REAGENT or RESOURCE	SOURCE	IDENTIFIER
Chemicals, Peptides, and Recombinant Proteins
cOmplete™, EDTA-free Protease Inhibitor Cocktail	Roche Applied Science	Cat # 11873580001
3X (DYKDDDDK) Peptide	APExBio	Cat # A6001
Critical Commercial Assays
Gibson Assembly® Master Mix	New England BioLabs	Cat # E2611S
ANTI-FLAG® M2 Affinity Gel	Sigma Aldrich	Cat # A2220
Octyl-beta-Glucoside Detergent	Thermo Fisher	Cat # 28310
18:1 (Δ9-Cis) PC (DOPC)	Avanti Polar Lipids	Cat # 850375
16:0–18:1 PE	Avanti Polar Lipids	Cat # 850757
PI3P diC16	Echelon Biosciences	Cat # P-3016
PI(3,5)P2 diC16	Echelon Biosciences	Cat # P-3516
PI3P diC8	Echelon Biosciences	Cat # P-3008
PI(3,5)P2 diC8	Echelon Biosciences	Cat # P-3508
C6-C6-BODIPY-FL-PI3P	Echelon Biosciences	Cat # C-03F6
C6-C6-BODIPY-FL-PI(3,5)P2	Echelon Biosciences	Cat # C-35F6
Malachite Green Phosphate Assay Kit	Cayman Chemical	Cat # 10009325
Deposited Data
PIKfyve complex centered on Vac14 - map1	This paper	EMD: 22634
Vac14 backbone models	This paper	PDB: 7K1Y
PIKfyve complex centered on PIKfyve - map2	This paper	EMD: 22647
PIKfyve Kinase domain, CCT domain models	This paper	PDB: 7K2V
PIKfyve complex centered on Fig4 – map3	This paper	EMD: 22631
Fig4 Sac homology domain model	This paper	PDB: 7K1W
Experimental Models: Cell Lines
Expi293F™ Cells	Gibco™ Thermo Fisher	Cat # A14527
Oligonucleotides
Primers for Recombinant DNA, see Table S2	This paper	N/A
Recombinant DNA
Construct of wildtype Vac14, Fig4, PIKfyve	L. Weisman	N/A
Plasmid: pcDNA3.1 – 2xStrepII-tagged Vac14	This paper	N/A
Plasmid: pcDNA3.1 – 2xStrepII-tagged Vac14 - MBP	This paper	N/A
Plasmid: pcDNA3.1 - His-tagged Fig4 -WT	This paper	N/A
Plasmid: pcDNA3.1 - His-tagged Fig4 -CS	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-Fig4 - WT	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-Fig4 - CS	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-PIKfyve - WT	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-PIKfyve – S2053E	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-PIKfyve – S2053A	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-PIKfyve – K1877E	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-PIKfyve – 722-MBP-723	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-PIKfyve – 1192-MBP-1193	This paper	N/A
Plasmid: pCMV-10 – 3xFLAG-PIKfyve – 1806-MBP-1807	This paper	N/A
Software and Algorithms
ImageJ	Schneider et al., 2012	https://imagej.nih.gov/ij/
RELION 3.0.4	Zivanov et al., 2018	DOI: 10.7554/eLife.42166
RaptorX	Kallberg et al., 2012	http://raptorx.uchicago.edu/
I-TASSER	Yang et al., 2015	https://zhanglab.ccmb.med.umich.edu/I-TASSER/
Other
Quantifoil®, Orthogonal Array of 1.2µm Diameter Holes - 1.3µm Separation, mounted on a 300M Cu grid	TED PELLA, INC	Cat # 658–300-Cu