Meeting Abstract
LBS4.2 Sunday, Jan. 6 Salinity effects on gene expression patterns in gills of the green crab Carcinus maenas TOWLE, David W.*; TERWILLIGER, Nora B.; HENRY, Raymond P.; Mt. Desert Island Biological Lab.; Oregon Institute of Marine Biology; Auburn University dtowle@mdibl.org
Osmoregulatory ion transport across crustacean gill epithelium is energized by basolateral Na+/K+-ATPase and is dependent upon a number of other transporters in the apical and basolateral membrane as well as cytosolic carbonic anhydrase. To further characterize transport mechanisms in this tissue, an analysis of gill transcript abundance following salinity reduction was carried out using DNA microarrays based on 15,637 expressed sequence tags generated from a multiple-tissue normalized cDNA library from the green crab Carcinus maenas. Clustering yielded 4,483 unique transcript sequences used to design 50-mer oligonucleotides probes that were printed in quadruplicate on treated glass slides. Total RNA was isolated from posterior gills at various time intervals following transfer of crabs from normal seawater (34 ppt) to dilute seawater (10 ppt). Messenger RNAs were reverse transcribed and the resulting cDNAs, labeled with Cy3 (controls) or Cy5 (experimental) dyes, were hybridized to the arrays. Analysis of the hybridization results identified 15 transcripts that were strongly induced following salinity transfer, many of which encoded unknown proteins. Examination of the expression of candidate transporters confirmed the induction of Na+/K+-ATPase alpha subunit and carbonic anhydrase mRNA, previously shown via quantitative PCR, and showed for the first time strong induction of a Cl–/HCO3– exchanger as well as a complex pattern of sequential induction and repression of three unique Ca2+-activated Cl– channels. Supported by NSF (IOB-0543860 to DWT and IBN-0230005 to RPH), NIH (P20 RR016463), and an MDIBL New Investigator Award to NBT.
