A Novel Intermediate in Transcription Initiation by Human Mitochondrial RNA Polymerase

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 2014 Jan 8

PMID 24393772

Citations 41

Authors

Yaroslav I Morozov

Karen Agaronyan

Alan C M Cheung

Michael Anikin

Patrick Cramer

Dmitry Temiakov

Affiliations

Soon will be listed here.

Abstract

The mitochondrial genome is transcribed by a single-subunit T7 phage-like RNA polymerase (mtRNAP), structurally unrelated to cellular RNAPs. In higher eukaryotes, mtRNAP requires two transcription factors for efficient initiation-TFAM, a major nucleoid protein, and TFB2M, a transient component of mtRNAP catalytic site. The mechanisms behind assembly of the mitochondrial transcription machinery and its regulation are poorly understood. We isolated and identified a previously unknown human mitochondrial transcription intermediate-a pre-initiation complex that includes mtRNAP, TFAM and promoter DNA. Using protein-protein cross-linking, we demonstrate that human TFAM binds to the N-terminal domain of mtRNAP, which results in bending of the promoter DNA around mtRNAP. The subsequent recruitment of TFB2M induces promoter melting and formation of an open initiation complex. Our data indicate that the pre-initiation complex is likely to be an important target for transcription regulation and provide basis for further structural, biochemical and biophysical studies of mitochondrial transcription.

Citing Articles

TEFM facilitates transition from RNA synthesis to DNA synthesis at H-strand replication origin of mtDNA.

Matsuda S, Nakayama M, Do Y, Ishiuchi T, Yagi M, Wanrooij S Commun Biol. 2025; 8(1):202.

PMID: 39922921 PMC: 11807126. DOI: 10.1038/s42003-025-07645-4.

Human mitochondrial RNA polymerase structures reveal transcription start-site and slippage mechanism.

Shen J, Goovaerts Q, Ajjugal Y, De Wijngaert B, Das K, Patel S bioRxiv. 2024; .

PMID: 39677640 PMC: 11642874. DOI: 10.1101/2024.12.02.626445.

Noncanonical Amino Acids in Biocatalysis.

Birch-Price Z, Hardy F, Lister T, Kohn A, Green A Chem Rev. 2024; 124(14):8740-8786.

PMID: 38959423 PMC: 11273360. DOI: 10.1021/acs.chemrev.4c00120.

Sequence-specific dynamic DNA bending explains mitochondrial TFAM's dual role in DNA packaging and transcription initiation.

Huh H, Shen J, Ajjugal Y, Ramachandran A, Patel S, Lee S Nat Commun. 2024; 15(1):5446.

PMID: 38937458 PMC: 11211510. DOI: 10.1038/s41467-024-49728-6.

Mechanisms and pathologies of human mitochondrial DNA replication and deletion formation.

Bernardino Gomes T, Vincent A, Menger K, Stewart J, Nicholls T Biochem J. 2024; 481(11):683-715.

PMID: 38804971 PMC: 11346376. DOI: 10.1042/BCJ20230262.

References

Kukat C, Larsson N . mtDNA makes a U-turn for the mitochondrial nucleoid. Trends Cell Biol. 2013; 23(9):457-63. DOI: 10.1016/j.tcb.2013.04.009. View

Fuda N, Ardehali M, Lis J . Defining mechanisms that regulate RNA polymerase II transcription in vivo. Nature. 2009; 461(7261):186-92. PMC: 2833331. DOI: 10.1038/nature08449. View

Cheetham G, Steitz T . Structure of a transcribing T7 RNA polymerase initiation complex. Science. 1999; 286(5448):2305-9. DOI: 10.1126/science.286.5448.2305. View

Savkina M, Temiakov D, McAllister W, Anikin M . Multiple functions of yeast mitochondrial transcription factor Mtf1p during initiation. J Biol Chem. 2009; 285(6):3957-3964. PMC: 2823537. DOI: 10.1074/jbc.M109.051003. View

