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m 1 A and m 1 G Potently Disrupt A-RNA Structure Due to the Intrinsic Instability of Hoogsteen Base Pairs

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ABSTRACT

The B-DNA double helix can dynamically accommodate G–C and A–T base pairs in either Watson-Crick or Hoogsteen configurations. Here, we show that G–C+ and A–U Hoogsteen base pairs are strongly disfavored in A-RNA. As a result, N1-methyl adenosine and N1-methyl guanosine, which occur in DNA as a form of alkylation damage, and in RNA as a posttranscriptional modification, have dramatically different consequences. They create G–C+ and A–U Hoogsteen base pairs in duplex DNA that maintain the structural integrity of the double helix, but block base pairing all together and induce local duplex melting in RNA, providing a mechanism for potently disrupting RNA structure through posttranscriptional modifications. The markedly different propensities to form Hoogsteen base pairs in B-DNA and A-RNA may help meet the opposing requirements of maintaining genome stability on one hand, and dynamically modulating the structure of the epitranscriptome on the other.

No MeSH data available.


Lack of detectable exchange across diverse RNA sequence and structural contexts. (a) Secondary structures with bps showing no detectable RD highlighted in red. (b) Off-resonance RD profiles for the highlighted bps with error bars representing experimental uncertainty (one s.d.) estimated from mono-exponential fitting using a Monte-Carlo based method (Methods).
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Figure 2: Lack of detectable exchange across diverse RNA sequence and structural contexts. (a) Secondary structures with bps showing no detectable RD highlighted in red. (b) Off-resonance RD profiles for the highlighted bps with error bars representing experimental uncertainty (one s.d.) estimated from mono-exponential fitting using a Monte-Carlo based method (Methods).

Mentions: So far, RD studies have provided evidence for µs–ms conformational exchange in non-coding RNAs involving localized changes in secondary structure in and around non-canonical motifs (reviewed in ref. 18). The RD contributions from such chemical exchange processes can mask the ability to detect WC⇄HG exchange. To hone in on WC⇄HG exchange in A-RNA, we carried out 13C and 15N R1ρ RD experiments21,22 on an RNA duplex (hp-A6-RNA) capped by a stabilizing apical loop lacking non-canonical motifs and containing the same sequence (A6-DNA) for which we first reported transient HG bps in B-DNA6 (Fig. 1c and Supplementary Fig. 1a). We targeted purine-C8, C-C6, G-N1, T-N3 and sugar purine-C1′ sites (highlighted in orange in Fig. 1b), all of which have previously been shown to exhibit significant RD due to WC⇄HG chemical exchange in B-DNA6,7,23 (Supplementary Table 1 and Supplementary Note). In stark contrast to B-DNA, all RD profiles measured in in hp-A6-RNA were flat with no signs of detectable conformational exchange on the µs–ms timescale (Fig. 1d). No RD was observed across a variety of rG–rC and rA–rU WC bps, under low pH conditions (pH = 5.4) that allow optimal RD detection of WC⇄HG exchange in B-DNA6,24, upon increasing the temperature (T = 35°C), and in the presence of 4 mM Mg2+ (at pH = 6.8 and T = 5 or 25°C) (Figs. 1d and Fig. 2b, Supplementary Fig. 1b).


m 1 A and m 1 G Potently Disrupt A-RNA Structure Due to the Intrinsic Instability of Hoogsteen Base Pairs
Lack of detectable exchange across diverse RNA sequence and structural contexts. (a) Secondary structures with bps showing no detectable RD highlighted in red. (b) Off-resonance RD profiles for the highlighted bps with error bars representing experimental uncertainty (one s.d.) estimated from mono-exponential fitting using a Monte-Carlo based method (Methods).
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Related In: Results  -  Collection

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Figure 2: Lack of detectable exchange across diverse RNA sequence and structural contexts. (a) Secondary structures with bps showing no detectable RD highlighted in red. (b) Off-resonance RD profiles for the highlighted bps with error bars representing experimental uncertainty (one s.d.) estimated from mono-exponential fitting using a Monte-Carlo based method (Methods).
Mentions: So far, RD studies have provided evidence for µs–ms conformational exchange in non-coding RNAs involving localized changes in secondary structure in and around non-canonical motifs (reviewed in ref. 18). The RD contributions from such chemical exchange processes can mask the ability to detect WC⇄HG exchange. To hone in on WC⇄HG exchange in A-RNA, we carried out 13C and 15N R1ρ RD experiments21,22 on an RNA duplex (hp-A6-RNA) capped by a stabilizing apical loop lacking non-canonical motifs and containing the same sequence (A6-DNA) for which we first reported transient HG bps in B-DNA6 (Fig. 1c and Supplementary Fig. 1a). We targeted purine-C8, C-C6, G-N1, T-N3 and sugar purine-C1′ sites (highlighted in orange in Fig. 1b), all of which have previously been shown to exhibit significant RD due to WC⇄HG chemical exchange in B-DNA6,7,23 (Supplementary Table 1 and Supplementary Note). In stark contrast to B-DNA, all RD profiles measured in in hp-A6-RNA were flat with no signs of detectable conformational exchange on the µs–ms timescale (Fig. 1d). No RD was observed across a variety of rG–rC and rA–rU WC bps, under low pH conditions (pH = 5.4) that allow optimal RD detection of WC⇄HG exchange in B-DNA6,24, upon increasing the temperature (T = 35°C), and in the presence of 4 mM Mg2+ (at pH = 6.8 and T = 5 or 25°C) (Figs. 1d and Fig. 2b, Supplementary Fig. 1b).

View Article: PubMed Central - PubMed

ABSTRACT

The B-DNA double helix can dynamically accommodate G–C and A–T base pairs in either Watson-Crick or Hoogsteen configurations. Here, we show that G–C+ and A–U Hoogsteen base pairs are strongly disfavored in A-RNA. As a result, N1-methyl adenosine and N1-methyl guanosine, which occur in DNA as a form of alkylation damage, and in RNA as a posttranscriptional modification, have dramatically different consequences. They create G–C+ and A–U Hoogsteen base pairs in duplex DNA that maintain the structural integrity of the double helix, but block base pairing all together and induce local duplex melting in RNA, providing a mechanism for potently disrupting RNA structure through posttranscriptional modifications. The markedly different propensities to form Hoogsteen base pairs in B-DNA and A-RNA may help meet the opposing requirements of maintaining genome stability on one hand, and dynamically modulating the structure of the epitranscriptome on the other.

No MeSH data available.