Zomerdijk J C, Kieft R, Duyndam M, Shiels P G, Borst P
Division of Molecular Biology, The Netherlands Cancer Institute, Amsterdam.
Nucleic Acids Res. 1991 Apr 11;19(7):1359-68. doi: 10.1093/nar/19.7.1359.
African trypanosomes evade the immune response of their host by periodically changing their variant surface glycoprotein (VSG) coat. Each coat is encoded by a separate VSG gene. Expressed genes are in a telomeric expression site (ES) and there are several sites in each trypanosome. To study the transcription control of VSG genes in Trypanosoma brucei we have analyzed an ES, called the dominant ES (DES), that readily switches off and on. The promoter area of the DES is very similar to that of the 221 ES (Zomerdijk et al., 1990). It can be switched off and on in vivo without detectable DNA alterations in the vicinity of the transcription start and it can drive high transient expression of a reporter gene in transfection experiments. However, there are also two major differences between the DES and the 221 ES. First, one version of the DES contains an additional upstream transcription unit overlapping the VSG gene ES promoter. The presence of this upstram transcription is dispensable, however, for the VSG gene ES promoter is active, even if transcription through this start from the upstream promoter is blocked using UV light. Moreover, a second version of the DES present in another trypanosome variant does not produce these upstream transcripts. Secondly, we find that the inactivation of DES transcription in one trypanosome variant is accompanied by DNA alterations in the DES upstream (greater than 2 kb) of the transcription start; reactivation of DES transcription is accompanied by another alteration far upstream. Although we cannot exclude that these DNA rearrangements are incidental, our results raise the possibility that the activity of ES promoters is negatively controlled in cis by far upstream sequences not included in transfection constructs and that alterations in these sequences may lead to (in)activation of the promoter.
非洲锥虫通过周期性改变其可变表面糖蛋白(VSG)外衣来逃避宿主的免疫反应。每一层外衣都由一个单独的VSG基因编码。表达的基因位于端粒表达位点(ES),每个锥虫中有多个这样的位点。为了研究布氏锥虫中VSG基因的转录调控,我们分析了一个称为显性ES(DES)的ES,它很容易开启和关闭。DES的启动子区域与221 ES的启动子区域非常相似(佐默迪克等人,1990年)。它可以在体内开启和关闭,而转录起始附近没有可检测到的DNA改变,并且在转染实验中它可以驱动报告基因的高瞬时表达。然而,DES和221 ES之间也存在两个主要差异。首先,DES的一个版本包含一个额外的上游转录单元,与VSG基因ES启动子重叠。然而,这个上游转录的存在对于VSG基因ES启动子的活性是可有可无的,因为即使使用紫外线阻断从上游启动子通过这个起始点的转录,VSG基因ES启动子仍然是活跃的。此外,另一个锥虫变体中存在的DES的第二个版本不会产生这些上游转录本。其次,我们发现一个锥虫变体中DES转录的失活伴随着转录起始点上游(大于2 kb)的DES中的DNA改变;DES转录的重新激活伴随着更远上游的另一种改变。虽然我们不能排除这些DNA重排是偶然的,但我们的结果提出了一种可能性,即ES启动子的活性受到转染构建体中未包含的远上游序列的顺式负调控,并且这些序列的改变可能导致启动子的(去)激活。