A Hybrid G-quadruplex Structure Formed Between RNA and DNA Explains the Extraordinary Stability of the Mitochondrial R-loop

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 2012 Sep 12

PMID 22965135

Citations 93

Authors

Paulina H Wanrooij

Jay P Uhler

Yonghong Shi

Fredrik Westerlund

Maria Falkenberg

Claes M Gustafsson

Affiliations

Soon will be listed here.

Abstract

In human mitochondria the transcription machinery generates the RNA primers needed for initiation of DNA replication. A critical feature of the leading-strand origin of mitochondrial DNA replication is a CG-rich element denoted conserved sequence block II (CSB II). During transcription of CSB II, a G-quadruplex structure forms in the nascent RNA, which stimulates transcription termination and primer formation. Previous studies have shown that the newly synthesized primers form a stable and persistent RNA-DNA hybrid, a R-loop, near the leading-strand origin of DNA replication. We here demonstrate that the unusual behavior of the RNA primer is explained by the formation of a stable G-quadruplex structure, involving the CSB II region in both the nascent RNA and the non-template DNA strand. Based on our data, we suggest that G-quadruplex formation between nascent RNA and the non-template DNA strand may be a regulated event, which decides the fate of RNA primers and ultimately the rate of initiation of DNA synthesis in human mitochondria.

Citing Articles

Mechanisms underlining R-loop biology and implications for human disease.

Liu J, Li F, Cao Y, Lv Y, Lei K, Tu Z Front Cell Dev Biol. 2025; 13:1537731.

PMID: 40061014 PMC: 11885306. DOI: 10.3389/fcell.2025.1537731.

Decoding the regulatory roles of circular RNAs in cardiac fibrosis.

You Q, Yu J, Pan R, Feng J, Guo H, Liu B Noncoding RNA Res. 2025; 11:115-130.

PMID: 39759175 PMC: 11697406. DOI: 10.1016/j.ncrna.2024.11.007.

Mitochondrial genome complexity in : nanopore sequencing reveals chloroplast gene transfer and DNA rearrangements.

Xie Y, Liu W, Guo L, Zhang X Front Genet. 2024; 15:1395805.

PMID: 38903753 PMC: 11188483. DOI: 10.3389/fgene.2024.1395805.

The initiation of mitochondrial DNA replication.

Liu Y, Liu H, Zhang F, Xu H Biochem Soc Trans. 2024; 52(3):1243-1251.

PMID: 38884788 PMC: 11346463. DOI: 10.1042/BST20230952.

CircR-loop: a novel RNA:DNA interaction on genome instability.

Su X, Feng Y, Chen R, Duan S Cell Mol Biol Lett. 2024; 29(1):89.

PMID: 38877420 PMC: 11177446. DOI: 10.1186/s11658-024-00606-5.

References

Xu B, Clayton D . A persistent RNA-DNA hybrid is formed during transcription at a phylogenetically conserved mitochondrial DNA sequence. Mol Cell Biol. 1995; 15(1):580-9. PMC: 232017. DOI: 10.1128/MCB.15.1.580. View

Ojala D, Montoya J, Attardi G . tRNA punctuation model of RNA processing in human mitochondria. Nature. 1981; 290(5806):470-4. DOI: 10.1038/290470a0. View

Wang J, Englund P, Jensen R . TbPIF8, a Trypanosoma brucei protein related to the yeast Pif1 helicase, is essential for cell viability and mitochondrial genome maintenance. Mol Microbiol. 2012; 83(3):471-85. PMC: 3262056. DOI: 10.1111/j.1365-2958.2011.07938.x. View

Farr C, Wang Y, Kaguni L . Functional interactions of mitochondrial DNA polymerase and single-stranded DNA-binding protein. Template-primer DNA binding and initiation and elongation of DNA strand synthesis. J Biol Chem. 1999; 274(21):14779-85. DOI: 10.1074/jbc.274.21.14779. View

Skourti-Stathaki K, Proudfoot N, Gromak N . Human senataxin resolves RNA/DNA hybrids formed at transcriptional pause sites to promote Xrn2-dependent termination. Mol Cell. 2011; 42(6):794-805. PMC: 3145960. DOI: 10.1016/j.molcel.2011.04.026. View

