Sun Shengming, Xuan Fujun, Fu Hongtuo, Zhu Jian, Ge Xianping, Wu Xugan
Key Laboratory of Genetic Breeding and Aquaculture Biology of Freshwater Fishes, Ministry of Agriculture, Freshwater Fisheries Research Centre, Chinese Academy of Fishery Sciences, Wuxi 214081, PR China.
Jiangsu Provincial Key Laboratory of Coastal Wetland Bioresources and Environmental Protection, Yancheng City, Jiangsu Province 224002, PR China.
Comp Biochem Physiol A Mol Integr Physiol. 2017 May;207:36-42. doi: 10.1016/j.cbpa.2017.02.010. Epub 2017 Feb 10.
Most hemoglobins are respiratory proteins and are ubiquitous in animals, bacteria, fungi, protists, and plants. In this study, we describe a membrane-bound hemoglobin in the oriental river prawn Macrobrachium nipponense (MnHb), which also expresses hemocyanin. MnHb cDNA was cloned using the rapid amplification of cDNA ends (RACE) approach, which afforded a 1201bp gene encoding a 193 amino acid polypeptide. Bioinformatic evaluation suggested MnHb is membrane anchored by N-myristoylation, and immunofluorescence confirmed its location in the membrane of chief cells in the gill. The effect of hypoxia on MnHb expression was investigated, and reverse transcription PCR (RT-PCR) and Western blotting showed that MnHb was expressed almost exclusively in the gill. Quantitative RT-PCR revealed a significant increase in expression after 6h of hypoxia, and levels peaked at 24h due to oxidative stress. Exposure of cultured prawns to the stress inducer HO significantly up-regulated the expression of MnHb in a dose-dependent manner. MnHb may have a role in protecting cell membrane lipids from damage by reactive oxygen species.
大多数血红蛋白都是呼吸蛋白，在动物、细菌、真菌、原生生物和植物中广泛存在。在本研究中，我们描述了日本沼虾（Macrobrachium nipponense）中的一种膜结合血红蛋白（MnHb），该虾也表达血蓝蛋白。使用cDNA末端快速扩增（RACE）方法克隆了MnHb cDNA，得到一个1201bp的基因，编码一个193个氨基酸的多肽。生物信息学评估表明，MnHb通过N-肉豆蔻酰化锚定在膜上，免疫荧光证实其位于鳃主细胞的膜中。研究了缺氧对MnHb表达的影响，逆转录PCR（RT-PCR）和蛋白质免疫印迹表明MnHb几乎只在鳃中表达。定量RT-PCR显示缺氧6小时后表达显著增加，由于氧化应激，24小时时表达水平达到峰值。将养殖的虾暴露于应激诱导剂HO中，MnHb的表达以剂量依赖的方式显著上调。MnHb可能在保护细胞膜脂质免受活性氧损伤方面发挥作用。
