Induction of Oxidative Stress in Sirtuin Gene-Disrupted Ashbya gossypii Mutants Overproducing Riboflavin

AgHST1 and AgHST3 genes encode sirtuins that are NAD(+)-dependent protein deacetylases. According to previous reports, their disruption leads to the overproduction of riboflavin in Ashbya gossypii. In this study, we investigated the potential causes of riboflavin overproduction in the AgHST1 Delta and AgHST3 Delta mutant strains of A. gossypii. The generation of reactive oxygen species was increasd in the mutants compared to in WT. Additionally, membrane potential was lower in the mutants than in WT. The NAD(+)/NADH ratio in AgHST1 Delta mutant strain was lower than that in WT; however, the NAD(+)/NADH ratio in AgHST3 Delta was slightly higher than that in WT. AgHST1 Delta mutant strain was more sensitive to high temperatures and hydroxyurea treatment than WT or AgHST3 Delta. Expression of the AgGLR1 gene, encoding glutathione reductase, was substantially decreased in AgHST1 Delta and AgHST3 Delta mutant strains. The addition of N-acetyl-L-cysteine, an antioxidant, suppressed the riboflavin production in the mutants, indicating that it was induced by oxidative stress. Therefore, high oxidative stress resulting from the disruption of sirtuin genes induces riboflavin overproduction in AgHST1 Delta and AgHST3 Delta mutant strains. This study established that oxidative stress is an important trigger for riboflavin overproduction in sirtuin gene-disrupted mutant strains of A. gossypii and helped to elucidate the mechanism of riboflavin production in A. gossypii.