Poster number 136 submitted by Avleen Chawla
The catalytic activity of H1 circRNA
Avleen K. Chawla (Department of Chemistry, Carnegie Mellon University), Anna M. Kietrys (Department of Chemistry, Carnegie Mellon University)
RPPH1 complex has been long known to cleave pre-tRNA from its 5´-end leading to the formation of its mature form. In human cells, it consists of a catalytic RNA subunit, known as H1 RNA, and various protein subunits, which assist the RNA in folding and interacting with the substrate pre-tRNA. In 2006, H1 RNA was found to be able to independently catalyze this cleavage in vitro under a high Mg2+ concentration. This ribozyme is crucial for proper translation and cell functioning.
In our work, we analyzed a pool of RNAs with altered expression in the human brain. Interestingly, we found that one of overexpressed circular RNAs has the partial sequence of the H1 RNA. We experimentally validated this circRNA and also found a circular RNA having the same sequence as the linear H1 RNA. This circular H1 RNA is expressed among cell lines such as HEK293T, HeLa, and U87MG cells. The predicted structures (using RNAfold) for both the linear and circular isoforms were found to be identical, which suggests catalytic activity of circRPPH1. Circular isoforms have been known to be more stable than linear isoforms. Circular H1 RNA present in cells may have a better catalytic efficiency comparing to the linear H1 RNA. To investigate this, we have synthesized the linear RNA using in vitro transcription and circularized it using T4 Ligase I. Further studies involve studying the catalytic activity of circular RNA in vitro and in vivo, along with the mechanistic and kinetic studies of the reaction. Interactions of this circular RNA will be investigated with the RPPH1 protein subunits to map the function of this H1 circRNA.
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Keywords: circular RNA, ribozymes, H1 RNA