Zhu Xiao-Jing, Feng Chen-Zhuo, Dai Zhong-Min, Zhang Ruo-Chao, Yang Wei-Jun
Institute of Cell Biology and Genetics, Zhejiang University, 317, College of Life Sciences, Zijingang Campus, Hangzhou, Zhejiang 310058, China.
Stress. 2007 Mar;10(1):53-63. doi: 10.1080/10253890601130773.
AMP-activated protein kinase (AMPK) plays a central role in maintaining the energy balance of organisms under physiological and environmental stresses. Here two AMPK alpha subunit gene transcripts (named Afr-AMPKalpha1 and Afr-AMPKalpha2) from Artemia franciscana were isolated and gene expression was characterized by semiquantitive reverse transcription-polymerase chain reaction (RT-PCR). Afr-AMPKalpha1 was differentially expressed during Artemia developmental stages as well as in response to stresses, such as heat-shock, starvation and a hyperosmotic environment. Afr-AMPKalpha1 mRNA expression in adult Artemia decreased under heat shock, but not in a time- and temperature-dependent manner. By contrast, the transcript sharply decreased in heat-shocked cysts in a time-dependent manner. Under hyperosmotic stress, however, the mRNA level in adults first declined and then increased with prolonged exposure. In the case of starvation, the gene expression in adults decreased and was undetectable after day 9. In addition, Afr-AMPKalpha2 mRNA expression was too low to be detected without nested PCR. Southern blot analysis, moreover, indicated AMPK alpha subunit was present in multiple copies in the Artemia genome. Furthermore, our results demonstrate that the Afr-AMPKalpha1 mRNA level sharply decreases in Artemia carrying diapause-destined embryos and this indicates the possibility that Afr-AMPKalpha1 is involved in determining the reproductive mode in Artemia.
AMP激活的蛋白激酶（AMPK）在维持生物体在生理和环境应激下的能量平衡中起着核心作用。在此，我们从卤虫中分离出两个AMPKα亚基基因转录本（命名为Afr-AMPKα1和Afr-AMPKα2），并通过半定量逆转录-聚合酶链反应（RT-PCR）对基因表达进行了表征。Afr-AMPKα1在卤虫发育阶段以及对热休克、饥饿和高渗环境等应激反应中差异表达。热休克下成年卤虫中Afr-AMPKα1 mRNA表达下降，但不是以时间和温度依赖的方式。相比之下，热休克后的包囊中该转录本以时间依赖的方式急剧下降。然而，在高渗应激下，成年卤虫中的mRNA水平先下降，然后随着暴露时间延长而上升。在饥饿情况下，成年卤虫中的基因表达下降，在第9天后无法检测到。此外，Afr-AMPKα2 mRNA表达太低，不进行巢式PCR就无法检测到。此外，Southern印迹分析表明AMPKα亚基在卤虫基因组中以多拷贝形式存在。此外，我们的结果表明，携带滞育胚胎的卤虫中Afr-AMPKα1 mRNA水平急剧下降，这表明Afr-AMPKα1参与决定卤虫生殖模式的可能性。
