APEX2 mediated proximity labeling to investigate the proximal proteome of mitochondria tethered ULK1 in wild type and fip200 knock down cells

The object of this project is to investigate the proximal proteome of ULK1 during mitophagy and the effects of a FIP200 knock down using the APEX2-mediated proximity labeling system. APEX2-ULK1 or APEX2 was fused to 2xFKBP-myc and stably expressed in a cell line stably expressing a truncated FIS1-FRB fusion protein. Treatment with rapalog induces the stable interaction between FRB and FKBP, resulting in tethering of FKBPx2-myc-APEX2-ULK1 or FKBPx2-myc-APEX2 to the outer mitochondrial membrane.