Rakheja Isha, Ansari Asgar Hussain, Ray Arjun, Chandra Joshi Dheeraj, Maiti Souvik
Chemical and Systems Biology Unit, Council of Scientific and Industrial Research-Institute of Genomics & Integrative Biology, New Delhi 110025, India.
Academy of Scientific & Innovative Research (AcSIR), CSIR-Human Resource Development Centre, Sector 19, Kamla Nehru Nagar, Ghaziabad, Uttar Pradesh 201002, India.
Mol Ther Nucleic Acids. 2022 Sep 23;30:241-256. doi: 10.1016/j.omtn.2022.09.016. eCollection 2022 Dec 13.
The triple-helix structure at the 3' end of metastasis-associated lung adenocarcinoma transcript 1 (), a long non-coding RNA, has been considered to be a target for modulating the oncogenic functions of . This study examines the binding of quercetin-a known triplex binding molecule-to the triplex. By employing UV-visible spectroscopy, circular dichroism spectroscopy, and isothermal titration calorimetry, we observed that quercetin binds to the triplex with a stoichiometry of 1:1 and of 495 ± 61 nM, along with a negative change in free energy, indicating a spontaneous interaction. Employing real-time PCR measurements, we observed around 50% downregulation of transcript levels in MCF7 cells, and fluorescence hybridization (FISH) experiments showed concomitantly reduced levels of in nuclear speckles. This interaction is likely a result of a direct interaction between the molecule and the RNA, as indicated by a transcription-stop experiment. Further, transcriptome-wide analysis of alternative splicing changes induced by quercetin revealed modulation of downstream genes. Collectively, our study shows that quercetin strongly binds to the triplex and modulates its functions. It can thus be used as a scaffold for further development of therapeutics or as a chemical tool to understand functions.
转移相关肺腺癌转录本1（MALAT1）是一种长链非编码RNA，其3'端的三螺旋结构被认为是调节MALAT1致癌功能的靶点。本研究检测了已知的三链结合分子槲皮素与MALAT1三链的结合情况。通过紫外可见光谱、圆二色光谱和等温滴定量热法，我们观察到槲皮素以1:1的化学计量比与MALAT1三链结合，解离常数为495±61 nM，同时自由能发生负变化，表明这是一种自发相互作用。通过实时PCR测量，我们观察到MCF7细胞中MALAT1转录水平下调了约50%，荧光原位杂交（FISH）实验表明核斑点中MALAT1水平也随之降低。转录终止实验表明，这种相互作用可能是该分子与RNA直接相互作用的结果。此外，槲皮素诱导的选择性剪接变化的全转录组分析揭示了MALAT1下游基因的调控情况。总的来说，我们的研究表明槲皮素能强烈结合MALAT1三链并调节其功能。因此，它可作为进一步开发治疗药物的支架，或作为理解MALAT1功能的化学工具。
