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首页> 外文期刊>The Journal of Experimental Biology >Evidence for an apical Na-Cl cotransporter involved in ion uptake in a teleost fish.
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Evidence for an apical Na-Cl cotransporter involved in ion uptake in a teleost fish.

机译：在硬骨鱼中涉及离子吸收的根尖Na-Cl共转运蛋白的证据。

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摘要

Cation-chloride cotransporters, such as the Na(+)/K(+)/2Cl(-) cotransporter (NKCC) and Na(+)/Cl(-) cotransporter (NCC), are localized to the apical or basolateral plasma membranes of epithelial cells and are involved in active ion absorption or secretion. The objectives of this study were to clone and identify ;freshwater-type' and ;seawater-type' cation-chloride cotransporters of euryhaline Mozambique tilapia (Oreochromis mossambicus) and to determine their intracellular localization patterns within mitochondria-rich cells (MRCs). From tilapia gills, we cloned four full-length cDNAs homologous to human cation-chloride cotransporters and designated them as tilapia NKCC1a, NKCC1b, NKCC2 and NCC. Out of the four candidates, the mRNA encoding NKCC1a was highly expressed in the yolk-sac membrane and gills (sites of the MRC localization) of seawater-acclimatized fish, whereas the mRNA encoding NCC was exclusively expressed in the yolk-sac membrane and gills of freshwater-acclimatized fish. We then generated antibodies specific for tilapia NKCC1a and NCC and conducted whole-mount immunofluorescence staining for NKCC1a and NCC, together with Na(+)/K(+)-ATPase, cystic fibrosis transmembrane conductance regulator (CFTR) and Na(+)/H(+) exchanger 3 (NHE3), on the yolk-sac membrane of tilapia embryos acclimatized to freshwater or seawater. The simultaneous quintuple-color immunofluorescence staining allowed us to classify MRCs clearly into four types: types I, II, III and IV. The NKCC1a immunoreactivity was localized to the basolateral membrane of seawater-specific type-IV MRCs, whereas the NCC immunoreactivity was restricted to the apical membrane of freshwater-specific type-II MRCs. Taking account of these data at the level of both mRNA and protein, we deduce that NKCC1a is the seawater-type cotransporter involved in ion secretion by type-IV MRCs and that NCC is the freshwater-type cotransporter involved in ion absorption by type-II MRCs. We propose a novel ion-uptake model by MRCs in freshwater that incorporates apically located NCC. We also reevaluate a traditional ion-uptake model incorporating NHE3; the mRNA was highly expressed in freshwater, and the immunoreactivity was found at the apical membrane of other freshwater-specific MRCs.
机译：阳离子氯化物共转运蛋白，例如Na（+）/ K（+）/ 2Cl（-）共转运蛋白（NKCC）和Na（+）/ Cl（-）共转运蛋白（NCC），位于顶或基底外侧质膜上上皮细胞的表达，并参与活性离子的吸收或分泌。本研究的目的是克隆和鉴定莫桑比克罗非鱼（Oreochromis mossambicus）的淡水型和海水型阳离子-氯化物共转运蛋白，并确定它们在富含线粒体的细胞（MRCs）中的细胞内定位模式。从罗非鱼ia中，我们克隆了四个与人类阳离子-氯化物共转运蛋白同源的全长cDNA，并将其命名为罗非鱼NKCC1a，NKCC1b，NKCC2和NCC。在这四种候选物中，编码NKCC1a的mRNA在海水适应的鱼的卵黄囊膜和g（MRC定位位点）中高表达，而编码NCC的mRNA仅在卵黄囊膜和g中表达。淡水适应鱼类。然后，我们生成了对罗非鱼NKCC1a和NCC特异的抗体，并对NKCC1a和NCC进行了整装免疫荧光染色，以及Na（+）/ K（+）-ATPase，囊性纤维化跨膜电导调节剂（CFTR）和Na（+）/ H（+）交换子3（NHE3），在罗非鱼胚胎的卵黄囊膜上适应了淡水或海水。同时进行的五色免疫荧光染色使我们可以将MRC清楚地分为四种类型：I，II，III和IV型。 NKCC1a免疫反应性局限于海水特异性IV型MRCs的基底外侧膜，而NCC免疫反应性限于淡水特异性II型MRCs的顶膜。考虑到mRNA和蛋白质水平的这些数据，我们推论NKCC1a是参与IV型MRC离子分泌的海水型共转运蛋白，而NCC是参与II型离子吸收的淡水型共转运蛋白。 MRC。我们提出了由MRCs在淡水中结合根尖定位的NCC的新型离子吸收模型。我们还重新评估了包含NHE3的传统离子吸收模型。 mRNA在淡水中高表达，并且在其他淡水特异性MRC的顶膜上发现了免疫反应性。

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