Lipid droplets accumulation and other biochemical changes induced in the fungal pathogen Ustilago maydis under nitrogen-starvation

Abstract

In many organisms, the growth under nitrogen-deprivation or a poor nitrogen source impacts on the carbon flow distribution and causes accumulation of neutral lipids, which are stored as lipid droplets (LDs). Efforts are in progress to find the mechanism of LDs synthesis and degradation, and new organisms capable of accumulating large amounts of lipids for biotechnological applications. In this context, when Ustilago maydis was cultured in the absence of a nitrogen source, there was a large accumulation of lipid bodies containing mainly triacylglycerols. The most abundant fatty acids in lipid bodies at the stationary phase were palmitic, linoleic, and oleic acids, and they were synthesized de novo by the fatty-acid synthase. In regard to the production of NADPH for the synthesis of fatty acids, the cytosolic NADP-dependent isocitrate dehydrogenase and the glucose-6-phosphate and 6-phosphogluconate dehydrogenases couple showed the highest specific activities, with a lower activity of the malic enzyme. The ATP-citrate lyase activity was not detected in any of the culture conditions, which points to a different mechanism for the transfer of acetyl-CoA into the cytosol. Protein and RNA contents decreased when U. maydis was grown without a nitrogen source. Due to the significant accumulation of triacylglycerols and the particular composition of fatty acids, U. maydis can be considered an alternative model for biotechnological applications.

AnUstilago maydischassis for itaconic acid production without by‐products

“…To stop glycolipid production, repair templates were designed to remove the whole gene clusters for MEL (Hewald et al , ) and UA (Teichmann et al , ) biosynthesis. The production of TAGs, forming intracellular lipid droplets as main energy storage in U. maydis (Aguilar et al , ), was reduced by single gene deletion of UMAG_03937. This gene, subsequently named dgat , encodes a putative diacylglycerol acyltransferase based on sequence alignment of DGAT2 from Saccharomyces cerevisiae (Liu et al , ).…”

“…The molecular structures of the three investigated lipid classes (MELs, UA and TAGs) feature different fatty acid chain length profiles: two short‐ to medium‐chain (C 2 to C 14 ) fatty acids for MELs (Hewald et al , ; Huang et al , ), one short‐chain (C 6 or C 8 ) and a C 16 fatty acid for UA (Teichmann et al , , ), and three long‐chain fatty acids, predominantly C 16 and C 18 , for TAGs (Aguilar et al , ). As expected from the molecular structures, the deletion of glycolipid production, especially ∆MEL, led to a drastic reduction or even complete absence of C 6 , C 12 and C 14 fatty acids (Fig.…”

“…It also has large biotechnological potential due to features such as natural production of a wide range of value‐added molecules, insensitivity to medium impurities and hydromechanic stress, and unicellular growth (Klement et al , ; Klement and Büchs, ; Maassen et al , ). Main products include organic acids (malate, succinate, itaconate, itatartarate and (S)‐2‐hydroxyparaconate), polyols (erythritol and mannitol), glycolipids (mannosylerythritol lipids and ustilagic acid), intracellular triacylglycerols (Guevarra and Tabuchi, ; Bölker et al , ; Moon et al , ; Feldbrügge et al , ; Aguilar et al , ) and proteins (Kämper et al , ; Mueller et al , ; Geiser et al , ). Culture conditions influence the composition of the product spectrum.…”

“…The gene cluster enabling UA biosynthesis is formed by 12 open reading frames including the cytochrome P450 monooxygenase encoded by cyp1 (Hewald et al , ; Teichmann et al , , ). Intracellular triacylglycerol (TAG) formation has not been studied in detail in Ustilago , but in other oleaginous species, diacylglycerol acyltransferases (DGAT) represent main contributors by mediating the final acylation of diacylglycerols to TAGs (Guo et al , ; Aguilar et al , ).…”

Ustilago Rabenhorstiana—An Alternative Natural Itaconic Acid Producer

“…For wildtype strains of the species Ustilago, low final titers of 44.5 g·L −1 itaconic acid, low yields up to 0.24 (w/w), and a low productivity of maximum 0.31 g (L·h) −1 are disadvantageous [19]. This is due to a variety of by-products like other organic acids, glycolipids, and intracellular triacylglycerols, which are produced in parallel to itaconic acid [16,18,20,21]. Nevertheless, in addition to the robustness of the yeasts, Fermentation 2020, 6, 4 3 of 15 The main cultures in shake flasks were carried out at 30 • C and 120 rpm in 250 mL shake flasks with three baffles and a filling volume of 100 mL Tabuchi-medium. To identify potential impurities and the utilization of monosaccharides based on lignocellulosic feedstock, test tubes with Kapsenberg caps were used and a working volume of 2 mL (ø 16 mm × 100 mm).…”