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Data processing, multi-omic pathway mapping, and metabolite activity analysis using XCMS Online

Nature Protocols volume 13, pages 633–651 (2018)Cite this article

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Abstract

Systems biology is the study of complex living organisms, and as such, analysis on a systems-wide scale involves the collection of information-dense data sets that are representative of an entire phenotype. To uncover dynamic biological mechanisms, bioinformatics tools have become essential to facilitating data interpretation in large-scale analyses. Global metabolomics is one such method for performing systems biology, as metabolites represent the downstream functional products of ongoing biological processes. We have developed XCMS Online, a platform that enables online metabolomics data processing and interpretation. A systems biology workflow recently implemented within XCMS Online enables rapid metabolic pathway mapping using raw metabolomics data for investigating dysregulated metabolic processes. In addition, this platform supports integration of multi-omic (such as genomic and proteomic) data to garner further systems-wide mechanistic insight. Here, we provide an in-depth procedure showing how to effectively navigate and use the systems biology workflow within XCMS Online without a priori knowledge of the platform, including uploading liquid chromatography (LC)–mass spectrometry (MS) data from metabolite-extracted biological samples, defining the job parameters to identify features, correcting for retention time deviations, conducting statistical analysis of features between sample classes and performing predictive metabolic pathway analysis. Additional multi-omics data can be uploaded and overlaid with previously identified pathways to enhance systems-wide analysis of the observed dysregulations. We also describe unique visualization tools to assist in elucidation of statistically significant dysregulated metabolic pathways. Parameter input takes 5–10 min, depending on user experience; data processing typically takes 1–3 h, and data analysis takes ∼30 min.

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Figure 1: Upload of mass spectrometry data (Steps 2–5).
Figure 2: Predictive pathway analysis parameter settings (Step 14).
Figure 3: Predictive metabolic pathway results (Steps 20–24).
Figure 4: Overlapping metabolite information (Steps 24–29).
Figure 5: Predictive metabolites results (Steps 36–40).
Figure 6: Pathway cloud plot (Steps 42–46).
Figure 7: Multi-omics integration (Steps 47–50).
Figure 8: Multi-omics results (Steps 51–59).

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Acknowledgements

The authors thank the following for funding assistance: Ecosystems and Networks Integrated with Genes and Molecular Assemblies (ENIGMA), a Scientific Focus Area Program at Lawrence Berkeley National Laboratory for the US Department of Energy, Office of Science, Office of Biological and Environmental Research under contract number DE-AC02-05CH11231 (G.S.); and the National Institutes of Health (grants R01 GM114368 (G.S.) and PO1 A1043376-02S1 (G.S.)).

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Authors and Affiliations

  1. Center for Metabolomics and Mass Spectrometry, The Scripps Research Institute, La Jolla, California, USA

    Erica M Forsberg, Tao Huan, Duane Rinehart, H Paul Benton, Benedikt Warth, Brian Hilmers & Gary Siuzdak

  2. Department of Chemistry and Biochemistry, San Diego State University, San Diego, California, USA

    Erica M Forsberg

  3. Department of Food Chemistry and Toxicology, University of Vienna, Vienna, Austria

    Benedikt Warth

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  1. Erica M Forsberg
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Contributions

E.M.F. and T.H. contributed equally to writing the manuscript. E.M.F., T.H., D.R., H.P.B., B.H. and G.S. contributed to platform development, and H.P.B., B.W. and G.S. contributed to manuscript writing.

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Correspondence to Gary Siuzdak.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Methods and Supplementary Table 1. (PDF 852 kb)

Supplementary Data 1

Demonstration transcriptomics data set. (ZIP 5 kb)

Supplementary Data 2

Significant protein data set. (ZIP 0 kb)

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Forsberg, E., Huan, T., Rinehart, D. et al. Data processing, multi-omic pathway mapping, and metabolite activity analysis using XCMS Online. Nat Protoc 13, 633–651 (2018). https://doi.org/10.1038/nprot.2017.151

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