Ribosome-associated chaperones integrate co-translational mitochondrial protein import, stress signaling and proteostasis in Saccharomyces cerevisiae.
TL;DR
It is shown that NAC and Ssb1/2 have distinct yet partially overlapping functions in the handling of mitochondrial precursor proteins, whereas NAC limits the accumulation of misfolded proteins at the mitochondrial surface.
Abstract
Mitochondrial biogenesis requires the coordinated synthesis, targeting, and import of nuclear-encoded mitochondrial precursor proteins. Although ribosome-associated chaperones support co-translational protein folding, their genetic contributions to mitochondrial protein import and cellular homeostasis remain incompletely defined. Here, we investigate the roles of the nascent polypeptide-associated complex (NAC) and the ribosome-associated Hsp70 system Ssb1/2 in Saccharomyces cerevisiae. We show that NAC and Ssb1/2 have distinct yet partially overlapping functions in the handling of mitochondrial precursor proteins. Loss of NAC activates the mitochondrial retrograde pathway and enhances growth on ethanol as a non-fermentable carbon source without compromising respiratory competence, indicating metabolic adaptation rather than overt mitochondrial dysfunction. In contrast, Ssb1/2 deficiency disrupts cytosolic proteostasis, sensitizes cells to TORC1 inhibition, and impairs autophagy and mitophagy. Using a TEV protease-based import reporter, we show that Ssb1/2 promotes efficient co-translational distribution of precursor proteins, whereas NAC limits the accumulation of misfolded proteins at the mitochondrial surface. Biochemical analyses further reveal that Ssb1/2 supports the association of translating cytosolic ribosomes with the mitochondrial outer membrane, while NAC loss partially restores this interaction in the absence of Ssb1/2. Together, these findings establish NAC and Ssb1/2 as key components of an integrated network linking co-translational targeting, mitochondrial signaling, and cellular homeostasis.