'Systems toxicology' approach identifies coordinated metabolic responses to copper in a terrestrial non-model invertebrate, the earthworm Lumbricus rubellus

doi:10.1186/1741-7007-6-25

BMC Biology

Volume 6

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Bundy JG
Sidhu JK
Rana F
Spurgeon DJ
Svendsen C
Wren JF
Stürzenbaum SR
Morgan AJ
Kille P

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Sidhu JK
Rana F
Spurgeon DJ
Svendsen C
Wren JF
Stürzenbaum SR
Morgan AJ
Kille P
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Research article

'Systems toxicology' approach identifies coordinated metabolic responses to copper in a terrestrial non-model invertebrate, the earthworm Lumbricus rubellus

Jacob G Bundy¹ , Jasmin K Sidhu¹ , Faisal Rana¹ , David J Spurgeon² , Claus Svendsen² , Jodie F Wren³^,4 , Stephen R Stürzenbaum⁵ , A John Morgan³ and Peter Kille³

¹Department of Biomolecular Medicine, Division of Surgery, Oncology, Reproductive Biology, and Anaesthetics (SORA), Faculty of Medicine, Imperial College London, SW7 2AZ, London, UK

²Centre for Ecology and Hydrology, Monks Wood, Abbots Ripton, Huntingdon PE28 2LS, UK

³School of Biosciences, University of Cardiff, Main Building, Park Place, Cardiff, CF10 3TL, UK

⁴School of Biological Sciences, University of Bristol, Woodland Road, Bristol, BS8 1UG, UK

⁵School of Biomedical and Health Sciences, Pharmaceutical Sciences Division, King's College London, Franklin Wilkins Building, Stamford Street, London, SE1 9NH, UK

author email corresponding author email

BMC Biology 2008, 6:25doi:10.1186/1741-7007-6-25


Published:	3 June 2008

Additional files

Additional file 1:

A 600 MHz ¹H NMR spectrum of typical earthworm extract, polar fraction. (A) and (B) have an expanded vertical scale compared with (C) and (D). Resonance from HEFS (compound 19) at 6.19 ppm is not represented at its full height. Metabolite labels correspond to numbers given in Table 1. * represents an unknown compound that is a probable breakdown product of HEFS.

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Additional file 2:

Chemical shift regions for integrals of lipid extract spectral data ('missing' int01 was for internal standard TMS).

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Additional file 3:

NMR integrals of lipid data, heatmap showing individual replicates.

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Additional file 4:

600 MHz ¹H spectra of lipid extracts. Samples only shown from control (blue) and highest (red) dose groups. One spectrum from red group was excluded as an outlier and is not shown here. (A) Vinylic protons from unsaturated fatty acids; (B) unassigned; (C) glycerol protons from triacylglycerols; (D) glycerol peaks from glycerophospholipids; (E) protons allylic to two double bonds; (G) protons allylic to one double bond; (H) and (I) terminal methyls.

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Additional file 5:

Table of transcripts showing more than twofold change in expression as a consequence of copper exposure.

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Additional file 6:

Table of transcripts significantly changed in earthworms exposed to copper. ANOVA, p < 0.05, Benjamini and Hochberg [56] FDR correction.

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Additional file 7:

Table of GO terms overrepresented in transcripts whose expression is altered by copper exposure by more than twofold.

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Additional file 8:

Table of the targeted functional transcript changes observed during copper exposure of adult earthworms.

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Additional file 9:

Comparison of 6% perchloric acid (red) and chloroform/methanol (blue) extraction methods for earthworm tissue: (A) aromatic region; (B) aliphatic region.

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Additional file 10:

Assessment of micro-array sensitivity and signal linearity. Representative analysis of the fluorescent signal generated by 10 RNAs introduced at known concentrations prior to labelling and detected by complementary reporter (10 replicates of each reporter spotted on the array). Data were generated from representative arrays selected from transcript analyses performed on RNA extracted from control and copper-dosed samples. (A)-(E) represent data from copper exposures of 0, 10, 40, 160 and 480 mg/kg, respectively. The average signal is indicated by closed circles with technical error bars representing the standard error of the measurements. A fitted regression line is shown for the linear portion of the response together with the R²value for the fitted line.

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Additional file 11:

Graphical representations of relative gene expression against fluorescence intensity from control and copper-exposed samples. Array data were normalised and filtered (as described in Methods) and the log₂of the average fold change (M) plotted against the log₂of the average mean signal intensity (A). (A)-(E) represent data from copper exposure of 0, 10, 40, 160 and 480 mg/kg, respectively.

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Additional file 12:

Table of metabolite concentrations as determined by 600 MHz ¹H spectroscopy (μmol/mg tissue dry weight).

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