Lam G, Thummel C S
Howard Hughes Medical Institute, Department of Human Genetics, University of Utah, Salt Lake City 84112-5331, USA.
Curr Biol. 2000 Aug 24;10(16):957-63. doi: 10.1016/s0960-9822(00)00631-x.
The introduction of double-stranded RNA (dsRNA) can selectively interfere with gene expression in a wide variety of organisms, providing an ideal approach for functional genomics. Although this method has been used in Drosophila, it has been limited to studies of embryonic gene function. Only inefficient effects have been seen at later stages of development.
When expressed under the control of a heat-inducible promoter, dsRNA interfered efficiently and specifically with gene expression during larval and prepupal development in Drosophila. Expression of dsRNA corresponding to the EcR ecdysone receptor gene generated defects in larval molting and metamorphosis, resulting in animals that failed to pupariate or prepupae that died with defects in larval tissue cell death and adult leg formation. In contrast, expression of dsRNA corresponding to the coding region of the betaFTZ-F1 orphan nuclear receptor had no effect on puparium formation, but led to an arrest of prepupal development, generating more severe lethal phenotypes than those seen with a weak betaFTZ-F1 loss-of-function allele. Animals that expressed either EcR or betaFTZ-F1 dsRNA showed defects in the expression of corresponding target genes, indicating that the observed developmental defects are caused by disruption of the genetic cascades that control the onset of metamorphosis.
These results confirm and extend our understanding of EcR and betaFTZ-F1 function. They also demonstrate that dsRNA expression can inactivate Drosophila gene function at later stages of development, providing a new tool for functional genomic studies in Drosophila.
双链RNA（dsRNA）的引入能够在多种生物体中选择性地干扰基因表达，为功能基因组学提供了一种理想的方法。尽管该方法已在果蝇中使用，但一直局限于胚胎基因功能的研究。在发育后期仅观察到低效的效果。
当在热诱导启动子的控制下表达时，dsRNA在果蝇幼虫和蛹前期发育过程中有效地、特异性地干扰了基因表达。与蜕皮激素受体（EcR）基因对应的dsRNA的表达在幼虫蜕皮和变态过程中产生缺陷，导致动物无法化蛹或蛹前期死亡，伴有幼虫组织细胞死亡和成虫腿部形成缺陷。相比之下，与βFTZ-F1孤儿核受体编码区对应的dsRNA的表达对蛹形成没有影响，但导致蛹前期发育停滞，产生比βFTZ-F1功能丧失弱等位基因所见更严重的致死表型。表达EcR或βFTZ-F1 dsRNA的动物在相应靶基因的表达上出现缺陷，表明观察到的发育缺陷是由控制变态起始的遗传级联反应的破坏引起的。
这些结果证实并扩展了我们对EcR和βFTZ-F1功能的理解。它们还表明dsRNA表达可以在果蝇发育后期使基因功能失活，为果蝇功能基因组学研究提供了一种新工具。
