How nitrogen sources influence Mortierella alpina aging: From the lipid droplet proteome to the whole-cell proteome and metabolome

Abstract

Arachidonic acid (ARA) is a valuable polyunsaturated fatty acid produced by Mortierella alpina. Although some strategies such as nitrogen supplementation have shown the potential to affect the aging of M. alpina in ways which enable it to produce more ARA, the underlying mechanism remains elusive. Herein, we conducted a systematical analysis of the lipid droplet proteome, as well as the whole-cell proteome and metabolome, in order to elucidate how and why two different nitrogen sources (KNO₃ and urea) affect the aging of M. alpina and the corresponding ARA concentration. We found that KNO₃ promoted the ARA concentration, while urea accelerated lipid consumption and stimulated the decomposition of mycelia. Although both KNO₃ and urea activated carbohydrate metabolic pathways, KNO₃ exerted a stronger promoting effect on the pentose phosphate pathway and induced the lipid droplets to participate in the citrate-pyruvate cycle. The activities of malic enzyme and isocitrate dehydrogenase were also promoted more by KNO₃. These pathways provided additional substrates and reducing power for ARA synthesis and ROS elimination. Accordingly, since urea showed a weaker promotion of the related pathways, it caused a depression of the antioxidant system and a consequent increase of ROS. These findings facilitate the design of nitrogen supplementation strategies to achieve higher ARA concentrations, and provide guidance for deciphering the mechanisms of similar aging phenomena in other oleaginous microorganisms.

Significance: Polyunsaturated fatty acids such as arachidonic acid (ARA) are valuable nutrients, which play important roles in preventing numerous diseases and facilitating development. Although it has been found for years that ARA production will be increased in the aging process of Mortierella alpina (M. alpina) and nitrogen sources are involved in this process, the underlying mechanism for this phenomenon remains unknown. In this work, we used the subcellular proteomics, whole-cell proteomics and metabolomics methods to explore the mechanisms by which two different nitrogen (KNO₃ and urea) affected the aging process of M. alpina. Finally, we gave some new insights for the mechanisms mentioned above. This finding will fuel the technology developments for the ARA production using microbes.

Keywords: Aging; Arachidonic acid; Fermentation; Lipid droplet; Nitrogen source; Proteomics.