Role of SLC12A10.2, a Na-Cl cotransporter-like protein, in a Cl uptake mechanism in zebrafish (Danio rerio).
The thiazide-sensitive Na(+)-Cl(-) cotransporter (NCC), a member of the SLC12 family, is mainly expressed in the apical membrane of the mammalian distal convoluted tubule (DCT) cells, is responsible for cotransporting Na(+) and Cl(-) from the lumen into DCT cells and plays a major role in the mammalian renal NaCl reabsorption. The NCC has also been reported in fish, but the functional role in fish ion regulation is yet unclear. The present study used zebrafish as an in vivo model to test the hypothesis of whether the NCC plays a role in Na(+) and/or Cl(-) uptake mechanisms. Four NCCs were cloned, and only one of them, zebrafish (z) slc12a10.2 was found to predominately and specifically be expressed in gills. Double in situ hybridization/immunocytochemistry in zebrafish skin/gills demonstrated that the specific expression of zslc12a10.2 mRNA in a novel group of ionocytes differed from those of the previously-reported H(+)-ATPase-rich (HR) cells and Na(+)-K(+)-ATPase-rich (NaR) cells. Gill mRNA expression of zslc12a10.2 was induced by a low-Cl environment that stimulated fish Cl(-) influx, while a low-Na environment suppressed this expression. Incubation with metolazone, a specific inhibitor of the NCC, impaired both Na(+) and Cl(-) influx in 5-day postfertilization (dpf) zebrafish embryos. Translational knockdown of zslc12a10.2 with a specific morpholino caused significant decreases in both Cl(-) influx and Cl(-) content of 5-dpf zebrafish embryos, suggesting that the operation of zNCC-like 2 results in a net uptake of Cl(-) in zebrafish. On the contrary, zslc12a10.2 morphants showed increased Na(+) influx and content that resulted from upregulation of mRNA expressions of Na(+)-H(+) exchanger 3b and carbonic anhydrase 15a in HR cells. These results for the first time provide in vivo molecular physiological evidence for the possible role of the NCC in the Cl(-) uptake mechanism in zebrafish skin/gills.
Wang
Yi-Fang
YF
Institute of Cellular and Organismic Biology, Academia Sinica, Nankang, Taipei, Taiwan 11529, ROC.
Tseng
Yung-Che
YC
Yan
Jia-Jiun
JJ
Hiroi
Junya
J
Hwang
Pung-Pung
PP
D000818:Animals; D002712:Chlorides / Q000378:metabolism / Q000494:pharmacology; D004232:Diuretics / Q000494:pharmacology; D004625:Embryo, Nonmammalian / Q000378:metabolism; D005880:Gills / Q000378:metabolism; D008788:Metolazone / Q000494:pharmacology; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D010802:Phylogeny; D020033:Protein Isoforms / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D012867:Skin / Q000378:metabolism; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology; D049992:Sodium Chloride Symporters / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D064466:Solute Carrier Family 12, Member 1; D015027:Zebrafish / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism
D002712:Chlorides; D004232:Diuretics; D016376:Oligonucleotides, Antisense; D020033:Protein Isoforms; D012333:RNA, Messenger; C541440:SLC12A10.2 protein, zebrafish; D049992:Sodium Chloride Symporters; D064466:Solute Carrier Family 12, Member 1; D029961:Zebrafish Proteins; D012964:Sodium; D006180:Proton-Translocating ATPases; D000254:Sodium-Potassium-Exchanging ATPase; D008788:Metolazone