Nitrogen Source Affects Glycolipid Production and Lipid Accumulation in the Phytopathogen Fungus Ustilago maydis


When cultured in medium limited of nitrogen sources, the phytopathogen Ustilago maydis produces two amphipathic glycolipids: Ustilagic acid (UA) and Mannosylerythritol lipid (MEL), which in addition to the hydrophilic moiety, contain dior tri-hydroxylated C16 fatty acids (UA), or C8 and C16 saturated fatty acids (MEL). We compared the growth and morphology of cells in YPD and in minimum media containing glucose and nitrogen sources such as nitrate or urea and those deprived of nitrogen. Nitrogen-starved cells showed a dramatic accumulation of internal lipids identified as lipid droplets when stained with the hydrophobic probe BODIPY; these lipid droplets were enriched in unsaturated fatty acids. Fatty acids in YPD or medium containing nitrate as nitrogen source showed a combination of saturated/unsaturated lipids, but when urea was the nitrogen source, cells only contained saturated fatty acids. The glycolipid profiles produced in the presence or absence of nitrogen showed preferences towards the production of one kind of glycolipid: cells in media containing nitrate or urea produced different proportions of UA/MEL, but under nitrogen starvation cells contained only UA. The emulsification capacity of the glycolipids produced in media with or without nitrogen was similar (72% - 76%). HPLC of the glycolipids allowed the separation of fractions with different emulsifying characteristics. Our results indicate that U. maydis accumulates lipid droplets when deprived of nitrogen source and confirm that UA is not under nitrogen control, but rather that MEL and lipid droplets are produced and oppositely regulated by nitrogen.

Share and Cite:

Zavala-Moreno, A. , Arreguin-Espinosa, R. , Pardo, J. , Romero-Aguilar, L. and Guerra-Sánchez, G. (2014) Nitrogen Source Affects Glycolipid Production and Lipid Accumulation in the Phytopathogen Fungus Ustilago maydis. Advances in Microbiology, 4, 934-944. doi: 10.4236/aim.2014.413104.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Hewald, S., Josephs, K. and Bolker, M. (2005) Genetic Analysis of Biosurfactant Production in Ustilago maydis. Applied and Environmental Microbiology, 71, 3033-3040.
[2] Brefort, T., Doehlemann, G., Mendoza-Mendoza, A., Reissman, S., Djamei, A. and Khaman, R. (2009) Ustilago maydis as a Pathogen. Annual Review of Phytopathology, 47, 423-445.
[3] Spoeckner, S., Wray, V., Nimtz, M. and Lang, S. (1999) Glycolipids of the Smut Fungus Ustilago maydis from Cultivation on Renewable Resources. Applied Microbiology and Biotechnology, 51, 33-39.
[4] Morita, T., Kitagawa, M., Suzuki, M., Yamamoto, S., Sogabe, A., Yanagidani, S., Imura, T., Fukuoka, T. and Kitamoto, D. (2009) A Yeast Glycolipid Biosurfactant, Mannosylerythritol Lipid, Shows Potential Moisturizing Activity toward Cultured Human Skin Cells: The Recovery Effect of MEL-A on the SDS-Damaged Human Skin Cells. Journal of Oleo Science, 58, 639-642.
[5] Morita, T., Fukuoka, T., Imura, T. and Kitamoto, D. (2013) Production of Mannosylerythritol Lipids and Their Application in Cosmetics. Applied Microbiology and Biotechnology, 97, 4691-4700.
[6] Kitamoto, D., Isoda, H. and Nakahara, T. (2002) Functions and Potential Applications of Glycolipid Biosurfactants from Energy-Saving Materials to Gene Delivery Carriers. Journal of Bioscience and Bioengineering, 94, 187-201.
[7] Makkar, R.S. and Cameotra, S.S. (2002) An Update on the Use of Unconventional Substrates for Biosurfactant Production and Their New Applications. Applied Microbiology and Biotechnology, 58, 428-434.
