Targeted Metabolomics Using LC-MS in Neurospora crassa.
LC-MS
Metabolomics
Neurospora crassa
Journal
Current protocols
ISSN: 2691-1299
Titre abrégé: Curr Protoc
Pays: United States
ID NLM: 101773894
Informations de publication
Date de publication:
May 2022
May 2022
Historique:
entrez:
26
5
2022
pubmed:
27
5
2022
medline:
31
5
2022
Statut:
ppublish
Résumé
The filamentous fungus Neurospora crassa has historically been a model for understanding the relationship between genes and metabolism-auxotrophic mutants of N. crassa were used by Beadle and Tatum to develop the one-gene-one-enzyme hypothesis for which they earned the Nobel Prize in 1958. In the ensuing decades, several techniques have been developed for the systematic analysis of metabolites in N. crassa and other fungi. Untargeted and targeted approaches have been used, with a focus on secondary metabolites over primary metabolism. Here, we describe a pipeline for sample preparation, metabolite extraction, Liquid Chromatography-Mass Spectrometry (LC-MS), and data analysis that can be used for targeted metabolomics of primary metabolites in N. crassa. Liquid cultures are grown with shaking in a defined minimal medium and then collected using filtration. Samples are lyophilized for 2 days at -80°C, pulverized, and mixed with a solution to extract polar metabolites. The metabolites are separated and identified using LC-MS, with downstream analysis using Skyline interpretive software. Relative levels of hundreds of metabolites can be detected and compared across strains. © 2022 Wiley Periodicals LLC. Basic Protocol: Metabolite extraction and detection from Neurospora crassa cell cultures using Liquid Chromatography-Mass Spectrometry.
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
e454Subventions
Organisme : National Institute of Food and Agriculture
ID : #CA-R-PPA-6980-H
Informations de copyright
© 2022 Wiley Periodicals LLC.
Références
Adpressa, D. A., Connolly, L. R., Konkel, Z. M., Neuhaus, G. F., Chang, X. L., Pierce, B. R., … Loesgen, S. (2019). A metabolomics-guided approach to discover Fusarium graminearum metabolites after removal of a repressive histone modification. Fungal Genetics and Biology, 132, 103256. doi: 10.1016/j.fgb.2019.103256.
Albright, J. C., Henke, M. T., Soukup, A. A., McClure, R. A., Thomson, R. J., Keller, N. P., & Kelleher, N. L. (2015). Large-scale metabolomics reveals a complex response of Aspergillus nidulans to epigenetic perturbation. ACS Chemical Biology, 10(6), 1535-1541. doi: 10.1021/acschembio.5b00025.
Bai, N., Zhang, G., Wang, W., Feng, H., Yang, X., Zheng, Y., … Yang, J. (2021). Ric8 acts as a regulator of G-protein signalling required for nematode-trapping lifecycle of Arthrobotrys oligospora. Environmental Microbiology, 24, 1714-1730. doi: 10.1111/1462-2920.15735.
Barrios-González, J. (2018). Chapter 13-Secondary metabolites production: Physiological advantages in solid-state fermentation. In A. Pandey, C. Larroche, & C. R. Soccol (Eds.), Current developments in biotechnology and bioengineering (pp. 257-283). Elsevier. doi: 10.1016/B978-0-444-63990-5.00013-X.
Beadle, G. W., & Tatum, E. L. (1941). Genetic control of biochemical reactions in Neurospora. Proceedings of the National Academy of Sciences of the United States of America, 27(11), 499-506. doi: 10.1073/pnas.27.11.499.
Bedair, M., & Sumner, L. W. (2008). Current and emerging mass-spectrometry technologies for metabolomics. TrAC Trends in Analytical Chemistry, 27(3), 238-250. doi: 10.1016/j.trac.2008.01.006.
Bunch, D. R., McShane, A. J., & Wang, S. (2018). Investigation of transition ion ratio variation for liquid chromatography-tandem mass spectrometry: A case study approach. Clinica Chimica Acta, 486, 205-208. doi: 10.1016/j.cca.2018.08.009.
