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Redistribution of immunofluorescence of CFTR anion channel and NKCC cotransporter in chloride cells during adaptation of the killifish Fundulus heteroclitus to sea water
Department of Biology, St Francis Xavier University, Antigonish, NS, Canada B2G 2W5
* Author for correspondence (e-mail: bmarshal{at}stfx.ca )
Accepted 22 February 2002
Cellular distribution of cystic fibrosis transmembrane conductance regulator (CFTR) immunofluorescence was detected by monoclonal antibody directed to the C terminus of killifish CFTR (kfCFTR) in chloride cells of fresh water (FW) adapted fish and animals transferred to sea water (SW) for 24h, 48h and 14+ days. Confocal microscopy allowed localization within mitochondria-rich (MR) cells to be determined as superficial (i.e. in the apical membrane) or deeper within the cytoplasm of the cells. In FW, 90 % of MR cells had diffuse kfCFTR immunofluorescence in the central part of the cytosol, with only 8.1 % having apical kfCFTR, which was 6.6±0.54 µm below the microridges of surrounding pavement cells. Curiously, FW but not SW pavement cells also had positive immunofluorescence to kfCFTR. After 24h in SW, a time when kfCFTR expression is elevated, a condensed punctate immunofluorescence appeared among 18.8 % of MR cells, 13.4±0.66 µm (mean ± S.E.M.) below the surface of the cells. By 48h, a majority (76.3 %) of MR cells had punctate kfCFTR distribution and the distance from the surface was less (7.8±0.2 µm), a distribution approaching the SW-acclimated condition (i.e. all MR cells showing kfCFTR immunofluorescence, 6.1±0.04 µm below the surface). In contrast, NKCC immunofluorescence was condensed and localized in lateral parts of MR cell complexes in FW animals and then redistributed to the whole basal cytoplasm after acclimation to SW. CFTR, the anion channel responsible for Cl- secretion in marine teleosts, redistributes in MR cells during SW acclimation by condensation of a diffuse distribution below the apical crypt, followed by translocation and insertion in the apical membrane. NKCC, the cotransporter that translocates Cl- across the basolateral membrane, moves from an eccentric cytosolic location in FW to a diffuse basolateral localization in SW chloride cells.
Key words: epithelial ion transport, cystic fibrosis transmembrane conductanctance regulator, protein trafficking, Na+,K+,2Cl- cotransporter, osmoregulation, euryhaline teleost, killifish, Fundulus heteroclitus
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