A piggybacking mechanism enables peroxisomal localization of the glyoxylate cycle enzyme Mdh2 in yeast

Abstract

Eukaryotic cells evolved organelles that allow the compartmentalization and regulation of metabolic processes. Knowledge on molecular mechanisms that allow temporal and spatial organization of enzymes within organelles is therefore critical for understanding eukaryotic metabolism. Here we show that the yeast malate dehydrogenase 2 (Mdh2) is dually localized to the cytosol and to peroxisomes and is targeted to peroxisomes via association with Mdh3 and a Pex5-dependent piggybacking mechanism. The dual localization of Mdh2 contributes to our understanding of the glyoxylate cycle and provides a new perspective on compartmentalization of cellular metabolism, which is critical for the perception of metabolic disorders and aging.

Identification of Pex34p as a component of the peroxisomal de novo biogenesis machinery in yeast

“…Mdh2 was described as an enzyme involved in both, the glyoxylate cycle and fatty acid β-oxidation (Kunze et al, 2006). Initially described as a cytosolic protein, recent data demonstrated a fraction of Mdh2p to be localized to peroxisome, depending on Pex5p as well as Mdh3p (Gabay-Maskit et al, 2020). Deletion of YAL016C-B - a dubious open reading frame that overlaps with the promotor region of TPD3 caused a mixed phenotype similar to those observed in doa4 Δ and mdh2 Δ cells.…”

“…The generated query strain was crossed into the yeast deletion, hypomorphic allele and mini-peroxi collections (Breslow et al, 2008, Gabay-Maskit et al, 2020, Giaever et al, 2002) by the synthetic genetic array method (Cohen & Schuldiner, 2011, Tong & Boone, 2006). This procedure resulted in a collection of haploid strains used for screening mutants that display peroxisomal abnormalities after re-expression of Pex19p.…”

The trypanosome UDP-glucose pyrophosphorylase is imported by piggybacking into glycosomes where unconventional sugar nucleotide synthesis takes place

“…Piggybacking has been described as an import mechanism with relatively low efficiency in four out of five examples of physiological hetero-oligomer import into peroxisomes reported so far, i.e. superoxide dismutase (SOD1) (24) and lactate dehydrogenase (LDH) (25) in mammals, pyrazinamidase/nicotinamidase (PNC1) (28) and malate dehydrogenase 2 (Mdh2) (27) in yeast, which are co-imported with the PTS-containing copper chaperone SOD1 (CCS), read-through-extended LDH (LDHBx), glycerol-3-phosphate dehydrogenase (GPD1) and Mdh3, respectively. These four co-imported proteins display dual peroxisomal and cytosolic localisation with the majority remaining within the cytosol (27, 50).…”

“…More importantly, the UGP/PEPCK association provides the first example of hetero-oligomeric import by piggybacking involving two proteins not functionally related, since PEPCK is involved in the maintenance of the glycosomal redox and ATP/ADP balances, as well as gluconeogenesis (29, 36). Indeed, among the other known examples of piggybacking, CCS is the chaperone of SOD1 (24), LDH and LDHBx are encoded by the same gene (25), Mdh2 and Mdh3 are Mdh isoforms (27), the PST1-containing phosphatase B subunit and phosphatases A/C subunits form an heterotrimeric enzymatic complex (26), however, the peroxisomal functions of PNC1 and GPD1 are unknown (28).…”

“…Alternatively, proteins lacking a PTS can be imported into the organelle by piggybacking through interaction with a PTS-containing protein. The very few examples of piggybacking described so far in peroxisomes of mammals (24, 25), plants (26) and yeast (27, 28) involve hetero-oligomeric complexes formed by protein isoforms or by functionally related proteins. This mechanism of import has been proposed as an explanation to the presence of some PTS-lacking proteins within glycosomes, but has not yet been reported in trypanosomatids so far.…”