Biomedical subjects

Olga Fernández-Miragall

Publications and source records attributed to Olga Fernández-Miragall.

4 recordsLinked to original sources

PubMed2005

Evidence of reciprocal tertiary interactions between conserved motifs involved in organizing RNA structure essential for internal initiation of translation.

Internal ribosome entry site (IRES) elements consist of highly structured RNA regions that determine internal initiation of translation. We have previously shown that the foot-and-mouth disease virus (FMDV) IRES contains a GNRA tetraloop spanning residues G178UAA181. Here we show that tertiary RNA interactions dependent on the GNRA motif determine the structural organization of the central domain. By using mutational analysis in combination with RNA probing, we have identified distant reciprocal interactions between the GNRA motif and the invariant region G240CACG244, termed motif A. Mutations in motif A caused a decrease in IRES activity as severe as the GUAG substitution in the GNRA motif. Substitutions in either GNRA or motif A sequences induced a common reorganization around the conserved R199AAA202 stem-loop, suggesting that the latter contributes to stabilize the GNRA-motif A interaction. This finding was also consistent with a significant increase in the efficiency of RNA-RNA interactions determined in gel shift assays using as probe the hairpin that contains the GNRA motif compared to a transcript encompassing the entire apical region of the central domain. Thus, we propose that the central domain of the FMDV IRES contains a structural conformation essential for IRES activity stabilized by a tertiary contact involving residues in the GNRA tetraloop and motif A conserved sequences.

Base Sequence↗

PubMed2004

Picornavirus IRES: structure function relationship.

Picornavirus infections have been a challenging problem in human health. Genome organisation of picornavirus is unique in having a long, heavily-structured, multifunctional 5'untranslated region, preceding a single open reading frame from which all viral proteins are produced. Within the 5'leader, an internal region termed ribosome entry site (IRES) regulates viral protein synthesis in a 5'-independent manner. The IRES element itself is a distinctive feature of the picornavirus mRNAs, allowing efficient viral protein synthesis in infected cells in spite of a severe modification of translation initiation factors induced by viral proteases that lead to a fast inhibition of cellular protein synthesis. Picornavirus IRES elements are strongly structured, bearing several motifs, phylogenetically conserved, which are essential for IRES activity. Together with RNA structure, RNA-binding proteins play an essential role in the activity of the IRES element, having a profound effect on viral pathogenesis. Recent data on the involvement of these conserved motifs in RNA structure and protein recognition is discussed in detail. Understanding the interplay between these two components of IRES function is crucial to develop viral strategies aimed to use the viral RNA as the target of antiviral approaches.

5' Untranslated Regions↗

PubMed2003

Structural organization of a viral IRES depends on the integrity of the GNRA motif.

Little is known about the tertiary structure of internal ribosome entry site (IRES) elements. The central domain of foot-and-mouth disease (FMDV) IRES, named 3 or I, contains a conserved GNRA motif, essential for IRES activity. We have combined functional analysis with RNA probing to define its structural organization. We have found that a UNCG motif does not functionally substitute the GNRA motif; moreover, binding of synthetic GNRA stem-loops to domain 3 was significantly reduced in RNAs bearing UCCG or GUAG substitutions. The apical region of domain 3 consists of a four-way junction where residues of the GNRA tetraloop are responsible for the organization of the adjacent stem-loops, as deduced from ribonucleases and dimethyl sulfate accessibility. A single A-to-G substitution in the fourth position of this motif led to a strong RNA reorganization, affecting several nucleotides away in the secondary structure of domain 3. The study of mutants bearing UNCG or GUAG tetraloops revealed lack of protection to chemical attack in native RNA at specific nucleotides relative to the parental GUAA, suggesting that the GNRA motif dictates the organization and stability of domain 3. This effect is likely mediated by the interaction with distant residues. Therefore, the GNRA motif plays a crucial role in the organization of IRES structure with important consequences on activity.

Base Sequence↗

PubMed2002

IRES elements: features of the RNA structure contributing to their activity.

The activity of internal ribosome entry site (IRES) elements depends on their structural organization. We have addressed here the study of conserved structural motifs in the foot-and-mouth disease virus (FMDV) IRES as an example to understand the relationship between RNA structure and function. The features of the RNA structure known to be functionally relevant are discussed in regards to the capacity to modulate interaction of translation initiation factors with the FMDV IRES element. Additionally, the contribution of non-canonical RNA-binding proteins to FMDV IRES organization as well as stimulation of its activity by other mRNA regions is discussed.

Base Sequence↗