SERRANO, L.A.**; HENRY, R.P.; Auburn Univ.; Auburn Univ.: Salinity-induced changes in Carbonic Anhydrase activity and mRNA expression in the blue crab Callinectes sapidus

Two isoforms of the enzyme carbonic anhydrase (CA) were cloned from the gills of the blue crab, Callinectes sapidus. Specific primers were designed to study the induction during low salinity acclimation of the isoform showing 70% identity with the cytoplasmic CA isoform from the gills of the green crab, Carcinus maenas. First, hemolymph osmolality, CA activity, and CA RNA expression were measured in crabs acclimated to 35 ppt and transferred to 15 ppt for a time course of between 2 hr and 7 days. In a second experiment, crabs were transferred directly to 25, 15, or 5 ppt salinity for 4 days. During the 7 d time course, osmolality decreased during the first 12 hr (1100 to 820 mOsm) and stabilized around 780 mOsm by 24 hr. CA RNA expression in posterior gills (G7) increased 6 fold by 6 hr post-transfer and then decreased at 12-24 hr. There was a second rise in CA RNA expression at 7 d after transfer. There were no changes in CA expression in anterior gills (G3). Induction of CA activity lagged behind changes in RNA expression. The initial increase in CA activity in G7 occurred at 24 hr, and activity continued to increase through 7 d. No changes in CA activity were observed in G3. These results indicate that low salinity-stimulated CA induction is under transcriptional regulation, with gene activation occurring first, followed by synthesis of new enzyme. For the step-wise transfers to low salinity, the maximum increase in CA RNA expression (8 fold) was seen at 5 ppt. Interestingly, the largest increase in CA activity occurred at 15 ppt, with no further increase occurring at 5 ppt. These results suggest that the magnitude of salinity change affects both CA expression and activity, but not in exactly the same way, and there may be other regulatory steps involved. Supported by NSF IBN 02-30005 to RPH.