Structural architecture of an RNA that competitively inhibits RNase L

Abstract

Activation of RNase L endonuclease activity is part of the mammalian innate immune response to viral infection. The poliovirus RNA genome contains a sequence in its protein-coding region that can act as a competitive inhibitor of RNase L. Mutation, sequence, and functional analysis of this competitive inhibitor RNA (ciRNA) revealed that its activity depends on specific sequences, showed that a loop-loop hairpin interaction forms in the ciRNA, and suggested the presence of a loop E motif. These features lead to the hypothesis that the ciRNA's function is conferred in part by a specific three-dimensional folded RNA architecture. By using a combination of biophysical, mutational, and functional studies, we have mapped features of the threedimensional architecture of the ciRNA in its unbound form. We show that the loop-loop interaction forms in the free ciRNA and affects the overall structure, perhaps forming long-range tertiary interactions with the loop E motif. Local tight RNA-RNA backbone packing occurs in parts of the structure, but the fold appears to be less stable than many other tightly packed RNAs. This feature may allow the ciRNA to accommodate the translocation of ribosomes and polymerase across this multifunctional region of the viral RNA but also to function as an RNase L inhibitor.

Biophysical properties, thermal stability and functional impact of 8-oxo-7,8-dihydroguanine on oligonucleotides of RNA—a study of duplex, hairpins and the aptamer for preQ1as models

“…All thermal denaturation experiments were carried out using CD ( 48 – 50 ) or absorbance [Optical Density, ( 51 , 52 )] by following the bands that can be unambiguously assigned to formation of an A-form duplex with negative ellipticity at 210 nm or positive ellipticity at 270 nm (Figure 2 ). Interestingly, each modification impacted the thermal stability with decreasing T m values of ca.…”

RNA structures that resist degradation by Xrn1 produce a pathogenic Dengue virus RNA

“…One possible explanation of this effect could be that Xrn1 remains bound to xrRNAs, perhaps locking the enzyme in an inactive conformation or sequestering it from other substrates. There is evidence that an RNase L-inhibiting RNA of poliovirus operates as a competitive inhibitor through this type of mechanism ( Keel et al, 2012 ). However, in several of our experiments Xrn1 processes over 150 copies of the xrRNA substrate and over 300 copies of the 24-mer control RNA, indicating multiple turnover by the enzyme.…”

Emergency Services of Viral RNAs: Repair and Remodeling

“…Picornavirus RNAs also have some additional conserved functional structures, such as the hairpin inhibiting the host antiviral enzyme RNase L (154,155) as well as some other potentially biologically relevant elements with poorly characterized functions (156)(157)(158).…”