Structure of Propeller-Type Parallel-Stranded RNA G-Quadruplexes, Formed by Human Telomeric RNA Sequences in K+ Solution (original) (raw)
Related papers
Structure of Human Telomeric RNA (TERRA): Stacking of Two G-Quadruplex Blocks in K+ Solution
Biochemistry, 2013
Telomeric repeat-containing RNAs (TERRA) are transcription products of the telomeres. Human TERRA sequences containing UUAGGG repeats can form parallelstranded G-quadruplexes. The stacking interaction of such structures was shown to be important for ligand targeting and higher-order arrangement of G-quadruplexes in long TERRA sequences. Here we report on the first high-resolution structure of a stacked G-quadruplex formed by the 10nucleotide human TERRA sequence r(GGGUUAGGGU) in potassium solution. This structure comprises two dimeric three-layer parallel-stranded G-quadruplex blocks, which stack on each other at their 5′-ends. The adenine in each UUA loop is nearly coplanar with the 5′-end G-tetrad forming an A•(G•G•G•G)•A hexad, thereby increasing the stacking contacts between the two blocks. Interestingly, this stacking and loop conformation is different from all structures previously reported for the free human TERRA but resembles the structure previously determined for a complex between a human TERRA sequence and an acridine ligand. This stacking conformation is a potential target for drugs that recognize or induce the stacking interface.
Biochemistry, 2014
We investigate the structure formed by the first 18-nt of the 5′-terminal region of the human telomerase RNA (hTERC or hTR) using gel electrophoresis and UV, CD, and NMR spectroscopy. Our data suggest that this 18-nt sequence, r(GGGUUGCGGAGGGUGGGC), can form a stacked dimeric G-quadruplex in potassium solution. The two subunits, each being a three-layer parallel-stranded G-quadruplex with a cytosine bulge, are stacked at their 5′-end. The formation of this stacked dimeric Gquadruplex containing bulges could be biologically relevant for the dimerization and other interactions of the active human telomerase.
Nucleic Acids Research, 2017
Human telomeric RNA has been identified as a key component of the telomere machinery. Recently, the growing evidence suggests that the telomeric RNA forms G-quadruplex structures to play an important role in telomere protection and regulation. In the present studies, we developed a 19 F NMR spectroscopy method to investigate the telomeric RNA G-quadruplex structures in vitro and in living cells. We demonstrated that the simplicity and sensitivity of 19 F NMR approach can be used to directly observe the dimeric and two-subunits stacked Gquadruplexes in vitro and in living cells and quantitatively characterize the thermodynamic properties of the G-quadruplexes. By employing the 19 F NMR in living cell experiment, we confirmed for the first time that the higher-order G-quadruplex exists in cells. We further demonstrated that telomere RNA G-quadruplexes are converted to the higher-order Gquadruplex under molecular crowding condition, a cell-like environment. We also show that the higherorder G-quadruplex has high thermal stability in crowded solutions. The finding provides new insight into the structural behavior of telomere RNA G-quadruplex in living cells. These results open new avenues for the investigation of G-quadruplex structures in vitro and in living cells.
Journal of the American Chemical Society, 2010
We report on the self-assembled structures formed by 12-mer, 22-mer, and 45-mer telomeric RNA (telRNA/TERRA) sequences compared to their DNA analogues, as studied by electrospray mass spectrometry, circular dichroism, and thermal denaturation. The major difference between telomeric RNA and DNA sequences is the ability of telomeric RNA to form higher-order dimeric assemblies, initiated by cation-mediated stacking of two parallel G-quadruplex subunits. The 5'-5' stacking had been observed recently by NMR for the r(GGGUUAGGGU) 10-mer (Martadinata, H.; Phan, A. T. J. Am. Chem. Soc. 2009, 131, 2570); the present work shows that stacking also occurs for the 22-mer containing four G-tracts and for the 45-mer containing eight G-tracts, suggesting a general structural feature of telomeric RNA. The importance of kinetic effects in multimer formation, unfolding, and structural rearrangements is also highlighted.
