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Structural Insights Reveal the Dynamics of the Repeating r(CAG) Transcript Found in Huntington's Disease (HD) and Spinocerebellar Ataxias (SCAs).

Tawani A, Kumar A - PLoS ONE (2015)

Bottom Line: Moreover, mutant huntingtin protein translated from expanded r(CAG) also yields toxic effects.The overall RNA structure has helical parameters intermediate to the A- and B-forms of nucleic acids due to the global widening of major grooves and base-pair preferences near internal AA loops.The comprehension of structural behaviour by studying the spectral features and the dynamics also supports the flexible nature of the r(CAG) motif.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biosciences and Biomedical Engineering, Indian Institute of Technology Indore, Indore, Madhya Pradesh, India.

ABSTRACT
In humans, neurodegenerative disorders such as Huntington's disease (HD) and many spinocerebellar ataxias (SCAs) have been found to be associated with CAG trinucleotide repeat expansion. An important RNA-mediated mechanism that causes these diseases involves the binding of the splicing regulator protein MBNL1 (Muscleblind-like 1 protein) to expanded r(CAG) repeats. Moreover, mutant huntingtin protein translated from expanded r(CAG) also yields toxic effects. To discern the role of mutant RNA in these diseases, it is essential to gather information about its structure. Detailed insight into the different structures and conformations adopted by these mutant transcripts is vital for developing therapeutics targeting them. Here, we report the crystal structure of an RNA model with a r(CAG) motif, which is complemented by an NMR-based solution structure obtained from restrained Molecular Dynamics (rMD) simulation studies. Crystal structure data of the RNA model resolved at 2.3 Å reveals non-canonical pairing of adenine in 5´-CAG/3´-GAC motif samples in different syn and anti conformations. The overall RNA structure has helical parameters intermediate to the A- and B-forms of nucleic acids due to the global widening of major grooves and base-pair preferences near internal AA loops. The comprehension of structural behaviour by studying the spectral features and the dynamics also supports the flexible nature of the r(CAG) motif.

No MeSH data available.


Related in: MedlinePlus

Temperature dependent 1H NMR spectra for 5´ r(CCGCAGCGG)2 showing imino region and sugar 1´ proton region.The upfield shift of G6H1´ resonance (left), downfield shift of A5H1´ resonance (right) and a rapid downfield shift of C1H1´ resonance (right) is clearly seen.
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pone.0131788.g005: Temperature dependent 1H NMR spectra for 5´ r(CCGCAGCGG)2 showing imino region and sugar 1´ proton region.The upfield shift of G6H1´ resonance (left), downfield shift of A5H1´ resonance (right) and a rapid downfield shift of C1H1´ resonance (right) is clearly seen.

Mentions: To validate the dynamic nature of the non-canonical adenine pairs observed by crystal structure, NMR spectroscopy was used as it allows structural determination in aqueous environments near physiological conditions. One-dimensional proton NMR spectra was recorded for 5´ r(CCGCAGCGG)2 at various temperatures. With an increase in temperature, the broadening of proton resonance peaks in the imino region (G6NH) was observed (Fig 5). The peak broadened with a gradual decay as the temperature was increased from 283 to 293 K, and it eventually disappeared at room temperature. Such a spectrum represents the fast exchange of G6NH with solvent as the imino proton becomes accessible to the solvent due to base flipping, which indicates the dynamic nature of the non-canonical adenine pair (A5) near guanine. The existence of multiple adenine conformations was reinforced by the A5H1´ resonance peak in the sugar 1´ proton region. With an increase in temperature, a slight downfield shift in A5H1´ resonance and upfield shift in G6H1´ resonance further corroborated evidence for the dynamics of adenine pairs. In addition, terminal C1H1´ resonance showed a rapid downfield shift due to the rapid exchange of exposed protons with solvent (Fig 5).


Structural Insights Reveal the Dynamics of the Repeating r(CAG) Transcript Found in Huntington's Disease (HD) and Spinocerebellar Ataxias (SCAs).

Tawani A, Kumar A - PLoS ONE (2015)

Temperature dependent 1H NMR spectra for 5´ r(CCGCAGCGG)2 showing imino region and sugar 1´ proton region.The upfield shift of G6H1´ resonance (left), downfield shift of A5H1´ resonance (right) and a rapid downfield shift of C1H1´ resonance (right) is clearly seen.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493008&req=5

pone.0131788.g005: Temperature dependent 1H NMR spectra for 5´ r(CCGCAGCGG)2 showing imino region and sugar 1´ proton region.The upfield shift of G6H1´ resonance (left), downfield shift of A5H1´ resonance (right) and a rapid downfield shift of C1H1´ resonance (right) is clearly seen.
Mentions: To validate the dynamic nature of the non-canonical adenine pairs observed by crystal structure, NMR spectroscopy was used as it allows structural determination in aqueous environments near physiological conditions. One-dimensional proton NMR spectra was recorded for 5´ r(CCGCAGCGG)2 at various temperatures. With an increase in temperature, the broadening of proton resonance peaks in the imino region (G6NH) was observed (Fig 5). The peak broadened with a gradual decay as the temperature was increased from 283 to 293 K, and it eventually disappeared at room temperature. Such a spectrum represents the fast exchange of G6NH with solvent as the imino proton becomes accessible to the solvent due to base flipping, which indicates the dynamic nature of the non-canonical adenine pair (A5) near guanine. The existence of multiple adenine conformations was reinforced by the A5H1´ resonance peak in the sugar 1´ proton region. With an increase in temperature, a slight downfield shift in A5H1´ resonance and upfield shift in G6H1´ resonance further corroborated evidence for the dynamics of adenine pairs. In addition, terminal C1H1´ resonance showed a rapid downfield shift due to the rapid exchange of exposed protons with solvent (Fig 5).

Bottom Line: Moreover, mutant huntingtin protein translated from expanded r(CAG) also yields toxic effects.The overall RNA structure has helical parameters intermediate to the A- and B-forms of nucleic acids due to the global widening of major grooves and base-pair preferences near internal AA loops.The comprehension of structural behaviour by studying the spectral features and the dynamics also supports the flexible nature of the r(CAG) motif.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biosciences and Biomedical Engineering, Indian Institute of Technology Indore, Indore, Madhya Pradesh, India.

ABSTRACT
In humans, neurodegenerative disorders such as Huntington's disease (HD) and many spinocerebellar ataxias (SCAs) have been found to be associated with CAG trinucleotide repeat expansion. An important RNA-mediated mechanism that causes these diseases involves the binding of the splicing regulator protein MBNL1 (Muscleblind-like 1 protein) to expanded r(CAG) repeats. Moreover, mutant huntingtin protein translated from expanded r(CAG) also yields toxic effects. To discern the role of mutant RNA in these diseases, it is essential to gather information about its structure. Detailed insight into the different structures and conformations adopted by these mutant transcripts is vital for developing therapeutics targeting them. Here, we report the crystal structure of an RNA model with a r(CAG) motif, which is complemented by an NMR-based solution structure obtained from restrained Molecular Dynamics (rMD) simulation studies. Crystal structure data of the RNA model resolved at 2.3 Å reveals non-canonical pairing of adenine in 5´-CAG/3´-GAC motif samples in different syn and anti conformations. The overall RNA structure has helical parameters intermediate to the A- and B-forms of nucleic acids due to the global widening of major grooves and base-pair preferences near internal AA loops. The comprehension of structural behaviour by studying the spectral features and the dynamics also supports the flexible nature of the r(CAG) motif.

No MeSH data available.


Related in: MedlinePlus