[8] Morita, T., Ishibashi, Y., Fukuoka, T., Imura, T., Sakai, H., Abe, M. and Kitamoto, D. (2009) Production of Glycolipid Biosurfactants, Mannosylerythritol Lipids, Using Sucrose by Fungal and Yeast Strains, and their Interfacial Properties. Bioscience, Biotechnology, and Biochemistry, 73, 2352-2355.
[9] Liu, Y., Koh, C.M. and Ji, L. (2011) Bioconversion of Crude Glycerol to Glycolipids in Ustilago maydis. Bioresource Technology, 102, 3927-3933.
[10] Amaral, P.F., Coelho, M.A., Marrucho, I.M. and Coutinho, J.A. (2008) Biosurfactants from Yeasts: Characteristics, Production and Application. Advances in Experimental Medicine and Biology, 627, 236-249.
[11] Hewald, S., Linne, U., Scherer, M., Marahiel, M.A., Kamper, J. and Bolker, M. (2006) Identification of a Gene Cluster for Biosynthesis of Mannosylerythritol Lipids in the Basidiomycetous Fungus Ustilago maydis. Applied and Environmental Microbiology, 72, 5469-5477.
[12] Teichmann, B., Linne, U., Hewald, S., Marahiel, M.A. and Bolker, M. (2007) A Biosynthetic Gene Cluster for a Secreted Cellobiose Lipid with Antifungal Activity from Ustilago maydis. Molecular Microbiology, 66, 525-533.
[13] Guo, Y., Cordes, K.R., Farese Jr., R.V. and Walther, T.C. (2009) Lipid Droplets at a Glance. Journal of Cell Science, 122, 749-752.
[14] Liu, Y.M., Zhang, C.Y., Shen, X.P., Zhang, X.L., Cichello, S., Guan, H.B. and Liu, P.S. (2013) Microorganism Lipid Droplets and Biofuel Development. BMB Reports, 46, 575-581.
[15] Finn, P.F. and Dice, J.F. (2006) Proteolytic and Lipolytic Response to Starvation. Nutrition, 22, 830-844.
[16] Holliday, R. (1961) The Genetics of Ustilago maydis. Genetical Research, 2, 204-230.
[17] Bozaquel-Morais, B.L., Madeira, J.B., Maya-Monteiro, C.M., Masuda, C.A. and Montero-Lomeli, M. (2010) A New Fluorescence-Based Method Identifies Protein Phosphatases Regulating Lipid Droplet Metabolism. PLoS ONE, 5, e13692.
[18] Knight, J.A., Anderson, S. and Rawle, J.M. (1972) Chemical Basis of the Sulfo-Phospho-Vanillin Reaction for Estimating Total Serum Lipids. Clinical Chemistry, 18, 199-202.
[19] Techaoei, S., Leelapornpisid, P., Santiarwarn, D. and Lumyong, S. (2007) Preliminary Screening of Biosurfactant-Producing Microorganisms Isolated from Hot Spring and Garages in Northern Thailand. KMITL Science and Technology Journal, 7, 38-43.
[20] Bolker, M., Basse, C.W. and Schirawsk, J. (2008) Ustilago maydis Secondary Metabolism—From Genomics to Biochemistry. Fungal Genetics and Biology, 45, S88-S93.
[21] Morita, T., Fukuoka, T., Imura, T. and Kitamoto, D. (2013) Accumulation of Cellobiose Lipids under Nitrogen-Limiting Conditions by Two Ustilaginomycetous Yeasts, Pseudozyma aphidis and Pseudozyma hubeiensis. FEMS Yeast Research, 13, 44-49.
[22] Horts, R.J., Zeh, C., Saur, A., Sonnewald, S., Sonnewald, U. and Voll, L.M. (2012) The Ustilago maydis Nit2 Homolog Regulates Nitrogen Utilization and Is Required for Efficient Induction of Filamentous Growth. Eukaryotic Cell, 11, 368-380.
[23] Ageitos, J.M., Vallejo, J.A., Veiga-Crespo, P. and Villa, T.G. (2011) Oily Yeasts as Oleaginous Cell Factories. Applied Microbiology and Biotechnology, 90, 1219-1227.
