•The formation and folding pattern of miR-3620-5p G-quadruplex was investigated.•Sanguinarine had high binding affinity and stabilization effects on the G-quadruplex.•Sanguinarine could block the base-pairing of miR-3620-5p with its target sequence.G-quadruplexes have been reported to exist both in human genome and transcriptome and are of great interests due to their important biological functions. Up to now, the formation and property of G-quadruplex in human mature microRNAs has not been explored yet. In this study, we discovered that hsa-miR-3620-5p, a guanine rich human mature microRNA, could fold into a stable parallel G-quadruplex in near physiological condition for the first time. We explored the formation, folding pattern and binding affinity of the miR-3620-5p G-quadruplex by ESI-MS, CD, NMR and SPR. The results indicated that its high-order structure was comprised of three G-quartets with two bases in each parallel loop stretching outward and two bases flanking at each end. In addition, sanguinarine, a natural alkaloid screened from traditional Chinese medicine was characterized to have high binding affinity and thermodynamic stabilization effects through π–π stacking interaction with the external G-quartets. Furthermore, the potent interaction of sanguinarine with miR-3620-5p G-quadruplex could block the base pairing between miR-3620-5p and its target sequence. Therefore, our study revealed the possibility of regulating microRNA functions using potent G-quadruplex binders, and could provide a new approach to affect the microRNA:mRNA interactions.

A natural flexible cyclic molecule, fangchinoline (FAN) from traditional Chinese medicine, was found with a good binding affinity and thermodynamic stabilization effect on a bcl2 G-quadruplex. Further studies showed that fangchinoline could induce the formation and the conformational conversion of G-quadruplexes from a mixed parallel/anti-parallel to a parallel stranded structure. Our study provided a novel G-quadruplex ligand facilitating the structural conversion of the G-quadruplex, which implied that this fangchinoline could be used to regulate biological activities of the G-quadruplex DNA.

•ESI–MS revealed the formation of three G-quadruplexes from the upstream region of c-Myb transcription start site.•A natural bioactive alkaloid was found to selectively bind with the three c-Myb G-quadruplexes other than long-chain duplex DNA.•HRMS was utilized to investigate the binding competition of dehydroevodiamine in the mixture solution of three G-quadruplexes.In this research, ESI mass spectrometry was used to probe the formation of G-quadruplexes from three G-rich sequences (S1–S3) in the upstream region of the transcription start site in human c-Myb proto-oncogene. A ligand, dehydroevodiamine, was found for its high binding affinities towards the c-Myb G-quadruplexes. In addition, dehydroevodiamine bound towards the Q1–Q3 with high selectivity over long-chain duplex DNA. High-resolution ESI–MS was utilized to investigate the binding competition of dehydroevodiamine in the mixture solution of Q1–Q3 G-quadruplexes, and it appeared to have the binding affinity in the following order: Q3 ≈ Q1 > Q2, which is consistent with the results in the single G-quadruplex solutions. The properties of dehydroevodiamine gave its potential in the future studies of c-Myb expression regulation.Download high-res image (100KB)Download full-size image

There is a putative G-quadruplex sequence (PGS), 5′-G3ACAG4TTG2TG4 TAG4CAG2TTG3-3′, in the promoter region of the PALB2 gene. According to the importance of PALB2 in the process of homologous recombination and breast cancer, and the hypothesis that G-quadruplexes might play a role in transcriptional regulation, we evaluated the formation of G-quadruplex from this complex sequence, as well as its structural information and importance for PALB2 transcription. The G-rich sequence was suggested to form G-quadruplex structure in the NH4OAc solution with 3NH4+ ion by ESI-MS spectrometry, and it was testified in the KCl solution by CD experiments which showed the typical 290 nm band and a high melting temperature. We also classified the guanines in the sequence into four types and demonstrated their roles in the formation of the G-quadruplex structure. In addition, a luciferase assay proved that the formation of the G-quadruplex down-regulate the transcription of the gene. Therefore, our findings provided a foundation for the formation and bioactivity of the G-quadruplex in the promoter region of PALB2 gene and a novel insight for understanding PALB2 gene regulation.

A natural flexible cyclic molecule, fangchinoline (FAN) from traditional Chinese medicine, was found with a good binding affinity and thermodynamic stabilization effect on a bcl2 G-quadruplex. Further studies showed that fangchinoline could induce the formation and the conformational conversion of G-quadruplexes from a mixed parallel/anti-parallel to a parallel stranded structure. Our study provided a novel G-quadruplex ligand facilitating the structural conversion of the G-quadruplex, which implied that this fangchinoline could be used to regulate biological activities of the G-quadruplex DNA.