Zollo O, Tiranti V, Sondheimer N . Transcriptional requirements of the distal heavy-strand promoter of mtDNA. Proc Natl Acad Sci U S A. 2012; 109(17):6508-12. PMC: 3340101. DOI: 10.1073/pnas.1118594109. View

Sato S, Mimasu S, Sato A, Hino N, Sakamoto K, Umehara T . Crystallographic study of a site-specifically cross-linked protein complex with a genetically incorporated photoreactive amino acid. Biochemistry. 2010; 50(2):250-7. DOI: 10.1021/bi1016183. View

Litonin D, Sologub M, Shi Y, Savkina M, Anikin M, Falkenberg M . Human mitochondrial transcription revisited: only TFAM and TFB2M are required for transcription of the mitochondrial genes in vitro. J Biol Chem. 2010; 285(24):18129-33. PMC: 2881736. DOI: 10.1074/jbc.C110.128918. View

Ptashne M, Gann A . Transcriptional activation by recruitment. Nature. 1997; 386(6625):569-77. DOI: 10.1038/386569a0. View

Kouzine F, Wojtowicz D, Yamane A, Resch W, Kieffer-Kwon K, Bandle R . Global regulation of promoter melting in naive lymphocytes. Cell. 2013; 153(5):988-99. PMC: 3684982. DOI: 10.1016/j.cell.2013.04.033. View

10.

Lodeiro M, Uchida A, Bestwick M, Moustafa I, Arnold J, Shadel G . Transcription from the second heavy-strand promoter of human mtDNA is repressed by transcription factor A in vitro. Proc Natl Acad Sci U S A. 2012; 109(17):6513-8. PMC: 3340051. DOI: 10.1073/pnas.1118710109. View

11.

Villemain J, Guajardo R, Sousa R . Role of open complex instability in kinetic promoter selection by bacteriophage T7 RNA polymerase. J Mol Biol. 1998; 273(5):958-77. DOI: 10.1006/jmbi.1997.1358. View

12.

Larsson N, Wang J, Wilhelmsson H, Oldfors A, Rustin P, Lewandoski M . Mitochondrial transcription factor A is necessary for mtDNA maintenance and embryogenesis in mice. Nat Genet. 1998; 18(3):231-6. DOI: 10.1038/ng0398-231. View

13.

Gill S, Yager T, von Hippel P . Thermodynamic analysis of the transcription cycle in E. coli. Biophys Chem. 1990; 37(1-3):239-50. DOI: 10.1016/0301-4622(90)88023-l. View

14.

Dorman G, Prestwich G . Benzophenone photophores in biochemistry. Biochemistry. 1994; 33(19):5661-73. DOI: 10.1021/bi00185a001. View

15.

Temiakov D, Mentesana P, Ma K, Mustaev A, Borukhov S, McAllister W . The specificity loop of T7 RNA polymerase interacts first with the promoter and then with the elongating transcript, suggesting a mechanism for promoter clearance. Proc Natl Acad Sci U S A. 2000; 97(26):14109-14. PMC: 18879. DOI: 10.1073/pnas.250473197. View

16.

Dairaghi D, Shadel G, Clayton D . Addition of a 29 residue carboxyl-terminal tail converts a simple HMG box-containing protein into a transcriptional activator. J Mol Biol. 1995; 249(1):11-28. DOI: 10.1006/jmbi.1995.9889. View

17.

Young T, Ahmad I, Yin J, Schultz P . An enhanced system for unnatural amino acid mutagenesis in E. coli. J Mol Biol. 2009; 395(2):361-74. DOI: 10.1016/j.jmb.2009.10.030. View

18.

Lu B, Lee J, Nie X, Li M, Morozov Y, Venkatesh S . Phosphorylation of human TFAM in mitochondria impairs DNA binding and promotes degradation by the AAA+ Lon protease. Mol Cell. 2012; 49(1):121-32. PMC: 3586414. DOI: 10.1016/j.molcel.2012.10.023. 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.

Gaspari M, Larsson N, Gustafsson C . The transcription machinery in mammalian mitochondria. Biochim Biophys Acta. 2004; 1659(2-3):148-52. DOI: 10.1016/j.bbabio.2004.10.003. View