人類RCK/p54 (DDX6)同源基因- Zp54參與斑馬魚早期發育之研究

Abstract

RCK/p54 (DDX6)是一種DEAD box 螺旋酶，在miRNA途徑中可和Ago2及其他蛋白質作用形成miRNA-induced silencing complex (miRISC)，抑制mRNA轉譯並幫助Processing body的形成。在酵母菌、線蟲、果蠅和非洲爪蟾上之研究顯示 RCK/p54的同源蛋白參與基因的轉譯抑制作用(translational repression)並影響RNA的穩定性。然而RCK/p54此類蛋白在胚胎發育上的影響仍然不甚清楚，因此本研究利用斑馬魚的Zp54 (人類 RCK/p54同源基因), 探討DDX6在早期發育上扮演的角色。實驗結果顯示，在斑馬魚發育早期時Zp54的mRNA與蛋白質的表現量都很高，但隨著發育的進行快速減少。利用morpholinos抑制zp54的表現時，產生胚胎發育延遲的現象，並造成原腸期的缺陷。同時打入yfp-zp54 mRNA可降低morpholinos所造成的高死亡率和腦部發育異常的現象。實驗結果也顯示Zp54和人類RCK/p54有相似的功能，說明了Zp54在脊索動物上是一個具有高度保留性的基因，在早期胚胎發育中具有重要的功能。

RCK/p54 (DDX6), a RNA helicase of DEAD box family, plays multiplex roles in RNA metabolism. Previous studies of RCK/p54 homologues in yeast, C. elegans, Drosophila and Xenopus showed that RCK/p54 functions as a translational repressor and modulates RNA stability. RCK/p54 also interacts with Ago2 in the miRNA-induced silencing complex (miRISC), represses mRNA translation and facilitates the formation of P-body in human cells. However, the role of RCK/p54 in animal development remains unclear. Here we examined the function of human RCK/p54 homologue, Zp54, in zebrafish embryogenesis. We monitored the mRNA and protein levels of Zp54 in zebrafish embryos. Our results showed that zp54 is highly expressed at early stages and the level is dramatically deceased from one-cell stage to 72hpf. Knockdown of zp54 by injecting zp54 morpholinos results in a severe delay of embryogenesis, and specifically affect on the gastrulation stage. The mortality was decreased and the defects were rescued when zebrafish embryos were co-injected with zp54 morpholinos and yfp-zp54 mRNA. Our data also indicates that the function of Zp54 is similar with human RCK/p54 suggesting that Zp54 is structural and functional conserved in vertebrates and might play important roles during Zebrafish development.