Abstract
The telomere repeats containing RNA (TERRA) is transcribed from the C-rich strand of telomere DNA and comprises of UUAGGG nucleotides repeats in humans. The TERRA RNA repeats can exist in single stranded, RNA-DNA hybrid and G-quadruplex forms in the cell. Interaction of TERRA RNA with hnRNPA1 has been proposed to play critical roles in maintenance of telomere DNA. hnRNPA1 contains an N-terminal UP1 domain followed by an RGG-box containing C-terminal region. RGG-motifs are emerging as key protein motifs that recognize the higher order nucleic acid structures as well as are known to promote liquid-liquid phase separation of proteins. In this study, we have shown that the RGG-box of hnRNPA1 specifically recognizes the TERRA RNA G-quadruplexes that have loops in their topology, whereas it does not interact with the single-stranded RNA. Our results show that the N-terminal UP1 domain in the presence of the RGG-box destabilizes the loop containing TERRA RNA G-quadruplex efficiently compared to the RNA G-quadruplex that lacks loops, suggesting that unfolding of G-quadruplex structures by UP1 is structure dependent. Furthermore, we have compared the telomere DNA and TERRA RNA G-quadruplex binding by the RGG-box of hnRNPA1 and discussed its implications in telomere DNA maintenance.
Highlights
The thermodynamic stabilities of TERRA-6, TERRA-12, and TERRA-24 RNA G-quadruplexes were monitored using CD thermal melting experiments in the presence of 100 mM KCl
The CD spectra of the G-quadruplexes formed by the TERRA-6, TERRA-12 and TERRA-24 RNA sequences were acquired in physiologically relevant 100 mM KCl containing buffer [3]
Dimeric TERRA-12 G-quadruplex exhibited a Tm of 63.4°C, whereas the tetrameric TERRA-6 G-quadruplex exhibited a Tm of 72.5°C
Summary
The thermodynamic stabilities of TERRA-6, TERRA-12, and TERRA-24 RNA G-quadruplexes were monitored using CD thermal melting experiments in the presence of 100 mM KCl. The CD spectra of the G-quadruplexes formed by the TERRA-6, TERRA-12 and TERRA-24 RNA sequences were acquired in physiologically relevant 100 mM KCl containing buffer [3]. TERRA-6 sequence forms a tetrameric, parallel G-quadruplex without loops in its structure. TERRA-12 sequence adopts a dimeric, parallel G-quadruplex structure with two loops and TERRA-24 sequence adopts a monomeric, intramolecular, parallel G-quadruplex structure with three loops [1, 2].
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