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Fig. 2. | BMC Biology

Fig. 2.

From: The C-terminal region of the oxidoreductase MIA40 stabilizes its cytosolic precursor during mitochondrial import

Fig. 2.

Lowered levels of C-terminally truncated MIA40 can be partially rescued by proteasomal inhibition. a Steady-state levels of MIA40 truncation variants in HEK293-based YME1L1 deletion cells. MIA40Δ108 and MIA40WT were expressed for 2 days stably, inducibly in YME1L1 knockout in medium containing 30 μg ml−1 cumate. Cells were lysed and analyzed by immunoblotting. MIA40Δ108 is present at decreased levels compared to MIA40WT and is not stabilized by loss of YME1L1. Quantification using Image Lab. Data from 2 experiments were combined. Black arrowhead, endogenous MIA40; blue arrowhead, signal of MIA40Δ108. b Emetine chase analyses of truncated MIA40 variants. Protein expression of cells stably expressing MIA40Δ108 and MIA40WT was induced for 24 h prior to the experiment with 1 μg ml−1 doxycycline. Emetine was added for indicated times, the cells were lysed after 8 h and analyzed by immunoblotting. Mature MIA40Δ108 is equally stable as MIA40WT. Quantification using Image lab. Data from 3 experiments were combined and standard deviations are presented. Black arrowhead, endogenous MIA40; blue arrowhead, signal of MIA40Δ108. c Steady-state levels of MIA40Δ108 and MIA40WT upon proteasomal inhibition. Expression of MIA40 variants in HEK293 cells was induced using 1 μg ml−1 doxycycline for 16 h. Concomitantly, cells were incubated with 1 μM of the proteasome inhibitor MG132. Then, cells were lysed and analyzed by immunoblotting. MIA40Δ108 is present at strongly decreased levels compared to MIA40WT but can be partially stabilized by proteasomal inhibition. Quantification using Image lab. Data from 4 experiments were combined and standard deviations are presented. Black arrowhead, endogenous MIA40; blue arrowhead, signal of MIA40Δ108

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