Mitochondrial carrier protein overloading and misfolding induce aggresomes and proteostatic adaptations in the cytosol

Abstract

Previous studies in yeast showed that mitochondrial stressors not directly targeting the protein import machinery can cause mitochondrial precursor overaccumulation stress (mPOS) in the cytosol independent of bioenergetics. Here, we demonstrate mPOS and stress responses in human cells. We show that overloading of mitochondrial membrane carrier, but not matrix proteins, is sufficient to induce cytosolic aggresomes and apoptosis. The aggresomes appear to triage unimported mitochondrial proteins. Interestingly, expression of highly unstable mutant variants of the mitochondrial carrier protein, Ant1, also induces aggresomes despite a greater than 20-fold reduction in protein level compared to wild type. Thus, overloading of the protein import machinery, rather than protein accumulation, is critical for aggresome induction. The data suggest that the import of mitochondrial proteins is saturable and that the cytosol is limited in degrading unimported mitochondrial proteins. In addition, we found that EGR1, eEF1a, and ubiquitin C are up-regulated by Ant1 overloading. These proteins are known to promote autophagy, protein targeting to aggresomes, and the processing of protein aggregates, respectively. Finally, we found that overexpression of the misfolded variants of Ant1 induces additional cytosolic responses including proteasomal activation. In summary, our work captured a profound effect of unimported mitochondrial proteins on cytosolic proteostasis and revealed multiple anti-mPOS mechanisms in human cells.

The mitochondrial carrier SFXN1 is critical for complex III integrity and cellular metabolism

“…Moreover, stably overexpressing SFXN1 in SFXN1 KO partially improved CIII function ( Figures S4D and S4E ). We speculate that reintroducing SFXN1 at a physiological level may provide a better rescue, as overexpression of MCs has been shown to induce an aggregation-based, cytosolic stress response ( Liu et al, 2019 ).…”

Increased levels of the mitochondrial import factor Mia40 prevent the aggregation of polyQ proteins in the cytosol

“…Presumably owing to their hydrophobic nature, carrier proteins were reported in the past as being a severe burden for cytosolic proteostasis when their mitochondrial import is impaired (Hoshino, Wang et al, 2019, Liu, Wang et al, 2019, Piard, Umanah et al, 2018, Wang & Chen, 2015). We employed an established assay of PROTEOSTAT staining (Pihlasalo, Kirjavainen et al, 2011) to monitor the formation of misfolded proteins in yeast cells.…”

“…However, major clients of the Mia40 pathway are the small Tim proteins, essential components for the import of inner membrane carriers (Koehler, Jarosch et al, 1998, Luciano, Vial et al, 2001). Carriers are very abundant hydrophobic proteins and it is well documented that misfolded or non-imported carriers pose a major threat to cytosolic proteostasis (Hoshino et al, 2019, Liu et al, 2019, Vartiainen, Chen et al, 2014, Wang & Chen, 2015). Consistent with this idea, a previous genome-wide overexpression screen for suppressors of polyQ toxicity identified a large number of components of the carrier import pathway, including Erv1, Tim9, Tim10, Tim12, and Tim22 (Mason et al, 2013).…”

Cytosolic Adaptation to Mitochondrial Precursor Overaccumulation Stress Induces Progressive Muscle Wasting

“…Our recent studies showed that overloading of mitochondria with carrier proteins leads to the accumulation of unimported mitochondrial proteins in the cytosol, followed by proteostatic stress (or mPOS) and cell death 5 . However, it is unknown whether mPOS can occur in vivo and whether it causes pathology.…”