Gill-specific transcriptional regulation of Na+/K+-ATPase -subunit in the euryhaline shore crab Pachygrapsus marmoratus: sequence variants and promoter structure (original) (raw)
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Characterization of transport Na+-ATPases in gills of freshwater tilapia
Fish Physiology and Biochemistry, 1988
Branchial plasma membranes from the freshwater cichlid teleost Oreochromis mossambicus (tilapia) contain two Na 1-dependent ATPases: Na + / K + ATPase, and an amilonde-sensitive ATPase which is postulated to operate as a Na + / H + (-N H 4 +) ATPase. It is suggested that both enzyme activities are located in the basolateral membrane system o f the chloride cells. K + has opposing effects on the two enzymes: it stimu lates Na + / K + ATPase and inhibits Na + / H + (-N H 4 +) ATPase activity. Na + / H + ATPase appears more sensitive to NH4+ at low concentrations than N a +/ K + ATPase and the stimulatory effect by NH4+ ions on the first enzyme could be im portant in facilitating NH4+ excretion by tilapia gills under physiological condi tions. In vitro maximum stimulation by NH4+ is similar for the two enzymes (200%). In contrast to N a + / K + ATPase, Na V H + ATPase activity is inhibited by supra-physiological (> 2 0 mM) concentrations o f NH4+. cells contain the highest levels o f N a +/K + ATPase of all branchial cells. In contrast to suggestions put forward by several authors (reviewed by Payan et al. 1984) that under freshwater conditions lamellar respiratory cells are the main sites of N a + influx, recent evidence seems to reinstate the chloride cells as the sites o f active N a + uptake in freshwater (Avella et al. 1987). To date, the electrochemical gradient o f N a + which is built up via Na + /K + ATPase activity is regarded as the driving force for transepithelial N a + uptake in a variety o f ion translocating epithelia (Skou 1973; Kirschner 1983; Jorgensen 1985), including fish gills (de Renzis and Bornancin 1984). Studies on the localization and biochemical characteristics o f branchial N a +/ K +
2014
The effects of fish weight on salinity tolerance were studied in Caspian salmon (Salmo trutta caspius) parr. 180 fish (all with 2 years old but with three weights; 5, 15, 25g) were selected and they reared in freshwater (FW) and brackish water (BW; 13ppt salinity) for 10 days. The mRNA expression of two α-subunit isoforms of Na + , K + -ATPase (α1a and α1b) and NKCC co-transporters were studied in their gill tissue. In all three weight groups, the mRNA levels for the α1a isoforms decreased following BW exposure, whereas α1b levels significantly increased in 15g and 25g groups. In addition, NKCC gene expression were significantly higher in the groups of BW than FW in 15g and 25g weights (P<0.05). The reciprocal expression of Na + , K + -ATPase isoforms (α1a and α1b) during salinity acclimation suggests that they may have different roles in the gill of FW and BW fishes; ion uptake in FW and ion secretion in BW. In conclusion, in the Caspian salmon, between parrs with the same age, ...
Journal of Experimental Biology, 2004
Freshwater crustaceans, fish and amphibians are continuously challenged to maintain the ionic concentrations of their body fluids within strict physiological limits, while exposed to an extremely dilute medium containing less than 0.2•mmol•l-1 NaCl. The epithelia of hyperosmoregulating crabs exhibit low permeabilities to ions and water, minimizing passive salt loss and osmotic water entry across the body surfaces in general (Mantel and Farmer, 1983). However, to counterbalance urinary salt loss and that from passive diffusion across permeable body surface areas, mainly NaCl is actively taken up from fresh water against a large gradient. While amphibians actively absorb NaCl across the skin, the gills of crustaceans and fish possess specialized cells responsible for active NaCl uptake (Larsen, 1988; Péqueux et al., 1988; Péqueux, 1995). Such cells are characterized by elaborate, highly amplified membrane surfaces, often intimately associated with an elevated number of mitochondria. An increased abundance of mitochondrial F-ATPase provides these cells with their substrate, ATP, which is sufficient to supply the ion-motive ATPases that drive active NaCl absorption. The Na + /K +-ATPase, which often consists of three different subunits, is an essential participant in osmoregulatory NaCl uptake (Therien and Blostein, 2000; Lucu and Towle, 2003). In most epithelia, this transmembrane protein is located in the basolateral membranes where it exchanges three intracellular Na + ions for two extracellular K + ions, generating electrochemical gradients used by secondary active transporters in both the apical and basolateral membranes. In
The Journal of experimental biology, 2001
The vacuolar-type H(+)-ATPase (V-ATPase) has been implicated in osmoregulatory ion uptake across external epithelia of a growing variety of species adapted to life in fresh water. In the present study, we investigated whether the V-ATPase may also function in a euryhaline species that tolerates brackish water (8 salinity) but not fresh water, the shore crab Carcinus maenas. cDNA coding for the regulatory B-subunit of the V-ATPase was amplified and sequenced from C. maenas gills and partially sequenced from four other crab species. Two isoforms differing in the 3'-untranslated region were found in C. maenas. In this species, the abundance of B-subunit mRNA was greater in the respiratory anterior gills than the ion-transporting posterior gills and was not increased by acclimation to dilute salinity. Immunocytochemical analysis showed that the B-subunit protein is not targeted to the apical membrane but is distributed throughout the cytoplasmic compartment. Physiological studies of...
