Fig 1: Interaction proteomics of the MICOS complex reveals QIL1 as a novel interactor.(A) Immunofluorescence analysis of the subcellular localization of tagged proteins. Bars, 20 µm. (B) Overview of the MICOS interaction network obtained from IP-MS analysis. (C) Validation of QIL1 protein interactions by IP-MS analysis. Figure 1—figure supplement 1 shows a schematic representation of the MICOS complex highlighting the subunits and interactors analyzed by IP-MS in red and the expression levels of C-terminally tagged proteins compared to the endogenous version. Figure 1—figure supplement 2 contains the entire IP-MS data set. (D) Immunofluorescence analysis of the subcellular localization of C-terminally tagged QIL1. Bars, 20 µm. Endogenous QIL1 (E), MIC60 (F), or MIC10 (G) was immunopurified from crude mitochondria isolated from 293T cells. Endogenous interactions with MIC60, MIC10, AFG3L2, TMEM11, or QIL1 were assessed. (H) Alignment of QIL1 orthologs using the ClustalW software. ‘H’ indicates amino acids present within a conserved predicted transmembrane region (TMPred algorithm). (I) Cytoplasmic and mitochondrial fractions were separated in 293T lysates. Soluble and membrane fractions were separated by alkaline extraction. S indicates soluble, P indicates pellet. (J) Alkaline extraction was performed at increasing pH (pH11, pH11.5, pH12). (K) Proteinase K (PK) and increasing concentrations of digitonin were used to assess QIL1 sub-mitochondrial localization. (L) Proteinase K (PK), osmotic shock (OS), and Triton X-100 (TX100) were used to assess QIL1 sub-mitochondrial localization.DOI: http://dx.doi.org/10.7554/eLife.06265.003