Biochemistry, 2011
b S Supporting Information G uanine-rich nucleic acids can form a four-stranded structure called the G-quadruplex, based on the stacking of multiple G 3 G 3 G 3 G tetrads. 1À3 Biological roles of G-quadruplex structures have been discussed for G-rich DNA in various genomic locations, 4 such as telomeres, 5 oncogenic promoters, 6,7 and immunoglobulin switches, 4 as well as for G-rich RNA at 5 0-untranslated regions of several oncogenes. 8 Telomeres, which are located at the chromosomal ends, act as protective caps that prevent chromosome loss and degradation. Telomeres had always been thought to be transcriptionally silent until the recent finding that they could be transcribed into RNA molecules with lengths ranging from 100 to 9000 nt. 9,10 It has further been shown that telomeric-repeat-containing RNA (TERRA) perform various cellular regulatory functions, such as regulation of telomere length, 10À12 inhibition of telomerase, 13 telomeric heterochromatin formation, 14,15 and telomere protection. 9,16À18 Similar to the DNA counterparts, short G-rich human telomeric RNA sequences were shown to form G-quadruplex structures. 19À29 It has been suggested that the structures of long human telomeric DNA and RNA sequences are based on multiple G-quadruplex blocks, each formed by a four-repeat segment. 20À22,29À34 So far, high-resolution structures could only be obtained for RNA sequences containing up to two human telomeric repeats. 21,25 Here, we address the question on the arrangement of G-quadruplexes in long human TERRA sequences (of up to 96 repeats) by performing ribonuclease protection assays and molecular dynamics (MD) simulations. We found that G-quadruplexes formed by four and eight human telomeric UUAGGG repeats are abundant and stable structural units that may serve as building blocks in long human TERRA, and loopÀloop interactions may contribute to the stability of higher-order assemblies. ' MATERIALS AND METHODS Construction of Plasmids and DNA Templates. The PRST5 plasmid 35 containing 96 human telomeric repeats (Figure S1, Supporting Information) was a gracious gift from Jack D. Griffith's laboratory. Another plasmid containing 9 human telomeric repeats (Figure S1, Supporting Information) was constructed according to previously published protocol 35 with some modifications. A DNA template containing nine human telomeric repeats 5 0-CCCGGATCCGTCTCA(CCCTAA) 9 GTCTTCAAGCTT-CCC-3 0 was used for PCR reaction. Oligonucletide containing BbsI and HindIII restriction sites 5 0-GGGAAGCTTGAAGACTT-3 0 was used as the forward primer. Another oligonucleotide containing BsmBI and BamHI restrictions sites 5 0-CCCGGATC-CGTCTCACC-3 0 was used as the reverse primer. All the oligonucleotides were synthesized in house on an ABI 394 DNA
Journal of Biological Chemistry, 2012
Background: Telomeric repeat-containing RNA has recently been found in mammalian cells. Results: Oligonucleotide models of telomeric DNA and RNA form a hybrid G-quadruplex structure. Conclusion: We suggest a model system for understanding the structure and function of human telomeres. Significance: Our finding provides valuable information for understanding the structure and function of human telomeric DNA and RNA. Telomeric repeat-containing RNA, a non-coding RNA molecule, has recently been found in mammalian cells. The detailed structural features and functions of the telomeric RNA at human chromosome ends remain unclear, although this RNA molecule may be a key component of the telomere machinery. In this study, using model human telomeric DNA and RNA sequences, we demonstrated that human telomeric RNA and DNA oligonucleotides form a DNA-RNA G-quadruplex. We next employed chemistry-based oligonucleotide probes to mimic the naturally formed telomeric DNA-RNA G-quadruplexes in living cells, suggesting that the process of DNA-RNA G-quadruplex formation with oligonucleotide models of telomeric DNA and RNA could occur in cells. Furthermore, we investigated the possible roles of this DNA-RNA G-quadruplex. The formation of the DNA-RNA G-quadruplex causes a significant increase in the clonogenic capacity of cells and has an effect on inhibition of cellular senescence. Here, we have used a model system to provide evidence about the formation of G-quadruplex structures involving telomeric DNA and RNA sequences that have the potential to provide a protective capping structure for telomere ends.
A crystallographic and modelling study of a human telomeric RNA (TERRA) quadruplex
Nucleic Acids Research, 2010
DNA telomeric repeats in mammalian cells are transcribed to guanine-rich RNA sequences, which adopt parallel-stranded G-quadruplexes with a propeller-like fold. The successful crystallization and structure analysis of a bimolecular human telomeric RNA G-quadruplex, folded into the same crystalline environment as an equivalent DNA oligonucleotide sequence, is reported here. The structural basis of the increased stability of RNA telomeric quadruplexes over DNA ones and their preference for parallel topologies is described here. Our findings suggest that the 2 0 -OH hydroxyl groups in the RNA quadruplex play a significant role in redefining hydration structure in the grooves and the hydrogen bonding networks. The preference for specific nucleotides to populate the C3 0 -endo sugar pucker domain is accommodated by alterations in the phosphate backbone, which leads to greater stability through enhanced hydrogen bonding networks. Molecular dynamics simulations on the DNA and RNA quadruplexes are consistent with these findings. The computations, based on the native crystal structure, provide an explanation for RNA G-quadruplex ligand binding selectivity for a group of naphthalene diimide ligands as compared to the DNA G-quadruplex.
Antiparallel RNA G-quadruplex Formed by Human Telomere RNA Containing 8-Bromoguanosine
Scientific reports, 2017
In this study, by combining nuclear magnetic resonance (NMR), circular dichroism (CD), liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS), and gel electrophoresis, we report an unusual topological structure of the RNA G-quadruplex motif formed by human telomere RNA r(UAGGGU) containing 8-bromoguanosine. Results showed that the RNA sequence formed an antiparallel tetramolecular G-quadruplex, in which each pair of diagonal strands run in opposite directions. Furthermore, guanosines were observed both in syn- and anti-conformations. In addition, two of these G-quadruplex subunits were found to be stacking on top of each other, forming a dimeric RNA G-quadruplex. Our findings provide a new insight into the behavior of RNA G-quadruplex structures.