Tang C H, Lee T H
Department of Life Sciences, National Chung-Hsing University, Taichung, Taiwan.
J Exp Zool A Ecol Genet Physiol. 2007 Jul 1;307(7):411-8. doi: 10.1002/jez.391.
A novel relationship between branchial carbonic anhydrase II (CAII) and anion exchanger 1 (AE1) was investigated in the euryhaline spotted green pufferfish (Tetraodon nigroviridis). The immunoblots revealed that AE1 was only detected in the membrane fraction of gills while CAII can be probed both in the membrane and cytosol fractions of gills. CAII protein abundance in the membrane fraction is salinity dependent. Immunological detection of the membrane fraction CAII protein in gills showed 3.9-fold higher in the hyposmotic (freshwater) group than the hyperosmotic (seawater;35 per thousand) group. In contrast, there was no change in the protein level of cytosolic CAII between seawater and freshwater groups. The whole-mount immunocytochemical staining demonstrated that both AE1 and CAII were colocalized to the Na(+)/K(+)-ATPase-immunoreactive cells in gill epithelium of the pufferfish. The interaction between CAII and AE1 was further identified by co-immunoprecipitation because AE1 was detected in the immunoprecipitates of CAII and vice versa. Our results showed that in pufferfish gills CAII was not only expressed in the cytosol to produce the substrate for AE1 transport during Cl(-) influx but also associated with the plasma membrane via AE1. Obviously, it is essential for the physiological function of AE1 to interact with CAII in the membrane of gill Na(+)/K(+)-ATPase-immunoreactive cells. To our knowledge, this is the first study to demonstrate the interaction of branchial CAII and AE1 in fish. The novel correlation proposed a new model of Cl(-)/HCO(3) (-) transport in gills of the teleosts.
在广盐性的斑纹绿河豚（黑点叉鼻鲀）中，研究了鳃碳酸酐酶II（CAII）与阴离子交换蛋白1（AE1）之间的新型关系。免疫印迹显示，AE1仅在鳃的膜部分被检测到，而CAII在鳃的膜部分和胞质部分均可被检测到。膜部分中CAII蛋白的丰度取决于盐度。对鳃膜部分CAII蛋白的免疫检测显示，低渗（淡水）组比高渗（海水；35‰）组高3.9倍。相比之下，海水组和淡水组之间胞质CAII的蛋白水平没有变化。整装免疫细胞化学染色表明，AE1和CAII都共定位于河豚鳃上皮中Na(+)/K(+)-ATP酶免疫反应性细胞。通过共免疫沉淀进一步确定了CAII与AE1之间的相互作用，因为在CAII的免疫沉淀物中检测到了AE1，反之亦然。我们的结果表明，在河豚鳃中，CAII不仅在胞质中表达以在Cl(-)内流期间产生AE1转运的底物，而且还通过AE1与质膜相关联。显然，AE1在鳃Na(+)/K(+)-ATP酶免疫反应性细胞膜中与CAII相互作用对其生理功能至关重要。据我们所知，这是第一项证明鱼类鳃中CAII与AE1相互作用的研究。这种新型关联提出了硬骨鱼鳃中Cl(-)/HCO(3) (-)转运的新模型。
