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Long-range RNA interactions between structural domains of the aphthovirus internal ribosome entry site (IRES)

  • RICARDO RAMOS (a1) and ENCARNACION MARTÍNEZ-SALAS (a1)
    • Published online: 06 September 2001
Abstract

Internal initiation of translation is promoted by internal ribosome entry site (IRES) cis-acting elements. Using transcripts that correspond to the structural domains of the foot-and-mouth disease virus (FMDV) IRES, we have identified RNA–RNA interactions between separated domains (1–2, 3, 4–5, or HH) of the IRES structure. All the assayed domains were able to interact with the full-length IRES as well as with domain 3, although to a different extent, with the most efficient interactions being those occurring between domains 3 and 4–5, and domains 3 and 1–2. RNA–RNA complexes were stable over 1 h of incubation at 37 °C, and depended on Mg2+ and RNA concentration. Neither the antisense domain 1–2 nor tRNA interacted with domain 3, providing experimental evidence of the specificity for the sense strand of the IRES sequence. Additionally, domain 1–2 did not interact with 4–5, leading to the suggestion that domain 3 acts as a scaffold structure where the other domains bind. The thermal disassociation profile of these complexes indicated different strength in these interactions. Whereas 50% of the complexes between domains 3 and 4–5 were destabilized at 45 °C, those formed by domain 1–2 and 3 required temperatures higher than 51 °C. Efficient self-dimerization of domains 3 and 4–5 was found in the absence of other transcripts. Formation of domain 3 homodimer competed with formation of heterocomplexes with other domains, and conversely, domain 3 homodimers were competed out by the presence of the other domains. RNA interactions were also observed at physiological concentrations of Mg2+ and K1+. The identification of the RNA–RNA complexes reported here provide direct experimental evidence of tertiary interactions within IRES elements.

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Corresponding author
Reprint requests to: Encarnacion Martínez-Salas, Centro de Biología Molecular “Severo Ochoa”. CSIC-UAM. Cantoblanco 28049 Madrid, Spain; e-mail: emartinez@cbm.uam.es.
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RNA
  • ISSN: 1355-8382
  • EISSN: 1469-9001
  • URL: /core/journals/rna
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