Emwas, A.-H. M. (2015). The strengths and weaknesses of NMR spectroscopy and mass spectrometry with particular focus on metabolomics research. Methods in Molecular Biology, 1277, 161-193. doi: 10.1007/978-1-4939-2377-9_13.
Fernie, A. R., Trethewey, R. N., Krotzky, A. J., & Willmitzer, L. (2004). Metabolite profiling: From diagnostics to systems biology. Nature Reviews. Molecular Cell Biology, 5(9), 763-769. doi: 10.1038/nrm1451.
Gika, H. G., Wilson, I. D., & Theodoridis, G. A. (2014). LC-MS-based holistic metabolic profiling. Problems, limitations, advantages, and future perspectives. Journal of Chromatography B, 966, 1-6. doi: 10.1016/j.jchromb.2014.01.054.
Han, T.-L., Cannon, R. D., Gallo, S. M., & Villas-Bôas, S. G. (2019). A metabolomic study of the effect of Candida albicans glutamate dehydrogenase deletion on growth and morphogenesis. NPJ Biofilms and Microbiomes, 5(1), 13. doi: 10.1038/s41522-019-0086-5.
Jewett, M. C., Hofmann, G., & Nielsen, J. (2006). Fungal metabolite analysis in genomics and phenomics. Current Opinion in Biotechnology, 17(2), 191-197. doi: 10.1016/j.copbio.2006.02.001.
Johnson, C. H., Ivanisevic, J., & Siuzdak, G. (2016). Metabolomics: Beyond biomarkers and towards mechanisms. Nature Reviews. Molecular Cell Biology, 17(7), 451-459. doi: 10.1038/nrm.2016.25.
Judge, M. T., Wu, Y., Tayyari, F., Hattori, A., Glushka, J., Ito, T., … Edison, A. S. (2019). Continuous in vivo metabolism by NMR. Frontiers in Molecular Biosciences, 6, 26. Retrieved from https://www.frontiersin.org/article/10.3389/fmolb.2019.00026.
Kasbekar, D. P., & McCluskey, K. (2013). Neurospora genomics and molecular biology. Caister Academic Press. Retrieved from https://ebookcentral.proquest.com/lib/uvic/detail.action?docID=5897863.
Kim, J. D., Kaiser, K., Larive, C. K., & Borkovich, K. A. (2011). Use of 1H nuclear magnetic resonance to measure intracellular metabolite levels during growth and asexual sporulation in Neurospora crassa. Eukaryotic Cell, 10(6), 820-831. doi: 10.1128/EC.00231-10.
Kolde, R. (2019). pheatmap: Pretty Heatmaps (1.0.12) [Computer software]. Retrieved from https://CRAN.R-project.org/package=pheatmap.
McCluskey, K., Wiest, A., & Plamann, M. (2010). The Fungal Genetics Stock Center: A repository for 50 years of fungal genetics research. Journal of Biosciences, 35(1), 119-126. doi: 10.1007/s12038-010-0014-6.
Mosunova, O., Navarro-Muñoz, J. C., & Collemare, J. (2021). The biosynthesis of fungal secondary metabolites: From fundamentals to biotechnological applications. In Ó. Zaragoza & A. Casadevall (Eds.), Encyclopedia of mycology (pp. 458-476). Elsevier. doi: 10.1016/B978-0-12-809633-8.21072-8.
Nakahata, Y., Kaluzova, M., Grimaldi, B., Sahar, S., Hirayama, J., Chen, D., … Sassone-Corsi, P. (2008). The NAD+-dependent deacetylase SIRT1 modulates CLOCK-mediated chromatin remodeling and circadian control. Cell, 134(2), 329-340. doi: 10.1016/j.cell.2008.07.002.
Ouyang, S., Beecher, C. N., Wang, K., Larive, C. K., & Borkovich, K. A. (2015). Metabolic Impacts of using nitrogen and copper-regulated promoters to regulate gene expression in Neurospora crassa. G3, 5(9), 1899-1908. doi: 10.1534/g3.115.020073.
