Non-coding RNAs play important roles in cellular homeostasis and are involved in many human diseases including cancer. Intermolecular RNA-RNA interactions are the basis for the diverse functions of many non-coding RNAs. Herein, we show how the presence of tRNA influences the equilibrium between hairpin and G-quadruplex conformations in the 5' untranslated regions of oncogenes and model sequences. Kinetic and equilibrium analyses of the hairpin to G-quadruplex conformational transition of purified RNA as well as during co-transcriptional folding indicate that tRNA significantly shifts the equilibrium toward the hairpin conformer. The enhancement of relative translation efficiency in a reporter gene assay is shown to be due to the tRNA-mediated shift in hairpin-G-quadruplex equilibrium of oncogenic mRNAs. Our findings suggest that tRNA is a possible therapeutic target in diseases in which RNA conformational equilibria is dysregulated.

We determined hybridization rates of DNA probes bound to the surfaces of a human cell, and compared the rates with those determined in solution. The rates were slower on the cell surface than in solution-phase. The position of the nucleation site but not the location of the formed duplex relative to the cell surface adversely affected the hybridization kinetics.

Cotranscriptional folding of an RNA transcript enables formation of metastable RNA structures. Thermodynamic and kinetic properties of RNA G-quadruplex formation have previously been investigated using purified guanine-rich oligonucleotides. Here, we describe a method for analysis of cotranscriptional dynamics of the G-quadruplex formation based on real-time monitoring of the fluorescence of G-quadruplex ligands. For RNA sequences with the potential to form mutually exclusive hairpin or G-quadruplex structures, the efficiency of G-quadruplex formation during transcription depended on position of the hairpin forming sequence. The real-time monitoring enabled evaluation of environmental effects on RNA dynamics, as we demonstrated facilitation of post-transcriptional G-quadruplex formation under molecular crowding conditions. The strategy demonstrated here provides folding insights into the G-quadruplex during transcription that should be involved in gene regulation.

Riboswitch-mediated control of gene expression depends on ligand binding properties (kinetics and affinity) of its aptamer domain. A detailed analysis of interior regions of the aptamer, which affect the ligand binding properties, is important for both understanding natural riboswitch functions and for enabling rational design of tuneable artificial riboswitches. Kinetic analyses of binding reaction between flavin mononucleotide (FMN) and several natural and mutant aptamer domains of FMN-specific riboswitches were performed. The strong dependence of the dissociation rate (52.6-fold) and affinity (100-fold) on the identities of base pairs in the aptamer stem suggested that the stem region, which is conserved in length but variable in base-pair composition and context, is the tuning region of the FMN-specific aptamer. Synthetic riboswitches were constructed based on the same aptamer domain by rationally modifying the tuning regions. The observed 9.31-fold difference in the half-maximal effective concentration (EC50) corresponded to a 11.6-fold difference in the dissociation constant (K(D)) of the aptamer domains and suggested that the gene expression can be controlled by rationally adjusting the tuning regions.

Molecular iodine contributes significantly to carry out organic transformations in synthetic organic chemistry. It works effectively due to its mild Lewis acidic character, ability as an oxidizing agent, good moisture...