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草鱼3个6-磷酸葡萄糖酶催化亚基的基因表达分析及高糖饲料对其表达的影响
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作者:徐晶  梁旭方  蔡文静  高俊杰  旷玉兰  魏君冉  何珊 
单位:华中农业大学水产学院, 华中农业大学鳜鱼研究中心/农业农村部鳜鱼育种创新基地, 农业农村部淡水生物繁育重点实验室, 湖北 武汉 430070
关键词:草鱼 g6pc 糖异生 组织表达 胰岛素 
分类号:S963
出版年·卷·期(页码):2020·27·1(24-34)
摘要:
为了探讨6-磷酸葡萄糖酶催化亚基(glucose-6-phosphatase catalytic subunit,G6PC)在草鱼(Ctenopharyngodon idellus)糖代谢中的作用,采用同源序列比对的方式,在草鱼基因组中获取了3个g6pc基因的序列,通过序列比对和进化树分析,将其分别命名为g6pcag6pcb1g6pcb2,其编码的氨基酸序列与斑马鱼(Danio rerio)、虹鳟(Oncorhynchus mykiss)和人(Homo sapiens)具有较高的同源性,相似度分别为85%~94%、64%~83%和54%~66%。同线性分析表明g6pc在草鱼染色体上的分布与斑马鱼等高度相似,表明草鱼g6pc基因在进化中具有较高的保守性。利用RT-PCR检测3个基因在鳃、脂肪、脑、心脏、肝、肾、前肠、中肠、后肠和肌肉10个组织中的表达,结果显示,g6pca在脑和肝中表达量较高,脂肪组织次之;g6pcb1在肝中表达量最高,中肠次之;g6pcb2在心脏表达量最高,其次为脂肪组织。同时探讨了高糖饲料对草鱼不同g6pc亚型转录水平的影响,以及不同浓度葡萄糖和胰岛素刺激草鱼肝细胞(L8824)后对g6pca转录水平的影响。结果显示,饲喂高糖饲料(7周)后,与对照组相比,草鱼肝脏g6pca mRNA水平显著升高,g6pcb1g6pcb2 mRNA水平无显著变化。离体情况,与5 mmol/L葡萄糖组相比,15 mmol/L葡萄糖显著增加了L8824的g6pca mRNA水平,且1 mol/L胰岛素可以抑制这种作用;30 mmol/L葡萄糖对L8824 g6pca mRNA水平无显著性影响。本研究表明,草鱼g6pc发生加倍后存在功能分化,高糖可以诱导g6pca mRNA的表达,而对g6pcb1g6pcb2的转录水平无影响,其具体功能还需进一步研究。
The glucose-6-phosphatase catalytic subunit (G6PC) is an important component of the glucose-6-phosphatase (G6Pase) system, which controls glucose production from glycogenolysis and gluconeogenesis, and plays a key role in blood glucose homeostasis. G6pc can catalyze the hydrolysis of glucose-6-phosphate to glucose and inorganic phosphate in the living body. Despite its importance, little is known about the function of different g6pc subtypes in the grass carp (Ctenopharyngodon idella). We obtained three g6pc gene sequences from the grass carp genome using homologous sequence alignment, which were named g6pca, g6pcb1, and g6pcb2 through sequence alignment and a phylogenetic tree. Deduced amino acid sequences of g6pc cDNA sequences in grass carp shared similarities of 85%-94%, 64%-83%, and 54%-66% with zebrafish (Danio rerio), rainbow trout (Oncorhynchus mykiss), and human (Homo sapiens), respectively. In addition, synteny analyses of g6pc genes revealed that adjacent genes of grass carp g6pc were identical with that of zebrafish. This indicates that g6pc genes of grass carp were highly conserved in evolution. Grass carp g6pc mRNA expression was detected in different tissues. The results showed that the mRNA expression level of g6pca was highest in the brain, followed by the liver and adipose tissue. Expression of g6pcb1 was most strongly observed in the liver, and weakly observed in the midgut and kidney. The g6pcb2 mRNA level was highest in the heart, followed by adipose tissue. To explore the role of different g6pc subtypes in the glucose metabolism of grass carp, we compared the g6pc mRNA levels in carps with different diets with normal or high level of carbohydrates. The results revealed that a high carbohydrate diet significantly increased the mRNA expression level of g6pca in the liver, while it had no effect on the g6pcb1 and g6pcb2 mRNA levels. Compared with the control group, 15 mmol/L glucose obviously increased the expression of g6pca mRNA in L8824, while 30 mmol/L glucose had no remarkable influence, and insulin at a concentration of 1 mmol/L could overcome the stimulatory effect of 15 mmol/L glucose. Our results suggest that the g6pc genes of grass carp might have functional differentiation after gene duplication events. In addition, high glucose levels induced the expression of g6pca mRNA, but had no significant effect on the transcription levels of g6pcb1 and g6pcb2, the specific functions of which require further study.
该文献标准引用格式:
XU Jing, LIANG Xufang, CAI Wenjing, GAO Junjie, KUANG Yulan, WEI Junran, HE Shan.Gene expression analysis of three 6-phosphoglucose catalytic subunits in grass carp and the effects of a high carbohydrate diet on gene expression[J].Journal of Fishery Sciences of China,2020,27(1):24-34.[徐晶, 梁旭方, 蔡文静, 高俊杰, 旷玉兰, 魏君冉, 何珊.草鱼3个6-磷酸葡萄糖酶催化亚基的基因表达分析及高糖饲料对其表达的影响[J].中国水产科学,2020,27(1):24-34.]
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