Transcriptional regulation of two stage-specifically expressed genes in the protozoan parasite Toxoplasma gondii
Transcriptional regulation of two stage-specifically expressed genes in the protozoan parasite Toxoplasma gondii
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DOI：
10.1093/nar/gki314
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发表时间：
2005-01-01
影响因子：
14.9
通讯作者：
Tomavo, S
中科院分区：
文献类型：
--
作者：
Kibe, MK;Coppin, A;Tomavo, S
原生动物寄生虫弓形虫差异表达两种不同的烯醇酶同工酶，分别称为 ENO1 和 ENO2。为了了解速殖子向缓殖子转化过程中的差异基因表达，我们对两个刚地弓形虫烯醇化酶启动子进行了表征。这些序列之间没有发现同源性，并且没有明显的 TATA 或 CCAAT 盒。通过对与氯霉素乙酰转移酶报告基因融合的 5'-侧翼区域进行缺失分析，然后进行瞬时转染，研究了 ENO1 和 ENO2 启动子在速殖子向缓殖子分化过程中的差异激活。我们的数据表明，在增殖的速殖子中，ENO1的抑制涉及启动子中的负远端调节区（核苷酸-1245至-625），而ENO2启动子中的近端调节区指导低水平的表达。相反，ENO1 的启动子活性在速殖子转化为静止缓殖子后被高度诱导。缓殖子中的ENO2启动子分析表明有两个上游抑制位点（核苷酸-1929至-1067和-456至-222）。此外，电泳迁移率变动分析表明，速殖子和缓殖子核裂解物中存在 DNA 结合蛋白，这些蛋白与 ENO1 和 ENO2 上游调控区的应激反应元件 (STRE)、热休克样元件 (HSE) 和其他顺式调控元件结合。 AGGGG 共有序列的突变完全消除了蛋白质与含有该元件的寡核苷酸的结合。这项研究首次定义了参与重要病原体弓形虫基因调控的顺式调控元件和推定转录因子的特征。
The protozoan parasite Toxoplasma gondii differentially expresses two distinct enolase isoenzymes known as ENO1 and ENO2, respectively. To understand differential gene expression during tachyzoite to bradyzoite conversion, we have characterized the two T. gondii enolase promoters. No homology could be found between these sequences and no TATA or CCAAT boxes were evident. The differential activation of the ENO1 and ENO2 promoters during tachyzoite to bradyzoite differentiation was investigated by deletion analysis of 5 '-flanking regions fused to the chloramphenicol acetyltransferase reporter followed by transient transfection. Our data indicate that in proliferating tachyzoites, the repression of ENO1 involves a negative distal regulatory region (nucleotides -1245 to -625) in the promoter whereas a proximal regulatory region in the ENO2 promoter directs expression at a low level. In contrast, the promoter activity of ENO1 is highly induced following the conversion of tachyzoites into resting bradyzoites. The ENO2 promoter analysis in bradyzoites showed that there are two upstream repression sites (nucleotides -1929 to -1067 and -456 to -222). Furthermore, electrophoresis mobility shift assays demonstrated the presence of DNA-binding proteins in tachyzoite and bradyzoite nuclear lysates that bound to stress response elements (STRE), heat shock-like elements (HSE) and other cis-regulatory elements in the upstream regulatory regions of ENO1 and ENO2. Mutation of the consensus AGGGG sequence, completely abolished protein binding to an oligonucleotide containing this element. This study defines the first characterization of cis-regulatory elements and putative transcription factors involved in gene regulation of the important pathogen T. gondii.