Sen D, Gilbert W . Formation of parallel four-stranded complexes by guanine-rich motifs in DNA and its implications for meiosis. Nature. 1988; 334(6180):364-6. DOI: 10.1038/334364a0. View

Korhonen J, Pham X, Pellegrini M, Falkenberg M . Reconstitution of a minimal mtDNA replisome in vitro. EMBO J. 2004; 23(12):2423-9. PMC: 423294. DOI: 10.1038/sj.emboj.7600257. View

Xu Y, Kimura T, Komiyama M . Human telomere RNA and DNA form an intermolecular G-quadruplex. Nucleic Acids Symp Ser (Oxf). 2008; (52):169-70. DOI: 10.1093/nass/nrn086. View

Korhonen J, Gaspari M, Falkenberg M . TWINKLE Has 5' -> 3' DNA helicase activity and is specifically stimulated by mitochondrial single-stranded DNA-binding protein. J Biol Chem. 2003; 278(49):48627-32. DOI: 10.1074/jbc.M306981200. View

10.

Belotserkovskii B, Liu R, Tornaletti S, Krasilnikova M, Mirkin S, Hanawalt P . Mechanisms and implications of transcription blockage by guanine-rich DNA sequences. Proc Natl Acad Sci U S A. 2010; 107(29):12816-21. PMC: 2919923. DOI: 10.1073/pnas.1007580107. View

11.

Mignotte B, MARSAULT J . Effects of the Xenopus laevis mitochondrial single-stranded DNA-binding protein on the activity of DNA polymerase gamma. Eur J Biochem. 1988; 174(3):479-84. DOI: 10.1111/j.1432-1033.1988.tb14123.x. View

12.

Wanrooij S, Fuste J, Farge G, Shi Y, Gustafsson C, Falkenberg M . Human mitochondrial RNA polymerase primes lagging-strand DNA synthesis in vitro. Proc Natl Acad Sci U S A. 2008; 105(32):11122-7. PMC: 2516254. DOI: 10.1073/pnas.0805399105. View

13.

Fuste J, Wanrooij S, Jemt E, Granycome C, Cluett T, Shi Y . Mitochondrial RNA polymerase is needed for activation of the origin of light-strand DNA replication. Mol Cell. 2010; 37(1):67-78. DOI: 10.1016/j.molcel.2009.12.021. View

14.

Lee D, Clayton D . Properties of a primer RNA-DNA hybrid at the mouse mitochondrial DNA leading-strand origin of replication. J Biol Chem. 1996; 271(39):24262-9. DOI: 10.1074/jbc.271.39.24262. View

15.

Futami K, Shimamoto A, Furuichi Y . Mitochondrial and nuclear localization of human Pif1 helicase. Biol Pharm Bull. 2007; 30(9):1685-92. DOI: 10.1248/bpb.30.1685. View

16.

Sanders C . Human Pif1 helicase is a G-quadruplex DNA-binding protein with G-quadruplex DNA-unwinding activity. Biochem J. 2010; 430(1):119-28. DOI: 10.1042/BJ20100612. View

17.

Cheng X, Dunaway S, Ivessa A . The role of Pif1p, a DNA helicase in Saccharomyces cerevisiae, in maintaining mitochondrial DNA. Mitochondrion. 2007; 7(3):211-22. DOI: 10.1016/j.mito.2006.11.023. View

18.

Boule J, Zakian V . The yeast Pif1p DNA helicase preferentially unwinds RNA DNA substrates. Nucleic Acids Res. 2007; 35(17):5809-18. PMC: 2034482. DOI: 10.1093/nar/gkm613. View

19.

Falkenberg M, Gaspari M, Rantanen A, Trifunovic A, Larsson N, Gustafsson C . Mitochondrial transcription factors B1 and B2 activate transcription of human mtDNA. Nat Genet. 2002; 31(3):289-94. DOI: 10.1038/ng909. View

20.

Xu B, Clayton D . RNA-DNA hybrid formation at the human mitochondrial heavy-strand origin ceases at replication start sites: an implication for RNA-DNA hybrids serving as primers. EMBO J. 1996; 15(12):3135-43. PMC: 450256. View