Transposition of the Tourist-MITE mPing in yeast: an assay that retains key features of catalysis by the class 2 PIF/Harbinger superfamily.
Transposition of the Tourist-MITE mPing in yeast: an assay that retains key features of catalysis by the class 2 PIF/Harbinger superfamily.
复制标题
DOI：
10.1186/1759-8753-1-5
复制
发表时间：
2010-02-01
期刊：
影响因子：
4.9
通讯作者：
Wessler SR
中科院分区：
文献类型：
--
作者：
Hancock CN;Zhang F;Wessler SR
PIF/Harbinger是最近发现的DNA转座子超家族，现在已知其在从真菌到植物再到动物的基因组中分布。超家族成员的动员需要两个独立的元件编码蛋白（ORF 1和TfR）。该超家族的成员还动员了类逆转录酶的微型反向重复转座因子（MITEs），其是与植物，特别是禾本科植物的基因相关的最丰富的转座因子。许多植物基因组的系统发育分析表明，MITE可以从一个或几个元件迅速扩增到数百个或数千个元件。迄今为止在植物或动物中鉴定出的最活跃的DNA转座子是mPing，一种类似水稻蚜虫的MITE，是自主Ping元件的缺失衍生物。Ping和与之密切相关的Pong是唯一已知的自然活跃的PIF/Harbinger元素。一些水稻品系每代每株植物积累约40个新mPing插入。在这项研究中，我们报告的酵母转座试验的发展作为第一步，在破译的基础上的放大mist-MITEs的机制。由Ping和Pong编码的ORF 1和Tclase蛋白已被证明在水稻和转基因拟南芥中动员mPing。在酵母测定中对天然蛋白质的初始测试导致非常低的转座。显著更高的活动，获得了一个假定的核输出信号（内斯）的突变，增加了细胞核中的T细胞的数量。当引入拟南芥中时，内斯突变蛋白还催化从gfp报告基因切除mPing的频率更高。我们的酵母测定保留了mPing的切除和插入的关键特征，包括精确切除、延伸的插入序列偏好以及对可以来自Ping或Pong或两种元件的两种蛋白质的要求。酵母转座试验为分析PIF/Harbinger元件的两种蛋白质催化转座的机制提供了一个强大的平台。它概括了在植物系统中看到的mPing的切除和重新插入的所有特征。此外，在Tclase中一个假定的内斯的突变增加了酵母和植物中的转座。
PIF/Harbinger is the most recently discovered DNA transposon superfamily and is now known to populate genomes from fungi to plants to animals. Mobilization of superfamily members requires two separate element-encoded proteins (ORF1 and TPase). Members of this superfamily also mobilize Tourist-like miniature inverted repeat transposable elements (MITEs), which are the most abundant transposable elements associated with the genes of plants, especially the cereal grasses. The phylogenetic analysis of many plant genomes indicates that MITEs can amplify rapidly from one or a few elements to hundreds or thousands. The most active DNA transposon identified to date in plants or animals is mPing, a rice Tourist-like MITE that is a deletion derivative of the autonomous Ping element. Ping and the closely related Pong are the only known naturally active PIF/Harbinger elements. Some rice strains accumulate ~40 new mPing insertions per plant per generation. In this study we report the development of a yeast transposition assay as a first step in deciphering the mechanism underlying the amplification of Tourist-MITEs. The ORF1 and TPase proteins encoded by Ping and Pong have been shown to mobilize mPing in rice and in transgenic Arabidopsis. Initial tests of the native proteins in a yeast assay resulted in very low transposition. Significantly higher activities were obtained by mutation of a putative nuclear export signal (NES) in the TPase that increased the amount of TPase in the nucleus. When introduced into Arabidopsis, the NES mutant protein also catalyzed higher frequencies of mPing excision from the gfp reporter gene. Our yeast assay retains key features of excision and insertion of mPing including precise excision, extended insertion sequence preference, and a requirement for two proteins that can come from either Ping or Pong or both elements. The yeast transposition assay provides a robust platform for analysis of the mechanism underlying transposition catalyzed by the two proteins of PIF/Harbinger elements. It recapitulates all of the features of excision and reinsertion of mPing as seen in plant systems. Furthermore, a mutation of a putative NES in the TPase increased transposition both in yeast and plants.
登录
查看更多内容
影响因子：
11.6
作者：
Jiang, N;Wessler, SR
通讯作者：
Wessler, SR
影响因子：
56.9
作者：
SUTTON, WD;GERLACH, WL;PEACOCK, WJ
通讯作者：
PEACOCK, WJ
影响因子：
1.5
作者：
Kapitonov, VV;Jurka, J
通讯作者：
Jurka, J
影响因子：
3.9
作者：
Izsvák, Z;Ivics, Z;Hackett, PB
通讯作者：
Hackett, PB
DOI：
10.1073/pnas.0605421103
发表时间：
2006-11-21
影响因子：
11.1
作者：
Naito, Ken;Cho, Eunyoung;Wessler, Susan R.
通讯作者：
Wessler, Susan R.