Smargiasso Nicolas, Gabelica Valérie, Damblon Christian, Rosu Frédéric, De Pauw Edwin, Teulade-Fichou Marie-Paule, Rowe J Alexandra, Claessens Antoine
Mass Spectrometry Laboratory, GIGA-Research, University of Liege, Liege, Belgium.
BMC Genomics. 2009 Aug 6;10:362. doi: 10.1186/1471-2164-10-362.
Guanine-rich nucleic acid sequences are capable of folding into an intramolecular four-stranded structure called a G-quadruplex. When found in gene promoter regions, G-quadruplexes can downregulate gene expression, possibly by blocking the transcriptional machinery. Here we have used a genome-wide bioinformatic approach to identify Putative G-Quadruplex Sequences (PQS) in the Plasmodium falciparum genome, along with biophysical techniques to examine the physiological stability of P. falciparum PQS in vitro.
We identified 63 PQS in the non-telomeric regions of the P. falciparum clone 3D7. Interestingly, 16 of these PQS occurred in the upstream region of a subset of the P. falciparum var genes (group B var genes). The var gene family encodes PfEMP1, the parasite's major variant antigen and adhesin expressed at the surface of infected erythrocytes, that plays a key role in malaria pathogenesis and immune evasion. The ability of the PQS found in the upstream regions of group B var genes (UpsB-Q) to form stable G-quadruplex structures in vitro was confirmed using 1H NMR, circular dichroism, UV spectroscopy, and thermal denaturation experiments. Moreover, the synthetic compound BOQ1 that shows a higher affinity for DNA forming quadruplex rather than duplex structures was found to bind with high affinity to the UpsB-Q.
This is the first demonstration of non-telomeric PQS in the genome of P. falciparum that form stable G-quadruplexes under physiological conditions in vitro. These results allow the generation of a novel hypothesis that the G-quadruplex sequences in the upstream regions of var genes have the potential to play a role in the transcriptional control of this major virulence-associated multi-gene family.
富含鸟嘌呤的核酸序列能够折叠成一种称为G-四链体的分子内四链结构。当在基因启动子区域发现时，G-四链体可能通过阻断转录机制来下调基因表达。在这里，我们使用全基因组生物信息学方法在恶性疟原虫基因组中鉴定推定的G-四链体序列（PQS），并结合生物物理技术在体外检测恶性疟原虫PQS的生理稳定性。
我们在恶性疟原虫克隆3D7的非端粒区域鉴定出63个PQS。有趣的是，其中16个PQS出现在恶性疟原虫var基因子集（B组var基因）的上游区域。var基因家族编码PfEMP1，这是寄生虫的主要变体抗原和粘附素，在感染红细胞表面表达，在疟疾发病机制和免疫逃避中起关键作用。使用1H NMR、圆二色性、紫外光谱和热变性实验证实了在B组var基因上游区域发现的PQS（UpsB-Q）在体外形成稳定G-四链体结构的能力。此外，发现对形成四链体而非双链体结构的DNA具有更高亲和力的合成化合物BOQ1与UpsB-Q具有高亲和力结合。
这是首次证明恶性疟原虫基因组中的非端粒PQS在体外生理条件下形成稳定的G-四链体。这些结果提出了一个新假设，即var基因上游区域中的G-四链体序列有可能在这个主要的毒力相关多基因家族的转录控制中发挥作用。
