Subnuclear localization of the active variant surface glycoprotein gene expression site in Trypanosoma brucei
Subnuclear localization of the active variant surface glycoprotein gene expression site in Trypanosoma brucei
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DOI：
10.1073/pnas.95.21.12328
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发表时间：
1998-10-13
影响因子：
11.1
通讯作者：
Borst, P
中科院分区：
文献类型：
--
作者：
Chaves, I;Zomerdijk, J;Borst, P
在布氏锥虫中，RNA聚合酶II的转录和信使RNA的5'加帽是解偶联的：加帽的剪接前导序列被反式剪接到每个RNA上。这种解耦使得由RNA聚合酶I驱动的蛋白质编码基因转录成为可能。事实上，间接证据表明，血液形式的锥虫中的主要表面糖蛋白、变体表面糖蛋白（VSG）的基因由RNA聚合酶I转录。在单个锥虫中，在任何一个时间只有一个CSG表达位点被最大程度地转录，并且已经推测转录发生在细胞核内的独特位点，可能在核仁中。我们用荧光原位杂交检测了这一点。用覆盖约50 kb的活性221表达位点的探针，我们在单个荧光点中检测到该位点的核转录物，其不与核仁共定位。标记基因标记的活性表达位点DNA的荧光DNA原位杂交分析证实了与核仁的关联的情况下。即使启动子被rDNA启动子取代的活性表达位点也不与核仁共定位。正如预期的那样，插入rDNA阵列的标记基因主要与核仁共定位，而微管蛋白基因阵列则不。我们的结论是，转录的活性VSG表达位点不发生在核仁。
In Trypanosoma brucei, transcription by RNA polymerase II and 5' capping of messenger RNA are uncoupled: a capped spliced leader is trans spliced to every RNA. This decoupling makes it possible to have protein-coding gene transcription driven by RNA polymerase I. Indeed, indirect evidence suggests that the genes for the major surface glycoproteins, variant surface glycoproteins (VSGs) in bloodstream-form trypanosomes, are transcribed by RNA polymerase I. In a single trypanosome, only one CSG expression site is maximally transcribed at any one time, and it has been speculated that transcription takes place at a unique site within the nucleus, perhaps in the nucleolus. We tested this by using fluorescence in situ hybridization. With probes that cover about 50 kb of the active 221 expression site, we detected nuclear transcripts of this site in a single fluorescent spot, which did not colocalize with the nucleolus. Analysis of marker gene-tagged active expression site DNA by fluorescent DNA in situ hybridization confirmed the absence of association with the nucleolus. Even an active expression site in which the promoter had been replaced by an rDNA promoter did not colocalize with the nulceolus, As expected, marker genes inserted in the rDNA array predominantly colocalize with the nucleolus, whereas the tubulin gene arrays do not. We conclude that transcription of the active VSG expression site does not take place in the nucleolus.