Dynamic Regulation of Mitochondrial Import by the Ubiquitin System

• Published in: Molecular cell 2020-03, Vol.77 (5), p.1107-1123.e10
• Main Authors: Phu, Lilian, Rose, Christopher M., Tea, Joy S., Wall, Christopher E., Verschueren, Erik, Cheung, Tommy K., Kirkpatrick, Donald S., Bingol, Baris
• Format: Article
• Language: English
• Subjects: Animals; Biological Transport; Carrier Proteins - genetics; Carrier Proteins - metabolism; Deubiquitinase; E3 Ubiquitin Ligase; Female; HEK293 Cells; HeLa Cells; Humans; Male; Membrane Proteins - genetics; Membrane Proteins - metabolism; Mice, Inbred C57BL; Mice, Knockout; Mitochondria; Mitochondria - enzymology; Mitochondria - genetics; Mitochondrial Import; Mitochondrial Proteins - genetics; Mitochondrial Proteins - metabolism; Proteasome Endopeptidase Complex - genetics; Proteasome Endopeptidase Complex - metabolism; Proteolysis; Thiolester Hydrolases - genetics; Thiolester Hydrolases - metabolism; Time Factors; TOM complex; Ubiquitin - metabolism; Ubiquitin System; Ubiquitin-Protein Ligases - genetics; Ubiquitin-Protein Ligases - metabolism; Ubiquitination
• ISSN: 1097-2765; 1097-4164
• DOI: 10.1016/j.molcel.2020.02.012

Mitochondria import nearly their entire proteome from the cytoplasm by translocating precursor proteins through the translocase of the outer membrane (TOM) complex. Here, we show dynamic regulation of mitochondrial import by the ubiquitin system. Acute pharmacological inhibition or genetic ablation of the mitochondrial deubiquitinase (DUB) USP30 triggers accumulation of Ub-substrates that are normally localized inside the mitochondria. Mitochondrial import of USP30 substrates is impaired in USP30 knockout (KO) cells, suggesting that deubiquitination promotes efficient import. Upstream of USP30, the E3 ligase March5 ubiquitinates mitochondrial proteins whose eventual import depends on USP30. In USP30 KOs, exogenous March5 expression induces accumulation of unimported translocation intermediates that are degraded by the proteasomes. In USP30 KO mice, TOM subunits have reduced abundance across multiple tissues. Together these data highlight how protein import into a subcellular compartment can be regulated by ubiquitination and deubiquitination by E3 ligase and DUB machinery positioned at the gate. [Display omitted] •USP30 and March5 reciprocally modify import substrates at the TOM complex•USP30 promotes mitochondrial import of ubiquitinated substrates•March5 directs degradation of import intermediates by proteasomes•USP30 KO mice have reduced abundance of the TOM complex Phu et al. show that two ubiquitin system enzymes located on mitochondria, the E3 Ub ligase March5 and the deubiquitinase USP30, associate with the translocase and regulate mitochondrial import. While March5 opposes mitochondrial import and directs degradation of substrates, USP30 deubiquitinates substrates to promote their import.