Abstract 1685 A Chemical Biology Approach to Investigate Alternative Structures in Cancer Relevant Precursor MicroRNAs

Abnormal expression of microRNAs (miRNAs), small non-coding RNAs involved in gene regulation, has been linked in the development and progression of various cancers. Our understanding of miRNA biogenesis and processing relies on RNA recognition and cleavage by a series of dedicated enzymes to produce the mature miRNA. Mature miRNAs then go on to target partially complementary mRNAs, regulating their expression. A critical step in maturation involves Dicer-mediated processing of a precursor miRNA (pre-miRNA) by recognition of features within its stem-loop structure and cleavage to produce the miRNA duplex. However, recent reports showed that some pre-miRNAs can form an alternative G-quadruplex (G4) motif, creating an equilibrium between the canonical and noncanonical structure. Formation of the G4 inhibits Dicer-mediated recognition and cleavage and thereby reduces mature miRNA expression. These alternative structures may present a new avenue to not only understand natural regulation of miRNA maturation but may also provide a new mechanism to manipulate miRNA formation and function. To investigate the biological role of these G4s in pre-miRNA processing, we used a small molecule microarray screen to identify molecules that bind specifically to either the G4 or stem-loop structure in target pre-miRNAs with identified G4 motifs. Selective compounds were analyzed for their ability to bind to the G4 or stem-loop and to modify the structural equilibrium in the target pre-miRNA via biochemical and biophysical techniques. We are further examining how small molecule induced changes in pre-miRNA structure can alter miRNA maturation by examining Dicer-mediated cleavage in vitro. These RNA-binding probes will be useful tools in exploring this structure-based mechanism for pre-miRNA regulation, particularly in cancers where these miRNAs are irregularly expressed.