厚壳贻贝鸟氨酸-尿素循环中的关键代谢物及基因分析

鸟氨酸-尿素循环(OUC)是生物新陈代谢过程中的重要循环过程,但在贝类中尚缺乏相关研究。为此,以厚壳贻贝为研究对象,分别采用氨基酸分析仪和荧光定量PCR研究了其外套膜和后闭壳肌组织中的鸟氨酸-尿素循环途径的主要代谢物和关键基因的含量及其表达量;进一步测试了在精氨酸注射条件下,各主要代谢物和关键基因的含量及表达量变化,以及¹³C标记尿素注射贻贝后,其贝壳中δ¹³C比值(¹³C/¹²C)变化。结果表明,厚壳贻贝外套膜和后闭壳肌均含有较高浓度的尿素;精氨酸注射导致其两种组织中尿素浓度显著上升(P<0.01),以及瓜氨酸浓度显著下降(P<0.01),但鸟氨酸浓度维持相对稳定的水平。精氨酸注射显著上调了两种组织中的脲酶基因的表达量(P<0.01),但其他基因表达量的变化在两种组织中存在差异,显示出鸟氨酸-尿素循环途径在其两种组织中具有复杂而不同的调控过程。¹³C标记尿素注射贻贝显著上调了贝壳中δ¹³C的比值(P <0.01),表明尿素分子可能参与了贻贝贝壳的生物矿化过程。上述研究为深入了解贻贝鸟氨酸-尿素途径与生物矿化之间的关联,以及探讨贻贝对海水酸化耐受性的内在分子机制奠定了基础。

Analysis of the Key Metabolites and Genes Involving in Ornithine-Urea Cycle in Mytilus coruscus

The ornithine-urea cycle (OUC) plays important roles in metabolism. However, there is a lack of study of OUC in shellfish. For filling this gap, the mussel Mytilus coruscus was selected, and the key genes together with the metabolites of OUC pathway were analyzed in the mantle and adductor muscle, respectively, using a real-time fluorescent quantitative PCR and an amino-acid analyzer. Moreover, the changes of the metabolite concentrations and the gene relative expression level of OUC were analyzed after arginine injection. The δ¹³C (¹³C/¹²C) of the mussel shell was calculated by an Isotope cube after injection of urea with ¹³C stable isotope labeling. The results showed that both of the mantle and the adductor muscle contain relative high concentration of urea, and arginine injection significantly up-regulated the urea concentration (P<0.01) and down-regulated the citrulline concentration (P<0.01). The other metabolites, such as ornithine, were observed with remaining stability. In addition, the arginine injection increased significantly the relative expression level of urease gene in both mantle and adductor muscle (P<0.01). However, the expression levels of other genes were different in two tissues, indicating that the ornithine-urea cycle pathway has complex and different regulatory processes in these two tissues. Moreover, injection of ¹³C labeled urea significantly increased the ratio of δ¹³C in the mussel shells (P <0.01), suggesting that urea may be involved in the biomineralization process of mussel. These studies provided clues for further understanding of the relationship between the OUC pathway and biomineralization of mussel, as well as exploring the molecular mechanism of the tolerance of Mytilus to seawater acidification.