Hung C Y C, Tsui K N T, Wilson J M, Nawata C M, Wood C M, Wright P A
Department of Integrative Biology, University of Guelph, Guelph, Ontario, N1G 2W1, Canada.
J Exp Biol. 2007 Jul;210(Pt 14):2419-29. doi: 10.1242/jeb.002568.
The mechanism(s) of ammonia excretion in the presence of elevated external ammonia are not well understood in fish. Recent studies in other organisms have revealed a new class of ammonia transporters, Rhesus glycoprotein genes (Rh genes), which may also play a role in ammonia excretion in fish. The first objective of this study was to clone and characterize Rh genes in a fish species with a relatively high tolerance to environmental ammonia, the mangrove killifish Kryptolebias marmoratus (formerly Rivulus marmoratus). We obtained full-length cDNAs of three Rh genes in K. marmoratus: RhBG (1736 bp), RhCG1 (1920 bp) and RhCG2 (2021 bp), which are highly homologous with other known Rh gene sequences. Hydropathy analysis revealed that all three Rh genes encode membrane proteins with 10-12 predicted transmembrane domains. RhBG, RhCG1 and RhCG2 are highly expressed in gill tissue, with RhBG also present in skin of K. marmoratus. Exposure to elevated environmental ammonia (2 mmol l(-1) NH(4)HCO(3)) for 5 days resulted in a modest (+37%) increase in whole-body ammonia levels, whereas gill RhCG2 and skin RhCG1 mRNA levels were upregulated by 5.8- and 7.7-fold, respectively. RhBG mRNA levels were also increased in various tissues, with 3- to 7-fold increases in the liver and skeletal muscle. In a separate group of killifish exposed to air for 24 h, RhCG1 and RhCG2 mRNA levels were elevated by 4- to 6-fold in the skin. Thus, the multifold induction of Rh mRNA levels in excretory tissues (gills and skin) and internal tissues in response to conditions that perturb normal ammonia excretion suggests that RhBG, RhCG1 and RhCG2 may be involved in facilitating ammonia transport in this species. Furthermore, the findings support earlier studies demonstrating that the skin is an important site of ammonia excretion in K. marmoratus.
在外界氨水平升高的情况下，鱼类氨排泄的机制尚未完全明确。近期对其他生物的研究揭示了一类新的氨转运体——恒河猴糖蛋白基因（Rh基因），其可能在鱼类氨排泄中发挥作用。本研究的首要目标是克隆并鉴定红树林鳉鱼（Kryptolebias marmoratus，原称Rivulus marmoratus）中Rh基因的特性，该物种对环境氨具有较高耐受性。我们获得了红树林鳉鱼三个Rh基因的全长cDNA：RhBG（1736 bp）、RhCG1（1920 bp）和RhCG2（2021 bp），它们与其他已知Rh基因序列高度同源。亲水性分析表明，这三个Rh基因均编码具有10 - 12个预测跨膜结构域的膜蛋白。RhBG、RhCG1和RhCG2在鳃组织中高度表达，RhBG在红树林鳉鱼的皮肤中也有表达。暴露于环境氨水平升高（2 mmol l(-1) NH(4)HCO(3)）5天后，鱼体全身氨水平适度升高（+37%），而鳃中RhCG2和皮肤中RhCG1的mRNA水平分别上调了5.8倍和7.7倍。RhBG的mRNA水平在各个组织中也有所增加，肝脏和骨骼肌中增加了3至7倍。在另一组暴露于空气中24小时的鳉鱼中，皮肤中RhCG1和RhCG2的mRNA水平升高了4至6倍。因此，排泄组织（鳃和皮肤）以及内部组织中Rh mRNA水平在干扰正常氨排泄的条件下出现多倍诱导，这表明RhBG、RhCG1和RhCG2可能参与促进该物种的氨转运。此外，这些发现支持了早期的研究，即皮肤是红树林鳉鱼氨排泄的重要部位。