Perez, E. R., Knapp, J. A., Horn, C. K., Stillman, S. L., Evans, J. E., & Arfsten, D. P. (2016). Comparison of LC-MS-MS and GC-MS analysis of benzodiazepine compounds included in the drug demand reduction urinalysis program. Journal of Analytical Toxicology, 40(3), 201-207. doi: 10.1093/jat/bkv140.
Perkins, D. D., Radford, A., & Sachs, M. S. (2001). The Neurospora compendium: Chromosomal loci. Academic Press. doi: 10.1007/BF02811419.
Radford, A. (2004). Metabolic highways of Neurospora crassa revisited. Advances in Genetics, 52, 165-207. doi: 10.1016/S0065-2660(04)52005-9.
Romsdahl, J., Blachowicz, A., Chiang, Y.-M., Venkateswaran, K., & Wang, C. C. C. (2020). Metabolomic analysis of Aspergillus niger isolated from the international space station reveals enhanced production levels of the antioxidant pyranonigrin A. Frontiers in Microbiology, 11, 931. doi: 10.3389/fmicb.2020.00931.
Stokes, J. L., Larsen, A., Woodward, C. R., & Foster, J. W. (1943). A Neurospora assay for pyridoxine. Journal of Biological Chemistry, 150(1), 17-24. doi: 10.1016/S0021-9258(18)51246-1.
Sumner, L. W., Mendes, P., & Dixon, R. A. (2003). Plant metabolomics: Large-scale phytochemistry in the functional genomics era. Phytochemistry, 62(6), 817-836. doi: 10.1016/s0031-9422(02)00708-2.
Takahashi, H., Ochiai, K., Sasaki, K., Izumi, A., Shinyama, Y., Mohri, S., … Goto, T. (2021). Metabolome analysis revealed that soybean-Aspergillus oryzae interaction induced dynamic metabolic and daidzein prenylation changes. PloS One, 16(7), e0254190. doi: 10.1371/journal.pone.0254190.
Theodoridis, G. A., Gika, H. G., Want, E. J., & Wilson, I. D. (2012). Liquid chromatography-mass spectrometry based global metabolite profiling: A review. Analytica Chimica Acta, 711, 7-16. doi: 10.1016/j.aca.2011.09.042.
Wellen, K. E., Hatzivassiliou, G., Sachdeva, U. M., Bui, T. V., Cross, J. R., & Thompson, C. B. (2009). ATP-citrate lyase links cellular metabolism to histone acetylation. Science, 324(5930), 1076-1080. doi: 10.1126/science.1164097.
Wishart, D. S. (2019). Metabolomics for investigating physiological and pathophysiological processes. Physiological Reviews, 99(4), 1819-1875. doi: 10.1152/physrev.00035.2018.
Wishart, D. S., Tzur, D., Knox, C., Eisner, R., Guo, A. C., Young, N., … Querengesser, L. (2007). HMDB: The human metabolome database. Nucleic Acids Research, 35(Database issue), D521-526. doi: 10.1093/nar/gkl923.
Xu, Y.-F., Lu, W., & Rabinowitz, J. D. (2015). Avoiding misannotation of in-source fragmentation products as cellular metabolites in liquid chromatography-mass spectrometry-based metabolomics. Analytical Chemistry, 87(4), 2273-2281. doi: 10.1021/ac504118y.
Zamani, A. I., Barig, S., Ibrahim, S., Mohd Yusof, H., Ibrahim, J., Low, J. Y. S., … Ng, C. L. (2020). Comparative metabolomics of Phialemonium curvatum as an omnipotent fungus cultivated on crude palm oil versus glucose. Microbial Cell Factories, 19(1), 179. doi: 10.1186/s12934-020-01434-w.
Zhang, S., Nagana Gowda, G. A., Ye, T., & Raftery, D. (2010). Advances in NMR-based biofluid analysis and metabolite profiling. The Analyst, 135(7), 1490-1498. doi: 10.1039/c000091d.