Analysis of the interaction between human RITA and Drosophila Suppressor of Hairless
Analysis of the interaction between human RITA and Drosophila Suppressor of Hairless
复制标题
DOI：
10.1111/hrd2.00074
复制
发表时间：
2014-12-01
期刊：
影响因子：
2.7
通讯作者：
Maier, Dieter
中科院分区：
文献类型：
--
作者：
Brockmann, Birgit;Mastel, Helena;Maier, Dieter
Notch信号传导介导细胞间通讯，其受转录因子CSL影响，CSL是脊椎动物CBF 1/RBP-J、无毛果蝇抑制因子[Su（H）]和C. elegans Lag1. CBF 1/RBP-J的核输入依赖于共激活子和共阻遏子，而输出依赖于RITA。RITA是微管蛋白和CBF 1/RBP-J结合蛋白，在脊椎动物中作为Notch信号传导的负调节剂。RITA蛋白在果蝇中是高度保守的，但尚未发现果蝇同源物。在这项工作中，解决了人类RITA在苍蝇中的活动。为此，我们产生了转基因苍蝇，允许组织特异性诱导的人RITA，这是证明了蛋白质印迹和苍蝇组织。出乎意料的是，在果蝇发育过程中RITA的过表达几乎没有表型后果，即使与Su（H）或Notch拮抗剂Hairless同时过表达。我们通过共免疫沉淀证明了人RITA与Su（H）和微管蛋白的体内结合。此外，RITA和微管蛋白共定位在某种程度上在几个果蝇组织。总的来说，我们的数据表明，人RITA，虽然结合果蝇Su（H）和微管蛋白，不能影响果蝇中的Notch信号通路，这表明Su（H）的核输出机制，如果存在于果蝇中，不依赖于RITA。
Notch signalling mediates intercellular communication, which is effected by the transcription factor CSL, an acronym for vertebrate CBF1/RBP-J, Drosophila Suppressor of Hairless [Su(H)] and C. elegans Lag1. Nuclear import of CBF1/RBP-J depends on co-activators and co-repressors, whereas the export relies on RITA. RITA is a tubulin and CBF1/RBP-J binding protein acting as a negative regulator of Notch signalling in vertebrates. RITA protein is highly conserved in eumatazoa, but no Drosophila homologue was yet identified. In this work, the activity of human RITA in the fly was addressed. To this end, we generated transgenic flies that allow a tissue specific induction of human RITA, which was demonstrated by Western blotting and in fly tissues. Unexpectedly, overexpression of RITA during fly development had little phenotypic consequences, even when overexpressed simultaneously with either Su(H) or the Notch antagonist Hairless. We demonstrate the in vivo binding of human RITA to Su(H) and to tubulin by co-immune precipitation. Moreover, RITA and tubulin co-localized to some degree in several Drosophila tissues. Overall our data show that human RITA, albeit binding to Drosophila Su(H) and tubulin, cannot influence the Notch signalling pathway in the fly, suggesting that a nuclear export mechanism of Su(H), if existent in Drosophila, does not depend on RITA.