Small molecule quercetin binds MALAT1 triplex and modulates its cellular function

• Published in: Molecular therapy. Nucleic acids 2022-12, Vol.30, p.241-256
• Main Authors: Rakheja, Isha, Ansari, Asgar Hussain, Ray, Arjun, Chandra Joshi, Dheeraj, Maiti, Souvik
• Format: Article
• Language: English
• Subjects: in silico docking; isothermal titration calorimetry; MALAT1; MT: Oligonucleotides: Therapies and Applications; Original; quercetin; RNA FISH; small-molecule binding; triple helix
• ISSN: 2162-2531; 2162-2531
• DOI: 10.1016/j.omtn.2022.09.016

The triple-helix structure at the 3′ end of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), a long non-coding RNA, has been considered to be a target for modulating the oncogenic functions of MALAT1. This study examines the binding of quercetin—a known triplex binding molecule—to the MALAT1 triplex. By employing UV-visible spectroscopy, circular dichroism spectroscopy, and isothermal titration calorimetry, we observed that quercetin binds to the MALAT1 triplex with a stoichiometry of 1:1 and Kd 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 MALAT1 transcript levels in MCF7 cells, and fluorescence in situ hybridization (FISH) experiments showed concomitantly reduced levels of MALAT1 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 MALAT1 downstream genes. Collectively, our study shows that quercetin strongly binds to the MALAT1 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 MALAT1 functions. [Display omitted] The small molecule quercetin and MALAT1 RNA 3′ triplex structure interaction indicates major groove binding. A physical interaction is predicted by in silico docking, biophysics, and a transcription-stop assay. The interaction decreases MALAT1 levels in cellulo, as seen by real-time PCR and RNA FISH. Decreased MALAT1 affects alternative splicing, as demonstrated by RNA sequencing.