Fish Physiology and Biochemistry, 2014
The present study was conducted to elucidate the osmoregulatory ability of the fish pearl spot (Etroplus suratensis) to know the scope of this species for aquaculture under various salinities. Juvenile pearl spot were divided into three groups and acclimated to freshwater (FW), brackish water (BW) or seawater (SW) for 15 days. The fish exhibited effective salinity tolerance under osmotic challenges. Although the plasma osmolality and Na ? , K ? and Cllevels increased with the increasing salinities, the parameters remained within the physiological range. The muscle water contents were constant among FW-, BW-and SW-acclimated fish. Two Na?/K?-ATPase a-isoforms (NKA a) were expressed in gills during acclimation in FW, BW and SW. Abundance of one isoform was up-regulated in response to seawater acclimation, suggesting its role in ion secretion similar to NKA a1b, while expression of another isoform was simultaneously up-regulated in response to both FW and SW acclimation, suggesting the presence of isoforms switching phenomenon during acclimation to different salinities. Nevertheless, NKA enzyme activities in the gills of the SW and FW individuals were higher (p \ 0.05) than in BW counterparts. Immunohistochemistry revealed that Na ? /K ?-ATPase immunoreactive (NKA-IR) cells were mainly distributed in the interlamellar region of the gill filaments in FW groups and in the apical portion of the filaments in BW and SW groups. The number of NKA-IR cells in the gills of the FW-acclimated fish was almost similar to that of SW individuals, which exceeded that of the BW individuals. The NKA-IR cells of BW and SW were bigger in size than their FW counterparts. Besides, the relative abundance of branchial Na ? /K ? /2Clco-transporter showed stronger evidence in favor of involvement of this protein in hypo-osmoregulation, requiring ion secretion by the chloride cells. To the best of our knowledge, this is the first study reporting the wide salinity tolerance of E. suratensis involving differential activation of ion transporters and thereby suggesting its potential as candidate for fish farming under different external salinities.
Journal of Experimental Biology, 2001
SUMMARYMany studies have shown that hyperosmoregulation in euryhaline crabs is accompanied by enhanced Na++K+-ATPase activity in the posterior gills, but it remains unclear whether the response is due to regulation of pre-existing enzyme or to increased gene transcription and mRNA translation. To address this question, the complete open reading frame and 3′ and 5′ untranslated regions of the mRNA coding for the α-subunit of Na++K+-ATPase from the blue crab Callinectes sapidus were amplified by reverse transcriptase/polymerase chain reaction (RT-PCR) and sequenced. The resulting 3828-nucleotide cDNA encodes a putative 1039-amino-acid protein with a predicted molecular mass of 115.6 kDa. Hydrophobicity analysis of the amino acid sequence indicated eight membrane-spanning regions, in agreement with previously suggested topologies. The α-subunit amino acid sequence is highly conserved among species, with the blue crab sequence showing 81–83 % identity to those of other arthropods and 74...
Journal of Experimental Zoology, 2002
The dynamics of branchial Na+,K+,2Cl− cotransporter (NKCC) and Na+,K+-ATPase (NKA) expression were investigated in brown trout and Atlantic salmon during salinity shifts and the parr-smolt transformation, respectively. In the brown trout, Western blotting revealed that NKCC and NKA abundance increased gradually and in parallel (30- and ten-fold, respectively) after transfer to seawater (SW). The NKA hydrolytic activity increased ten-fold after SW-transfer. Following back-transfer to fresh water (FW), the levels of both proteins and NKA activity decreased. The NKCC immunostaining in the gill of SW-acclimated trout was strong, and mainly localized in large cells in the filament and around the bases of the lamellae. In FW-acclimated trout, immunostaining was less intense and more diffuse. Partial cDNAs of the secretory NKCC1 isoform were cloned and sequenced from both brown trout and Atlantic salmon gills. Two differently sized transcripts were detected by Northern blotting in the gill but not in other osmoregulatory tissues (kidney, pyloric caeca, intestine). The abundance in the gill of these transcripts and of the associated NKCC protein increased four- and 30-fold, respectively, during parr-smolt transformation. The abundance of NKA α-subunit protein also increased in the gill during parr-smolt transformation though to a lesser extent than enzymatic activity (2.5- and eight-fold, respectively). In separate series of in vitro experiments, cortisol directly stimulated the expression of NKCC mRNA in gill tissue of both salmonids. The study demonstrates the coordinated regulation of NKCC and NKA proteins in the gill during salinity shifts and parr-smolt transformation of salmonids. © 2002 Wiley-Liss, Inc.
Physiological and Biochemical Zoology, 2016
The copepod Eurytemora affinis has an unusually broad salinity range, as some populations have recently invaded freshwater habitats independently from their ancestral saline habitats. Prior studies have shown evolutionary shifts in ion transporter activity during freshwater invasions and localization of ion transporters in newly discovered "Crusalis organs" in the swimming legs. The goals of this study were to localize and quantify expression of ion transport enzymes V-type H 1-ATPase (VHA) and Na 1 /K 1-ATPase (NKA) in the swimming legs of E. affinis and determine the degree of involvement of each leg in ionic regulation. We confirmed the presence of two distinct types of ionocytes in the Crusalis organs. Both cell types expressed VHA and NKA, and in the freshwater population the location of VHA and NKA in ionocytes was, respectively, apical and basal. Quantification of in situ expression of NKA and VHA established the predominance of swimming leg pairs 3 and 4 in ion transport in both saline and freshwater populations. Increases in VHA expression in swimming legs 3 and 4 of the freshwater population (in fresh water) relative to the saline population (at 15 PSU) arose from an increase in the abundance of VHA per cell rather than an increase in the number of ionocytes. This result suggests a simple mechanism for increasing ion uptake in fresh water. In contrast, the decline in NKA expression in the freshwater population arose from a decrease in ionocyte area in legs 4, likely resulting from decreases in number or size of ionocytes containing NKA. Such results provide insights into mechanisms of ionic regulation for this species, with added insights into evolutionary mechanisms underlying physiological adaptation during habitat invasions.