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Abstract Upon sensing cytosolic- and/or viral double-stranded (ds)DNA, absent-in-melanoma-2 (AIM2)-like-receptors (ALRs) assemble into filamentous signaling platforms to initiate inflammatory responses. The versatile yet critical roles of ALRs in host innate defense are increasingly appreciated; however, the mechanisms by which AIM2 and its related IFI16 specifically recognize dsDNA over other nucleic acids remain poorly understood (i.e. single-stranded (ss)DNA, dsRNA, ssRNA and DNA:RNA hybrid). Here, we find that although AIM2 can interact with various nucleic acids, it preferentially binds to and assembles filaments faster on dsDNA in a duplex length-dependent manner. Moreover, AIM2 oligomers assembled on nucleic acids other than dsDNA not only display less ordered filamentous structures, but also fail to induce the polymerization of downstream ASC. Likewise, although showing broader nucleic acid selectivity than AIM2, IFI16 binds to and oligomerizes most readily on dsDNA in a duplex length-dependent manner. Nevertheless, IFI16 fails to form filaments on single-stranded nucleic acids and does not accelerate the polymerization of ASC regardless of bound nucleic acids. Together, we reveal that filament assembly is integral to nucleic acid distinction by ALRs.
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Triplex‐Forming Peptide Nucleic Acids with Extended Backbones
Peptide nucleic acid (PNA) forms a triple helix with double‐stranded RNA (dsRNA) stabilized by a hydrogen‐bonding zipper formed by PNA’s backbone amides (N‐H) interacting with RNA phosphate oxygens. This hydrogen‐bonding pattern is enabled by the matching ~5.7 Å spacing (typical for A‐form dsRNA) between PNA’s backbone amides and RNA phosphate oxygens. We hypothesized that extending the PNA’s backbone by one ‐CH2‐ group may bring the distance between PNA amide N‐H closer to 7 Å, favourable for hydrogen‐bonding to the B‐form dsDNA phosphate oxygens. Extension of PNA backbone was expected to selectively stabilize PNA‐DNA triplexes compared to PNA‐RNA. To test this hypothesis, we synthesized triplex‐forming PNAs that had the pseudopeptide backbones extended by an additional ‐CH2‐ group in three different positions. Isothermal titration calorimetry measurements of the binding affinity of these extended PNA analogues for the matched dsDNA and dsRNA showed that, contrary to our structural reasoning, extending the PNA backbone at any position had a strong negative effect on triplex stability. Our results suggest that PNA may have an inherent preference for A‐form‐like conformations when binding double‐stranded nucleic acids. It appears that the original six atoms long PNA backbone is an almost perfect fit for binding to A‐form nucleic acids.
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- Award ID(s):
- 1708761
- PAR ID:
- 10175281
- Date Published:
- Journal Name:
- ChemBioChem
- ISSN:
- 1439-4227
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript
- Journal Article:
- https://doi.org/10.1002/cbic.202000432
