The intermembrane space protein Mix23 is a novel stress-induced mitochondrial import factor

Abstract

The biogenesis of mitochondria requires the import of hundreds of precursor proteins. These proteins are transported post-translationally with the help of chaperones, meaning that the overproduction of mitochondrial proteins or the limited availability of chaperones can lead to the accumulation of cytosolic precursor proteins. This imposes a severe challenge to cytosolic proteostasis, and triggers a specific transcription program, called the mitoprotein-induced stress response, which activates the proteasome system. This coincides with the repression of mitochondrial proteins, including many proteins of the intermembrane space (IMS). In contrast, herein we report that the so far uncharacterized IMS protein Mix23 is considerably up-regulated when mitochondrial import is perturbed. Mix23 is evolutionarily conserved and a homolog of the human coiled-coil domain containing 58, CCDC58. We found that, like the subunits of the proteasome, Mix23 is under control of the transcription factor Rpn4. It is imported into mitochondria by the mitochondrial disulfide relay. Mix23 is critical for the efficient import of proteins into the mitochondrial matrix particularly if the function of the TIM23 translocase is compromised such as in temperature-sensitive mutants of Tim17. Our observations identify Mix23 as a novel regulator or stabilizer of the mitochondrial protein import machinery that is specifically upregulated upon mitoprotein-induced stress conditions.

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

“…Is the modulation of Mia40 levels relevant under physiological conditions? In contrast to other components of the mitochondrial import machinery, the expression of Mia40 is upregulated upon stress conditions induced by mitochondrial defects (Boos et al , 2019; Zöller, Laborenz et al , 2020) but also by problems outside mitochondria (Metzger & Michaelis, 2009). Transcription of MIA40 is under control of Rpn4, a transcription factor that adjusts proteasome levels (Xie & Varshavsky, 2001).…”

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

“…Is the modulation of Mia40 levels relevant under physiological conditions? In contrast to other components of the mitochondrial import machinery, the expression of Mia40 is upregulated upon stress conditions induced by mitochondrial defects (Boos et al, 2019, Zöller, Laborenz et al, 2020) but also by problems outside mitochondria (Metzger & Michaelis, 2009). Transcription of MIA40 is under control of Rpn4, a transcription factor that adjusts proteasome levels (Xie & Varshavsky, 2001).…”

“…The following methods were performed according to published methods: Isolation of mitochondria and oxygen consumption measurements (Saladi, Boos et al, 2020); analysis of mRNA levels by qRT-PCR (Zöller et al, 2020); preparation of semi-intact cells and their use for in vitro import experiments (Laborenz et al, 2019); pulse chase assay and alkylation for in vivo analysis of Mia40-mediated import (Peleh et al, 2016); the analysis of mitochondrial protein import using the Oxa1-Ura3 reporter assay (Hansen et al, 2018); growth and analysis of MIA40-expressing HEK293T cells (Fischer, Horn et al, 2013, Murschall, Gerhards et al, 2020).…”

The ER protein Ema19 facilitates the degradation of nonimported mitochondrial precursor proteins

“…Precursor proteins that accumulate in the cytosol can be highly toxic (Wang and Chen, 2015;Wrobel et al, 2015) and elicit response programs to counteract these deleterious consequences (Nargund et al, 2012;Weidberg and Amon, 2018;Boos et al, 2019;Song et al, 2020;Zöller et al, 2020). The stability of many precursor proteins in the cytosol is low thanks to surveillance of the ubiquitin/proteasome system for uninserted precursors (Bragoszewski et al, 2017;Kowalski et al, 2018;Paasch et al, 2018;Saladi et al, 2020;Shakya et al, 2020).…”