TOWLE, David W.; Mount Desert Island Biol. Lab., Salsbury Cove, ME: Functional Genomics of Environmental Adaptations in Marine Crustaceans

The expression of genes in response to changes in salinity and temperature is being examined in our laboratory through the generation of expressed sequence tags and subsequent microarray analysis. Using the traditional gene-by-gene approach and quantitative PCR, the expression of ten different genes has been monitored in gills and other tissues of crabs and lobsters challenged with an environmental stimulus. For example, in the European shore crab Pachygrapsus marmoratus acclimated to normal seawater, salinity dilution leads to rapid up-regulation of transporter and heat-shock protein mRNA levels in all gills tested. In contrast, salinity increase produces an up-regulation of transporter gene expression only in gill number 7 (of 9). Current expressed sequence tag (EST) and microarray programs are developing tools for the global analysis of changes in gene expression. Starting with mixed tissue cDNA libraries normalized to reduce the abundance of highly expressed messages, we have produced 4,604 ESTs for the American lobster Homarus americanus and 5,362 ESTs for the green shore crab Carcinus maenas, available for searching at www.ncbi.nlm.nih.gov. Clustering has identified 3,773 unique sequences for H. americanus and 1,928 unique sequences for C. maenas. Oligonucleotides based on these ESTs are being arrayed for studies of transcriptome changes following environmental perturbation. In addition, the ESTs are being used directly to identify specific targets for further study, including the recent characterization of a zinc transporter in gills of C. maenas. Supported by NSF (DBI-0100394 and IBN-0340622) and NCRR (1-P20-RR16463).