[24] Hsieh, H.J., Su, C.H. and Chien, L.J. (2012) Accumulation of Lipid Production in Chlorella minutissima by Triacylglycerol Biosynthesis-Related Genes Cloned from Saccharomyces cerevisiae and Yarrowia lipolytica. Journal of Microbiology, 50, 526-534.
[25] Beopoulos, A., Chardot, T. and Nicaud, J.M. (2009) Yarrowia lipolytica: A Model and a Tool to Understand the Mechanisms Implicated in Lipid Accumulation. Biochimie, 91, 692-696.
[26] Hynes, M.J. and Murray, S.L. (2010) ATP-Citrate Lyase Is Required for Production of Cytosolic Acetyl Coenzyme A and Development in Aspergillus nidulans. Eukaryotic Cell, 9, 1039-1048.
[27] Liu, Z.J., Gao, Y., Chen, J., Imanaka, T., Bao, J. and Hua, Q. (2013) Analysis of Metabolic Fluxes for Better Understanding of Mechanisms Related to Lipid Accumulation in Oleaginous Yeast Trichosporon cutaneum. Bioresource Technology, 130, 144-151.
[28] Klement, T., Milker, S., Jager, G., Grande, P.M., Domínguez de María, P. and Büchs, J. (2012) Biomass Pretreatment Affects Ustilago maydis in Producing Itaconic Acid. Microbial Cell Factories, 11, 43.
[29] Walther, T.C. and Farese Jr., R.V. (2009) The Life of Lipid Droplets. Biochimica et Biophysica Acta, 1791, 459-466.
[30] Morita, E., Kumon, Y., Nakahara, T., Kagiwada, S. and Noguchi, T. (2010) Docosahexaenoic Acid Production and Lipid-Body Formation in Schizochytrium limacinum SR21. Marine Biotechnology, 8, 319-327.
[31] Siaut, M., Cuiné, S., Cagnon, C., Fessler, B., Nguyen, M., Carrier, P., Beyly, A., Beisson, F., Triantaphylidès, C., Li-Beisson, Y. and Peltier, G. (2011) Oil Accumulation in the Model Green Alga Chlamydomonas reinhardtii: Characterization, Variability between Common Laboratory Strains and Relationship with Starch Reserves. BMC Biotechnology, 11, 7.
[32] Levine, B. and Klionsky, D.J. (2004) Development by Self-Digestion: Molecular Mechanisms and Biological Functions of Autophagy. Developmental Cell, 6, 463-477.
[33] Lemieux, R.U. and Charanduk, R. (1951) Biochemistry of the Ustilaginales VI. The Acyl Groups of Ustilagic Acid. Canadian Journal of Chemistry, 29, 759-766.
[34] Bhattacharjee, S.S., Haskins, R.H. and Gorin, P.A.J. (1970) Location of Acyl Groups on Two Acylated Glycolipids from Strains of Ustilago (Smut Fungi). Carbohydrate Research, 13, 235-246.
[35] Buerth, C., Kovacic, F., Stock, J., Terfüchete, M., Wilhelm, S., Jaeger, K.E., Feldbrügge, M., Schipper, K., Ernest, J.F. and Tielker, D. (2014) Uml2 Is a Novel CalB-Type Lipase of Ustilago maydis with Phospholipase A Activity. Applied Microbiology and Biotechnology, 98, 4963-4973.
[36] Morita, T., Konishi, M., Fukuoka, T., Imura, T. and Kitamoto, D. (2007) Physiological Differences in the Formation of the Glycolipid Biosurfactants, Mannosylerythritol Lipids, between Pseudozyma antarctica and Pseudozyma aphidis. Applied Microbiology and Biotechnology, 74, 307-315.
[37] Irudayaraj, J., Bhaduri, S., Uppara, P.V. and Doble, M. (2008) Mannosylerythritol Lipids: A Review. Journal of Industrial Microbiology & Biotechnology, 35, 1559-1570.

Copyright © 2022 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.