Poster abstracts

Poster number 80 submitted by Yizhu Lin

High­throughput probing of human lncRNA secondary structures

Yizhu Lin (Department of Biological Sciences, Carnegie Mellon University), Gemma May (Department of Biological Sciences, Carnegie Mellon University), Joel McManus (Department of Biological Sciences, Carnegie Mellon University)

Abstract:
Long noncoding RNAs (lncRNAs) comprise a large proportion of the human transcriptome. Over the past decade, lncRNAs have been increasingly recognized as important regulators of multiple gene expression processes, and a growing number have been associated with human development and diseases. Different from mRNAs which are coding for proteins, lncRNA structures and their interactions with proteins are believed to be crucial in lncRNA regulatory functions. To experimentally determine the secondary structures of lncRNAs, a recently developed method, Mod-seq, is applied. In Mod-seq, RNAs are first chemical probed by SHAPE reagent, and then undergo high throughput sequencing to determine their secondary structures. Mod-seq provides substantial improvements in throughput than traditional RNA secondary structure determination methods, allowing quick structural probing of the full length lncRNAs that are thousands nucleotides long. We applied Mod-seq method on two in vitro transcribed human lncRNAs, sno-lncRNA and NEAT1. Sno-lncRNA is found to be deleted in an important human disease, Prader-Willi Syndrome (PWS). Sno-lncRNAs are produced from introns with two imbedded snoRNA genes, which leads to transcription of lncRNAs flanked by snoRNA sequences at both 5’ and 3’ ends without 5’ cap and 3’ poly(A) tails. Our Mod-seq probing showed that the 5’ and 3’ snoRNA sequences in sno-lncRNA have similar secondary structural patterns, and the 4 predicted fox2 protein binding sites share a partial single stranded partial double stranded structure motif that resembles internal loop. NEAT1 is another nuclear lncRNA, it serves as the structural scaffold for paraspeckles. Our Mod-seq probing on NEAT1 is aimed to reveal stable secondary structures and conserved secondary structural motifs across species.

References:
Talkish, J., May, G., Lin, Y., Woolford, J. L. & McManus, C. J. Mod­seq: high­throughput sequencing for chemical probing of RNA structure. RNA 20, 713­20 (2014).

Keywords: lncRNA, RNA secondary structure, high-throughput sequencing