Wellcome Centre for Molecular Parasitology, Glasgow Biomedical Research Centre, University of Glasgow, Glasgow, G12 8TA, UK.
BMC Genomics. 2010 Feb 22;11:124. doi: 10.1186/1471-2164-11-124.
The evolutionarily ancient parasite, Trypanosoma brucei, is unusual in that the majority of its genes are regulated post-transcriptionally, leading to the suggestion that transcript abundance of most genes does not vary significantly between different life cycle stages despite the fact that the parasite undergoes substantial cellular remodelling and metabolic changes throughout its complex life cycle. To investigate this in the clinically relevant sub-species, Trypanosoma brucei gambiense, which is the causative agent of the fatal human disease African sleeping sickness, we have compared the transcriptome of two different life cycle stages, the potentially human-infective bloodstream forms with the non-human-infective procyclic stage using digital gene expression (DGE) analysis.
Over eleven million unique tags were generated, producing expression data for 7360 genes, covering 81% of the genes in the genome. Compared to microarray analysis of the related T. b. brucei parasite, approximately 10 times more genes with a 2.5-fold change in expression levels were detected. The transcriptome analysis revealed the existence of several differentially expressed gene clusters within the genome, indicating that contiguous genes, presumably from the same polycistronic unit, are co-regulated either at the level of transcription or transcript stability.
DGE analysis is extremely sensitive for detecting gene expression differences, revealing firstly that a far greater number of genes are stage-regulated than had previously been identified and secondly and more importantly, this analysis has revealed the existence of several differentially expressed clusters of genes present on what appears to be the same polycistronic units, a phenomenon which had not previously been observed in microarray studies. These differentially regulated clusters of genes are in addition to the previously identified RNA polymerase I polycistronic units of variant surface glycoproteins and procyclin expression sites, which encode the major surface proteins of the parasite. This raises a number of questions regarding the function and regulation of the gene clusters that clearly warrant further study.
古老的寄生虫布氏锥虫的进化非常独特，其大多数基因都是在转录后进行调控的，这表明尽管寄生虫在其复杂的生命周期中经历了大量的细胞重塑和代谢变化，但大多数基因的转录物丰度在不同的生命周期阶段之间并没有显著差异。为了在临床上相关的亚种中研究这一点，即引起致命的人类疾病非洲昏睡病的布氏锥虫冈比亚亚种，我们使用数字基因表达（DGE）分析比较了两种不同生命周期阶段的转录组，即具有潜在人类感染性的血液形式与非人类感染性的前循环阶段。
产生了超过 1100 万个独特的标签，为 7360 个基因生成了表达数据，涵盖了基因组中 81%的基因。与相关的 T. b. brucei 寄生虫的微阵列分析相比，大约检测到了 10 倍以上表达水平变化 2.5 倍的基因。转录组分析显示，基因组内存在几个差异表达的基因簇，表明连续基因，大概来自同一多顺反子单元，在转录或转录稳定性水平上受到共同调控。
DGE 分析非常灵敏，能够检测到基因表达差异，首先揭示了比以前鉴定的更多数量的基因受到阶段调控，其次更重要的是，这种分析揭示了存在几个差异表达的基因簇，这些基因簇似乎位于同一多顺反子单元上，这一现象在微阵列研究中以前没有观察到。这些差异调节的基因簇除了先前鉴定的 RNA 聚合酶 I 多顺反子单元的变体表面糖蛋白和前环表达位点之外，这些基因编码寄生虫的主要表面蛋白。这引发了一些关于基因簇的功能和调节的问题，这些问题显然值得进一步研究。
