Talk abstracts
Talk on Friday 01:45-02:00pm submitted by Mika Schievelbein
Global RNA modifications to the MALAT1 triple helix affect thermostability and weaken binding to METTL16
Mika Schievelbein (Department of Chemistry and Biochemistry, University of Notre Dame), Carlos Resende (Department of Chemistry and Biochemistry, University of Notre Dame), Madeline M. Glennon (Department of Chemistry and Biochemistry, University of Notre Dame), Jessica A. Brown (Department of Chemistry and Biochemistry, University of Notre Dame)
Abstract:
Therapeutic mRNAs are generated using modified nucleotides, namely N1-methylpseudouridine (m1Ψ), to minimize/abrogate detection by the immune system. RNA modifications, even at a single nucleotide position, perturb RNA structure, although it is not well understood how structure and function is impacted for global-modified RNAs. Therefore, we examined the MALAT1 triple helix, a highly structured stability element that includes single-, double- and triple-stranded RNA, globally modified with N6-methyladenosine (m6A), pseudouridine (Ψ), or m1Ψ. UV thermal denaturation assays showed that m6A destabilizes both the Hoogsteen and Watson-Crick faces of the RNA by ~20°C, Ψ stabilizes the Hoogsteen and Watson-Crick faces of the RNA by ~12°C, and m1Ψ has minimal effect on the stability of the Hoogsteen face of RNA but increases the stability of the Watson-Crick face by ~9°C. Native gel-shift assays revealed that METTL16 binding to the MALAT1 triple helix was weakened by at least 4-, 12- and 8-fold, respectively, when RNA is globally modified with m6A, Ψ or m1Ψ. These results demonstrate that a more thermostable RNA structure does not lead to tighter RNA-protein interactions, thereby highlighting the regulatory power of RNA modifications at multiple levels.
Keywords: RNA Triple Helix, RNA Modifications, MALAT1