Functional genomics of the North Atlantic right whale (Eubalaena glacialis): Microarray development and transcriptome analysis of cadmium exposure
Ierardi, Jessalyn L.
A cDNA library has been constructed from a skin biopsy as an initial step in defining the transcriptome of the North Atlantic right whale (Eubalaena glacialis) and developing functional genomic tools to study right whale health at the molecular physiological level. 2496 randomly selected clones (expressed sequence tags, ESTs) have been sequenced, and genes identified as important in the response to stress and immune challenges have been cloned by targeted RT-PCR from skin cDNA. The analysis of the EST collection (archived at www.marinegenomics.org and GenBank) showed a 34.79% redundancy, yielding 1578 unigenes and 27 potential microsatellite markers. 96 genes were cloned by targeted PCR with 52 of these genes related to stress and immune function. Gene Ontology analysis of the unigene collection indicates that the skin is a rich source of expressed genes with diverse functions, suggesting an important role in multiple physiological processes including those related to immunity and stress response. An 8X15k oligonucleotide microarray was developed consisting of 2334 E. glacialis probes and 2166 Tursiops truncatus probes and used to measure the transcriptome level effects of right whale kidney fibroblast cells exposed to cadmium. Cells were exposed to three concentrations of cadmium chloride (CdCl2) for three exposure times. Cells exposed to 10-6M CdCl2 for 4 hours and 24 hours showed upregulated genes involved in protection from metal toxicity, oxidative stress, protein renaturation, apoptosis inhibition, and several regulators of cellular processes. Downregulated genes represented a suite of functions including cell proliferation, transcription regulation, actin polymerization, and stress fiber synthesis. The collection of differentially expressed genes in this study support proposed mechanisms of cadmium-induced apoptosis such as mitochondrial membrane potential collapse, reactive oxygen species (ROS) influx, and cell cycle arrest. The results confirm the right whale microarray as a reproducible tool in measuring differentiated gene expression and should be a valuable asset for transcriptome analysis of other baleen whales and potential health assessment protocols.
Northern right whale; Genomics -- Methodology