Poster abstracts

Poster number 94 submitted by Yaping Liu

Structural elucidation of the 3’ oncomiR-1 RNA by NMR spectroscopy

Yaping Liu (Biophysics Program), Somaye Badieyan (Department of Biological Chemistry), Michael Cianfrocco (Department of Biological Chemistry), Sarah C. Keane (Biophysics Program; Department of Chemistry)

Abstract:
MicroRNAs are small non-coding RNAs that post-transcriptionally regulate gene expression. To maintain proper microRNA expression levels, the enzymatic processing of primary and precursor microRNA elements must be strictly controlled. However, the molecular determinants underlying this strict regulation of microRNA biogenesis are not fully understood. We are investigating the differential processing of oncomiR-1, a polycistronic primary microRNA that encodes six individual miRNAs (miR-17, miR-18a, miR-19a, miR-20a, miR-19b and miR-20a) and is overexpressed in many cancers.
Chemical probing of oncomiR-1 revealed that the Drosha cleavage sites of pri-miR-92a are sequestered in a four-way junction. NPSL2, an independent stem loop element, is positioned just upstream of pri-miR-92a and sequesters a crucial part of the sequence that constitutes the basal helix of pri-miR-92a. Disruption of the NPSL2 hairpin structure could promote the formation of a pri-miR-92a basal helix that is primed for processing by Drosha. Thus, NPSL2 is predicted to function as a structural switch, regulating pri-miR-92a processing.
Studying the structure of the 3’ domain of oncomiR-1 is crucial for understanding the molecular basis of miR-92a biogenesis. 3’ oncomiR-1 is 234 nts long (~ 77 KDa), and therefore represents a significant technical challenge for structure elucidation by NMR.
By using a divide and conquer strategy along with deuterium editing approach, we were able to complete chemical shift assignments of 3’ oncomiR-1. To tackle the challenge of determining such a large RNA structure, we have also adopted a multidisciplinary approach combing NMR and cryo-EM. The future direction involves refining the 3’ oncomiR-1 structure using these hybrid methods.

References:
Ha, Kim Biogenesis 2014, Ketting et al. Genes and Development 2001, György et al. Science 2001, Hammond et al. Science 2001, Humphreys et al. PNAS 2005, Lim et al. Nature 2005
He et al., Nature 2005, O'Donnell et al. Nature 2005, Chaulk et al., RNA Biol. 2011, Chakraborty et al., RNA 2012, Chakraborty et al., NAR 2017

Keywords: miRNA, oncomiR-1, NMR