Two NAD-linked redox shuttles maintain the peroxisomal redox balance in Saccharomyces cerevisiae

Nadal A Al-Saryi,Ronald J A Wanders,Carlo W T Van Roermund,Lakhan Ekal,Chantell Payton,Murtakab Y Al-Hejjaj,Georgia E Hulmes,Ewald H Hettema

doi:10.1038/s41598-017-11942-2

Abstract

In Saccharomyces cerevisiae, peroxisomes are the sole site of fatty acid β-oxidation. During this process, NAD+ is reduced to NADH. When cells are grown on oleate medium, peroxisomal NADH is reoxidised to NAD+ by malate dehydrogenase (Mdh3p) and reduction equivalents are transferred to the cytosol by the malate/oxaloacetate shuttle. The ultimate step in lysine biosynthesis, the NAD+-dependent dehydrogenation of saccharopine to lysine, is another NAD+-dependent reaction performed inside peroxisomes. We have found that in glucose grown cells, both the malate/oxaloacetate shuttle and a glycerol-3-phosphate dehydrogenase 1(Gpd1p)-dependent shuttle are able to maintain the intraperoxisomal redox balance. Single mutants in MDH3 or GPD1 grow on lysine-deficient medium, but an mdh3/gpd1Δ double mutant accumulates saccharopine and displays lysine bradytrophy. Lysine biosynthesis is restored when saccharopine dehydrogenase is mislocalised to the cytosol in mdh3/gpd1Δ cells. We conclude that the availability of intraperoxisomal NAD+ required for saccharopine dehydrogenase activity can be sustained by both shuttles. The extent to which each of these shuttles contributes to the intraperoxisomal redox balance may depend on the growth medium. We propose that the presence of multiple peroxisomal redox shuttles allows eukaryotic cells to maintain the peroxisomal redox status under different metabolic conditions.

Highlights

Eukaryotic cells are compartmentalised into distinct, membrane-bound organelles which contain their own unique enzyme content and internal milieu
We provide evidence that lysine biosynthesis can be maintained by both the Mdh3p- and the Gpd1p-dependent shuttles during growth on glucose medium
It has been suggested that peroxisomal Gpd1p is part of a redox shuttle required to regenerate NAD+ formed during fatty acid oxidation

Summary

Introduction

Eukaryotic cells are compartmentalised into distinct, membrane-bound organelles which contain their own unique enzyme content and internal milieu. We provide evidence that lysine biosynthesis can be maintained by both the Mdh3p- and the Gpd1p-dependent shuttles during growth on glucose medium. We further show that when NAD+-linked saccharopine dehydrogenase is mislocalised to the cytosol, lysine biosynthesis occurs independently of the peroxisomal redox shuttles.

Results

Conclusion