516-32 |
| Trueblot anti-rabbit DyLight 800 |
Rockland |
18-8816-31 |
| Alexa Fluor 488 secondary antibody |
ThermoFisher |
A-11029 |
| Alexa Fluor 568 secondary antibody |
ThermoFisher |
A-11036 |
| Bacterial and Virus Strains |
| NEB 5-alpha Competent E.coli (High Efficiency) |
New England Biolabs |
C2987 |
| One Shot Mach1 Phage-Resistant Competent E.coli
|
ThermoFisher |
C862003 |
| SoluBL21 Competent E.coli
|
Genlantis |
C700200 |
| LentiCRISPRv2 |
GeneScript |
N/A |
| Chemicals, Peptides, and Recombinant Proteins |
| LR Clonase II Plus Enzyme Mix |
ThermoFisher |
11791100 |
| BP Clonase II Plus Enzyme Mix |
ThermoFisher |
11789100 |
| Lambda Protein Phosphatase |
NEW ENGLAND BioLabs Inc. |
P0753S |
| Bafilomycin A1 |
InvivoGen |
tlrl-baf1 |
| 3-Methyladenine |
Sigma |
M9281 |
| mouse macrophage colony stimulating factor (mM-CSF) |
Cell Signaling Technology |
#5228 |
| OptiPrep Density Gradient Medium |
Sigma |
D1556 |
| Critical Commercial Assays |
| Gateway Vector Conversion System |
ThermoFisher |
11828–029 |
| ProFection Mammalian Transfection System |
Promega |
E1200 |
| Amaxa Cell Line Nucleofector Kit R |
Lonza |
VCA-1001 |
| Lipofectamine 2000 |
ThermoFisher |
11668019 |
| LDH Assay kit (CytoTox 96® Non-Radioactive Cytotoxicity Assay) |
Promega |
G1780 |
| GFP-TRAP |
Chromotek |
Gtm-20 |
| Deposited Data |
| Raw MS data |
https://massive.ucsd.edu |
MSV000083251 |
| Experimental Models: Cell Lines |
| HeLa |
ATCC |
CCL-2 |
| HEK-293T |
ATCC |
CRL-11268 |
| Bone marrow macrophages |
From C57BL/6 Mouse femur |
N/A |
| HeLa Stx17KO
|
This work |
N/A |
| HeLa TBK1KO
|
This work |
N/A |
| HeLa ATG13KO
|
This work |
N/A |
| HeLa WT |
(Nguyen et al., 2016) |
N/A |
| HeLa LC3TKO
|
(Nguyen et al., 2016) |
N/A |
| HeLa GABARAPTKO
|
(Nguyen et al., 2016) |
N/A |
| HeLaHexaKO |
(Nguyen et al., 2016) |
N/A |
| 293T TBK1KO
|
This work |
N/A |
| Experimental Models: Organisms/Strains |
| C57BL/6J mice |
Jackson Laboratories |
C57BL/6J |
| Oligonucleotides |
| Primers for Stx17 S202A sense: 5′GTTGACATGGTCTGCAATGGCGTCAATCTTCT CCTGCTGA-3′ |
Integrated DNA Technologies |
N/A |
| Primers for Stx17 S202A anti-sense: 5′TCAGCAGGAGAAGATTGACGCCATTGCAGAC CATGTCAAC-3′ |
Integrated DNA Technologies |
N/A |
| Primers for Stx17 S202D sense: 5′GTTGACATGGTCTGCAATGTCGTCAATCTTCT CCTGCTGA-3′ |
Integrated DNA Technologies |
N/A |
| Primers for Stx17 S202D anti-sense: 5′TCAGCAGGAGAAGATTGACGACATTGCAGAC CATGTCAAC-3′ |
Integrated DNA Technologies |
N/A |
| ATG13 CRISPR sgRNA: TCACCCTAGTTATAGCAAGA |
Custom order (Harvard GEPG) |
N/A |
| Stx17 CRISPR sgRNA: GATGAAGATCAACCTGGAAG |
Custom order (Harvard GEPG) |
N/A |
| TBK1 CRISPR sgRNA: GATAGTAATCCCAACAGACC |
Custom order (Invitrogen) |
N/A |
| Recombinant DNA |
| pDest-GFP-Stx17 |
(Kumar et al., 2018) |
N/A |
| pDest-GFP-Stx17S202A
|
This work |
N/A |
| pDest-GFP-Stx17S202D
|
This work |
N/A |
| pDest-FLAG-Stx17 |
(Kumar et al., 2018) |
N/A |
| pDest-FLAG-Stx17S202A
|
This work |
N/A |
| pDest-FLAG-Stx17S202D
|
This work |
N/A |
| pDest-Myc-TBK1 |
(Pilli et al., 2012) |
N/A |
| pDest-Myc-TBK1K38D
|
(Pilli et al., 2012) |
N/A |
| pDest-mCherry-EGFP-LC3B |
(Pankiv et al., 2007) |
N/A |
| EGFP-WIPI2 |
Tassula Proikas-Cezanne (Eberhard Karls University Tübingen, Tübingen, Germany) |
N/A |
| GFP-ULK1 |
N. Mizushima (The University of Tokyo, Tokyo, Japan) |
N/A |
| pMXs-IP-EGFP-hFIP200 |
Addgene |
#38192 |
| pEGFP-DFCP1 |
(Axe et al., 2008) |
N/A |
| pDest-EGFP-LC3B |
(Kirkin et al., 2009) |
N/A |
| Keima-LC3 |
(An and Harper, 2018) |
N/A |
| LDH-Keima |
(An and Harper, 2018) |
N/A |
| Software and Algorithms |
| iDEV software |
ThermoFisher |
N/A |
| AIM software |
Carl Zeiss |
N/A |
| Scaffold software |
Proteome Software Inc |
N/A |
| Image Studio Software |
LI-COR |
N/A |
| Prism 6 |
GraphPad |
N/A |
| ImageJ |
National Institute of Health, USA |
N/A |
| Other |
| RIPA Lysis Buffer |
ThermoFisher |
89900 |
| NP40 Cell Lysis Buffer |
ThermoFisher |
FNN0021 |
| Protease Inhibitor Cocktail Tablets |
Roche |
11697498001 |
| PMSF |
Sigma Aldrich |
93482 |
| Dynabeads Protein G |
ThermoFisher |
10003D |
