Understanding the molecular mechanisms of redox-dependent mitochondrial homeostasis mediated by DJ-1 paralogs in Saccharomyces cerevisiae

Abstract

Mitochondrial integrity is a crucial determinant of overall cellular health. Mitochondrial dysfunction and impediments in regulating organellar homeostasis contribute majorly to the pathophysiological manifestation of several neurological disorders. Mutations in human DJ-1 (PARK7) have been implicated in the deregulation of mitochondrial homeostasis, a classical cellular etiology observed in the pathogenesis of Parkinson’s disease. DJ-1 is a multifunctional protein that belongs to a highly conserved class of proteins across phylogeny, constitutively called the DJ-1/ThiJ/PfpI superfamily. Although the pathophysiological significance of DJ-1 has been extensively studied, the underlying molecular mechanism by which DJ-1 paralogs contribute to mitochondrial maintenance and response to oxidative stress remains elusive. By utilizing genetic approaches in Saccharomyces cerevisiae as the model organism, we unravel the intricate mechanisms by which DJ-1 paralogs modulate mitochondrial health and redox homeostasis. Mitochondria adapt to a highly tubular network (due to enhanced Fzo1 levels) in the absence of DJ-1 paralogs, leading to an oxidative stress-sensitive cellular population. Our study highlights a synthetic interaction between Ubp2, a cysteine-dependent deubiquitinase, and DJ-1 paralogs. Intriguingly, the loss of Ubp2 restores the mitochondrial mass and integrity in the DJ-1 deletion background by modulating the ubiquitination status of Fzo1. Further, Ubp2 deletion makes cells less sensitive to oxidative stress without DJ-1 paralogs. In conclusion, the first part of our study dissects the crosstalk between Ubp2 and DJ-1 in regulating mitochondrial health and redox homeostasis. The DJ-1 paralogs in Saccharomyces cerevisiae consist of four homologs- Hsp31, Hsp32, Hsp33, and Hsp34, of which Hsp31 shares maximum sequence similarity with human DJ-1. Although the cytoprotective functions of Hsp31 are characterised, the mechanisms regulating its subcellular localization remain elusive. Interestingly, Hsp31 localizes to mitochondria under stress conditions despite lacking a canonical mitochondrial targeting signal. The second part of the current study identifies Tae1, an N-terminal methyltransferase, as a key regulator of Hsp31 redistribution into mitochondria. Tae1 specifically targets proteins with an X-Pro-Lys/Arg motif, catalysing methylation of Hsp31 at the N-terminus. The study highlights that mutating the residues at the N-terminal of Hsp31 modulates mitochondrial redistribution. This suggests that methylation at the N-terminal is integral in regulating the subcellular localisation of Hsp31. These findings uncover a novel post-translational mechanism for non-canonical mitochondrial targeting of proteins and emphasize the role of N-terminal modifications in oxidative stress response. In summary, the current work identifies the molecular mediators and mechanisms underlying DJ-1-dependent maintenance of mitochondrial integrity and response to oxidative stress.