海七鳃鳗pma-miR-200 c-3 p对斑马鱼心脏发育的作用研究

摘要

The pma-miR-200 c-3 p mimics from sea lamprey Petromyzon marinus were injected into zebrafish Da-nio rerio embryos by microinjection, and qRT-PCR and Western blot were used to analyze the marker gene expres-sion of the development of heart, and the expression levels of several genes in BMP/Smad pathway to study the bio-logical function of pma-miR-200c-3p of the sea lamprey. Besides, target gene bio-informatics and GFP fluores-cence expression analysis were used to study the function of pma-miR-200c-3p. The results showed that the gene expression levels of some cardiac development markers including bmp4 , bmp5 , smad1 , gata5 , tbx2 b and notch1 a were significantly increased in the pma-miR-200 c-3 p-injected embryos ( P<0 . 01 ) . The protein expression levels of the BMP4, Smad1 and GATA5 were significantly increased bya BMP/Smad pathway, and it appears that cdx1b is a target gene of pma-miR-200c-3p. Therefore, the findings indicate that pma-miR-200c-3p might repress the gene expression of cdx1b, and be involved in the BMP/Smadpathway to promote the cardiac development process.%为研究海七鳃鳗Petromyzon marinus pma-miR-200c-3p在心脏发育过程中的生物学功能，采用显微注射技术，将pma-miR-200c-3p模拟体注射到斑马鱼Danio rerio胚胎，过表达pma-miR-200c-3p后，使用qRT-PCR及Western blot法检测了一些与心脏发育相关的标志性基因的表达水平，以及BMP/Smad通路重要基因蛋白水平的变化，并利用生物信息学预测、 GFP荧光表达分析了pma-miR-200c-3p的生物学作用。结果表明：过表达pma-miR-200c-3p的斑马鱼胚胎中，与心脏发育相关的标志性bmp4、 bmp5、 smad1、gata5、 tbx2b和notch1a基因的表达水平极显著上调(P<0.01)，同时BMP/Smad通路中BMP4、 Smad1、 GA-TA5蛋白表达水平也明显升高；细胞试验表明， cdx1b基因可能是pma-miR-200c-3p的一个靶基因。研究表明， pma-miR-200c-3p可能通过抑制cdx1b基因的表达，参与BMP/Smad通路以促进心脏发育。