Publication Abstract

Development of a stickleback (Gasterosteus aculeatus) cDNA microarray and gene expression responses to dibenzanthracene, ethinyl-estradiol and copper.

Development of a stickleback (Gasterosteus aculeatus) cDNA microarray and gene expression responses to dibenzanthracene, ethinyl-estradiol and copper

T.D. Williams, M. Brown, J.K. Chipman, F. Falciani, F. Ortega, F. Geoghegan, J.A. Craft, I. Katsiadaki, J. Ball, C. Tyler and E. Santos

The Three-spined stickleback (Gasterosteus aculeatus) is a fish species which inhabits European estuarine and freshwater environments and is a useful model for ecotoxicology and, as a genome-sequenced species, ecotoxicogenomics.

We have constructed a cDNA array for the stickleback using SSH and normalised liver cDNA libraries (Abstract Brown et al) and a whole body cDNA library (Prof. Kingsley, Stanford, USA). Bioinformatic analyses of the sequenced clones have been carried out with a view to identification and functional annotation by Gene Ontology category (GO). This has used the Blast2GO program (A. Conesa, S. Goetz). Additional annotation has been derived from the Stickleback genome project.

Our initial microarray experiments with the stickleback have characterised their acute responses to dibenzanthracene (DbA), ethinyl-estradiol (EE2) and copper (Cu) treatment at a range of concentrations. Male sticklebacks were exposed to waterborne DbA at 10 to 50,000 ng/l, EE2 at 0.1 to 100 ng/l or Cu at 3.2 to 128 ug/l for 4 days. The microarray was used to compare gene expression of exposed fish to those of relevant controls. A dual sample preparation procedure was developed allowing metabolomic analysis of the same samples. Both pooled samples and individuals were investigated. Dose responses were found for known biomarker genes CYP1A (r²=0.90), vitellogenin (r²=0.97) and metallothionein (r²=0.88) to DbA, EE2 and Cu, respectively. Additional responsive genes and Gene Ontology categories will be described, providing insights into the mechanistic modes of action of these three model pollutants and highlighting additional biomarkers of exposure. Future experiments will study gene expression in environmentally-sampled fish and in fish exposed to contaminated water for six months in flumes. We therefore describe the establishment and evaluation of a useful resource for toxicogenomics and ecotoxicogenomics.

Reference

T.D. Williams, M. Brown, J.K. Chipman, F. Falciani, F. Ortega, F. Geoghegan, J.A. Craft, I. Katsiadaki, J. Ball, C. Tyler and E. Santos (2009) Development of a stickleback (Gasterosteus aculeatus) cDNA microarray and gene expression responses to dibenzanthracene, ethinyl-estradiol and copper. PRIMO 14

T.D. Williams, M. Brown, J.K. Chipman, F. Falciani, F. Ortega, F. Geoghegan, J.A. Craft, I. Katsiadaki*, J. Ball, C. Tyler and E. Santos

PRIMO 14