Rh glycoprotein expression is modulated in pufferfish (Takifugu rubripes) during high environmental ammonia exposure
Rh glycoprotein expression is modulated in pufferfish (Takifugu rubripes) during high environmental ammonia exposure
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DOI：
10.1242/jeb.044719
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发表时间：
2010-09-15
影响因子：
2.8
通讯作者：
Kato, Akira
中科院分区：
文献类型：
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作者：
Nawata, C. Michele;Hirose, Shigehisa;Kato, Akira
恒河（Rh）蛋白在淡水鱼氨转运过程中的参与受到了相当大的关注；然而，对海水物种的类似研究很少。我们将河豚鱼暴露在高环境氨（HEA； 1和5 mmol l(-1) NH4HCO3）中，并评估了氨排泄和鳃Rh mRNA和蛋白表达的模式。同时定量测定鳃H+- atp酶、NHE1、NHE2、NHE3、Na+/K+- atp酶（NKA）、Na+/K+/2Cl（-）共转运体（NKCC1） mRNA、H+- atp酶活性、NKA蛋白及活性。体外证实了NH4+对NKA的激活作用。Rhbg mRNA的下调和Rhcg1、H+-ATPase、NHE3、NKA、NKCC1 mRNA、H+-ATPase活性、NKA蛋白和活性水平的上调表明，HEA过程中氨的排泄主要由NKA驱动的富线粒体细胞（MRCs）介导，NH4+通过NKA和/或NKCC1进入基底侧，NH3通过Rhcg1挤出根尖。NHE3和/或H+- atp酶对NH3的再增殖将使通过Rh通道的反通量最小化。在HEA期间，在鳃中观察到下调的Rhbg和Rhag mRNA，这表明了一种协调的保护性反应，分别通过铺路细胞和柱细胞减少外部氨的流入，同时通过MRCs引导氨排泄。暴露于高碳酸血症（空气中二氧化碳含量为1%）导致鳃和红细胞Rhag mRNA下调。令人惊讶的是，Rhag、Rhbg、Rhcg1和Rhcg2蛋白对高碳酸血症和HEA都有反应，其表观分子质量发生了变化。因此，我们认为河豚Rh蛋白具有双重NH3/CO2转运功能。该结果支持并扩展了早先提出的河豚鱼鳃氨排泄模型，该模型基于Rh蛋白的免疫定位。路面细胞中的被动过程和/或Rhbg和Rhcg2可以维持血浆氨的基础水平，但升高的水平可能需要通过MRCs中的NKA和Rhcg1主动排泄。
Rhesus (Rh) protein involvement in ammonia transport processes in freshwater fish has received considerable attention; however, parallel investigations in seawater species are scant. We exposed pufferfish to high environmental ammonia (HEA; 1 and 5 mmol l(-1) NH4HCO3) and evaluated the patterns of ammonia excretion and gill Rh mRNA and protein expression. Gill H+-ATPase, NHE1, NHE2, NHE3, Na+/K+-ATPase (NKA), Na+/K+/2Cl(-) co-transporter (NKCC1) mRNA, H+-ATPase activity, NKA protein and activity, were also quantified. Activation of NKA by NH4+ was demonstrated in vitro. The downregulation of Rhbg mRNA and simultaneous upregulations of Rhcg1, H+-ATPase, NHE3, NKA, NKCC1 mRNA, H+-ATPase activity, and NKA protein and activity levels suggested that during HEA, ammonia excretion was mediated mainly by mitochondria-rich cells (MRCs) driven by NKA with basolateral NH4+ entry via NKA and/or NKCC1, and apical NH3 extrusion via Rhcg1. Reprotonation of NH3 by NHE3 and/or H+-ATPase would minimise back flux through the Rh channels. Downregulated Rhbg and Rhag mRNA observed in the gill during HEA suggests a coordinated protective response to minimise the influx of external ammonia via the pavement cells and pillar cells, respectively, while routing ammonia excretion through the MRCs. Exposure to hypercapnia (1% CO2 in air) resulted in downregulated gill and erythrocyte Rhag mRNA. Surprisingly, Rhag, Rhbg, Rhcg1 and Rhcg2 proteins responded to both hypercapnia and HEA with changes in their apparent molecular masses. A dual NH3/CO2 transport function of the pufferfish Rh proteins is therefore suggested. The results support and extend an earlier proposed model of pufferfish gill ammonia excretion that was based on immunolocalisation of the Rh proteins. Passive processes and/or Rhbg and Rhcg2 in the pavement cells may maintain basal levels of plasma ammonia but elevated levels may require active excretion via NKA and Rhcg1 in the MRCs.