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. Author manuscript; available in PMC: 2023 Nov 17.
Published in final edited form as: Mol Cell. 2022 Oct 24;82(22):4277–4289.e10. doi: 10.1016/j.molcel.2022.09.035

Figure 1. Mitochondrial TA proteins mislocalize to the ER via the EMC.

Figure 1.

(A) EMC depletion rescues the localization of FLAG-tagged OMP25 (F-OMP25), a mitochondrial TA protein, in ATP13A1 knockout (KO) cells. Immunofluorescence of F-OMP25 (green) and the ER marker TRAPα (magenta) in wildtype (WT) and ATP13A1 KO Flp-In HeLa T-REx cells treated with control siRNAs or siRNAs targeting EMC2, EMC6 or GET1. Scale bar, 10 μm.

(B) Pearson’s correlation coefficients (PCC; mean ± sd and individual points for indicated sample size) measuring colocalization of F-OMP25 and TRAPα. ****, p<0.0001; ns, not significant.

(C) Knocking out the EMC reduces F-OMP25 insertion into ER-derived rough microsomes (RMs). SDS-PAGE and autoradiography of total (tot.), membrane-inserted (pel.), and soluble (sup.) radiolabeled F-OMP25 from insertion reactions with RMs obtained from WT, ATP13A1 KO, or ATP13A1 and EMC6 double knockout (DKO) Flp-In 293 T-REx cells (left). Ratios of pelleted to soluble (pel.:sup.) F-OMP25 (top right) or a matched reporter containing the TM of the ER TA protein SQS (F-SQS; bottom right) were normalized to values obtained with WT RMs (mean + sem) for 3 replicates. **, p<0.01; *, p<0.05.

See also Figure S1.