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. 2018 Jan 8;7:e30015. doi: 10.7554/eLife.30015

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© 2018, Simões et al

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

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Figure 7. — (A) Effect of Ubp2^C745S on Fzo1^K464R ubiquitylation. HA-Fzo1 or HA-Fzo1^K464R were expressed in the presence of Ubp2 (∆fzo1 cells plus empty vector) or instead in the presence of Ubp2^C745S (∆ubp2 ∆fzo1 plus Ubp2^C745S-Flag), as indicated. Crude mitochondrial extracts were solubilized and HA-tagged Fzo1 was analyzed by SDS-PAGE and immunoblotting using an HA-specific antibody. Unmodified and ubiquitylated forms of HA-Fzo1 are indicated as in 2B. (B) Effect of UBP2 deletion on the steady state levels of Fzo1^K464R. Total cellular extracts of indicated strains expressing HA-Fzo1 or HA-Fzo1^K464R as indicated were analyzed by SDS-PAGE and immunoblotting using HA- and Tom40-specific antibodies. Bottom panel, quantification of five independent experiments, including SD. (C) Effect of Ubp2 and Mdm30 on the steady state levels of Fzo1. Total cellular extracts of wt, Δubp2 and Δubp2 Δmdm30 cells expressing HA-tagged Fzo1 endogenously (HA-Fzo1^int) were analyzed by SDS-PAGE and immunoblotting using HA- and Tom40-specific antibodies. Bottom panel, quantification of three independent experiments, including SD. PoS, Ponceau S staining.