Activation of an AMP-activated protein kinase is involved in post-diapause development of Artemia franciscana encysted embryos.
Activation of an AMP-activated protein kinase is involved in post-diapause development of Artemia franciscana encysted embryos.
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AMP 激活蛋白激酶的激活参与了卤虫包囊胚胎的滞育后发育
DOI：
10.1186/1471-213x-9-21
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发表时间：
2009-03-16
影响因子：
--
通讯作者：
Yang WJ
中科院分区：
文献类型：
--
作者：
Zhu XJ;Dai JQ;Tan X;Zhao Y;Yang WJ
卤虫的包囊可以保持在休眠状态很长一段时间，代谢率很低，只有在接近有利条件时才能恢复发育。滞育后的发育是一个非常复杂的过程，涉及多种代谢和生化事件。然而，调节这一过程的内在机制尚不清楚。本文报道了卤虫滞育后发育过程中AMP激活蛋白激酶（AMPK）的特异性激活。使用磷酸化AMPK α抗体，发现AMPK在滞育后发育过程中被磷酸化。激酶分析的结果表明，这种磷酸化是必不可少的AMPK激活。使用整装免疫组织化学，磷酸化AMPK被证明是主要位于外胚层的早期发育的胚胎在一个环形，然而，激活区域的位置和形状随着发展的进行而改变。此外，对包囊提取物不同部分的Western印迹分析表明，磷酸化AMPKα定位于细胞核，并且该位置不受细胞内pH的影响。免疫荧光染色包囊细胞核的共聚焦显微镜分析进一步表明，AMPKα在激活时定位于细胞核。此外，细胞内AMP，ADP和ATP水平在发育中的包囊，并通过HPLC测定，结果表明，卤虫AMPK的激活可能不与细胞内AMP：ATP的比例，提示其他途径调节卤虫AMPK活性。总之，我们报告的证据表明激活的AMPK在卤虫发育中的包囊，并提出了一个论点，其作用在某些细胞中的发育相关的基因表达和能量控制在滞育后的卤虫发展。
Cysts of Artemia can remain in a dormant state for long periods with a very low metabolic rate, and only resume their development with the approach of favorable conditions. The post-diapause development is a very complicated process involving a variety of metabolic and biochemical events. However, the intrinsic mechanisms that regulate this process are unclear. Herein we report the specific activation of an AMP-activated protein kinase (AMPK) in the post-diapause developmental process of Artemia. Using a phospho-AMPKα antibody, AMPK was shown to be phosphorylated in the post-diapause developmental process. Results of kinase assay analysis showed that this phosphorylation is essential for AMPK activation. Using whole-mount immunohistochemistry, phosphorylated AMPK was shown to be predominantly located in the ectoderm of the early developed embryos in a ring shape; however, the location and shape of the activation region changed as development proceeded. Additionally, Western blotting analysis on different portions of the cyst extracts showed that phosphorylated AMPKα localized to the nuclei and this location was not affected by intracellular pH. Confocal microscopy analysis of immunofluorescent stained cyst nuclei further showed that AMPKα localized to the nuclei when activated. Moreover, cellular AMP, ADP, and ATP levels in developing cysts were determined by HPLC, and the results showed that the activation of Artemia AMPK may not be associated with cellular AMP:ATP ratios, suggesting other pathways for regulation of Artemia AMPK activity. Together, we report evidence demonstrating the activation of AMPK in Artemia developing cysts and present an argument for its role in the development-related gene expression and energy control in certain cells during post-diapause development of Artemia.