Systematic multi‐level analysis of an organelle proteome reveals new peroxisomal functions

Abstract

Seventy years following the discovery of peroxisomes, their complete proteome, the peroxi-ome, remains undefined. Uncovering the peroxiome is crucial for understanding peroxisomal activities and cellular metabolism. We used high-content microscopy to uncover peroxisomal proteins in the model eukaryote -Saccharomyces cerevisiae. This strategy enabled us to expand the known peroxi-ome by ~40% and paved the way for performing systematic, whole-organellar proteome assays. By characterizing the sub-organellar localization and protein targeting dependencies into the organelle, we unveiled non-canonical targeting routes. Metabolomic analysis of the peroxi-ome revealed the role of several newly identified resident enzymes. Importantly, we found a regulatory role of peroxisomes during gluconeogenesis, which is fundamental for understanding cellular metabolism. With the current recognition that peroxisomes play a crucial part in organismal physiology, our approach lays the foundation for deep characterization of peroxisome function in health and disease.

Saccharomyces cerevisiaeDmo2p is involved in the mitochondrial copper metabolism and is required for the stability of newly translated Cox2p

“…Indeed, most IMS proteins contain a conserved pattern of cysteine residues present in the “twin CX 3 C or CX 9 C motifs” (Vögtle et al, 2012) with a direct impact on the regulation of redox-sensitive cellular processes between the mitochondria and other compartments of the cell (Nuebel et al, 2016). Similarly to Dmo2p, many IMS products present dual localization (Yifrach et al, 2022); in this study, we corroborate these previous observations about Dmo2p localization.…”

“…Oleate is an oxidative medium known to induce peroxisome biogenesis and activity (Venhuis et al, 1987). This experiment was motivated by previous findings that suggested the presence of Dmo2p associated with peroxisome membranes or membrane periphery (Yifrach et al, 2022). In yeast, peroxisomes carry out the β-oxidation of fatty acids to generate acetyl-CoA, which produces high levels of reactive oxygen species (ROS) as byproducts; indeed, mitochondria and peroxisome play central roles in the metabolism of ROS, with a common evolutionary origin (Bolte et al, 2015), they also share components of the fusion-fission machinery (Fransen et al, 2017).…”

Integrative Omics reveals changes in the cellular landscape of yeast without peroxisomes

“…Horizontal and vertical lines indicate a p-value of 0.05 and a fold-change of ≤ 0.66 or ≥ 1.5, respectively. Information about a peroxisomal localization of proteins or association with peroxisomes is based on (David et al, 2022; Elbaz-Alon et al, 2014; Marelli et al, 2004; Motley et al, 2012; Tanaka et al, 2014; Yifrach et al, 2022). PO, peroxisomal and peroxisome-associated proteins or transcripts.…”

“…Recent studies on S. cerevisiae peroxisomes (Yifrach et al, 2022) and mitochondria (Morgenstern et al, 2017) showed that the proteomes of both organelles contain numerous multi-localized proteins. Thus, we hypothesized that solute transporters of other cellular compartments might partially localize to peroxisomes.…”

“…Mutant screens were performed to isolate strains defective in various aspects of peroxisome biology. The analysis of yeast libraries by high throughput fluorescence microscopy (FM) and proteomics studies of isolated peroxisomes or affinity-purified protein complexes of peroxisomal proteins, mostly peroxins, resulted in the discovery of many additional proteins and processes related to peroxisome biology (David et al, 2013; Grunau et al, 2009; Marelli et al, 2004; Mast et al, 2016; Yifrach et al, 2022). Peroxisomes cannot function in isolation but must communicate and cooperate with their environment to exchange metabolites and coordinate cellular responses (Silva et al, 2020).…”