Kwok Chun Kit, Ding Yiliang, Shahid Saima, Assmann Sarah M, Bevilacqua Philip C
*Department of Chemistry, Pennsylvania State University, University Park, PA 16802, U.S.A.
†Center for RNA Molecular Biology, Pennsylvania State University, University Park, PA 16802, U.S.A.
Biochem J. 2015 Apr 1;467(1):91-102. doi: 10.1042/BJ20141063.
Guanine quadruplex structures (GQSs) play important roles in the regulation of gene expression and cellular processes. Recent studies provide strong evidence for the formation and function of DNA and RNA GQSs in human cells. However, whether GQSs form and are functional in plants remains essentially unexplored. On the basis of circular dichroism (CD)-detected titration, UV-detected melting, in-line probing (ILP) and reporter gene assay studies, we report the first example of a plant RNA GQS that inhibits translation. This GQS is located within the 5'-UTR of the ATAXIA TELANGIECTASIA-MUTATED AND RAD3-RELATED (ATR) mRNA of Arabidopsis thaliana (mouse-ear cress). We show that this GQS is highly stable and is thermodynamically favoured over a competing hairpin structure in the 5'-UTR at physiological K⁺ and Mg²⁺ concentrations. Results from ILP reveal the secondary structure of the RNA and support formation of the GQS in vitro in the context of the complete 5'-UTR. Transient reporter gene assays performed in living plants reveal that the GQS inhibits translation but not transcription, implicating this GQS as a translational repressor in vivo. Our results provide the first complete demonstration of the formation and function of a regulatory RNA GQS in plants and open new avenues to explore potential functional roles of GQS in the plant kingdom.
鸟嘌呤四链体结构（GQSs）在基因表达调控和细胞过程中发挥着重要作用。最近的研究为人类细胞中DNA和RNA GQSs的形成及功能提供了有力证据。然而，GQSs在植物中是否形成且具有功能基本上仍未得到探索。基于圆二色性（CD）检测滴定、紫外检测熔解、在线探针（ILP）和报告基因分析研究，我们报道了首个抑制翻译的植物RNA GQS的实例。这个GQS位于拟南芥（鼠耳芥）共济失调毛细血管扩张症突变和Rad3相关（ATR）mRNA的5'-非翻译区（UTR）内。我们表明，在生理K⁺和Mg²⁺浓度下，这个GQS高度稳定，且在热力学上比5'-UTR中与之竞争的发夹结构更具优势。ILP的结果揭示了RNA的二级结构，并支持在完整5'-UTR背景下该GQS在体外的形成。在活体植物中进行的瞬时报告基因分析表明，该GQS抑制翻译但不抑制转录，这意味着该GQS在体内是一种翻译抑制因子。我们的结果首次完整证明了植物中调控性RNA GQS的形成和功能，并为探索GQS在植物界的潜在功能作用开辟了新途径。
