| 1. |
- Fusté, Javier Miralles, et al.
(författare)
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Mitochondrial RNA polymerase is needed for activation of the origin of light-strand DNA replication.
- 2010
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Ingår i: Molecular cell. - : Elsevier BV. - 1097-4164 .- 1097-2765. ; 277, s. 225-225
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Tidskriftsartikel (refereegranskat)abstract
- Mitochondrial DNA is replicated by a unique enzymatic machinery, which is distinct from the replication apparatus used for copying the nuclear genome. We examine here the mechanisms of origin-specific initiation of lagging-strand DNA synthesis in human mitochondria. We demonstrate that the mitochondrial RNA polymerase (POLRMT) is the primase required for initiation of DNA synthesis from the light-strand origin of DNA replication (OriL). Using only purified POLRMT and DNA replication factors, we can faithfully reconstitute OriL-dependent initiation in vitro. Leading-strand DNA synthesis is initiated from the heavy-strand origin of DNA replication and passes OriL. The single-stranded OriL is exposed and adopts a stem-loop structure. At this stage, POLRMT initiates primer synthesis from a poly-dT stretch in the single-stranded loop region. After about 25 nt, POLRMT is replaced by DNA polymerase gamma, and DNA synthesis commences. Our findings demonstrate that POLRMT can function as an origin-specific primase in mammalian mitochondria.
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| 2. |
- Nicholls, Thomas J., et al.
(författare)
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Dinucleotide Degradation by REXO2 Maintains Promoter Specificity in Mammalian Mitochondria
- 2019
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765 .- 1097-4164. ; 76:5, s. 784-
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Tidskriftsartikel (refereegranskat)abstract
- Oligoribonucleases are conserved enzymes that degrade short RNA molecules of up to 5 nt in length and are assumed to constitute the final stage of RNA turnover. Here we demonstrate that REXO2 is a specialized dinucleotide-degrading enzyme that shows no preference between RNA and DNA dinucleotide substrates. A heart- and skeletal-muscle-specific knockout mouse displays elevated dinucleotide levels and alterations in gene expression patterns indicative of aberrant dinucleotide-primed transcription initiation. We find that dinucleotides act as potent stimulators of mitochondrial transcription initiation in vitro. Our data demonstrate that increased levels of dinucleotides can be used to initiate transcription, leading to an increase in transcription levels from both mitochondrial promoters and other, nonspecific sequence elements in mitochondrial DNA. Efficient RNA turnover by REXO2 is thus required to maintain promoter specificity and proper regulation of transcription in mammalian mitochondria.
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| 3. |
- Nicholls, Thomas J., et al.
(författare)
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Topoisomerase 3α Is Required for Decatenation and Segregation of Human mtDNA
- 2018
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765 .- 1097-4164. ; 69:1, s. 9-23.e6
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Tidskriftsartikel (refereegranskat)abstract
- How mtDNA replication is terminated and the newly formed genomes are separated remain unknown. We here demonstrate that the mitochondrial isoform of topoisomerase 3α (Top3α) fulfills this function, acting independently of its nuclear role as a component of the Holliday junction-resolving BLM-Top3α-RMI1-RMI2 (BTR) complex. Our data indicate that mtDNA replication termination occurs via a hemicatenane formed at the origin of H-strand replication and that Top3α is essential for resolving this structure. Decatenation is a prerequisite for separation of the segregating unit of mtDNA, the nucleoid, within the mitochondrial network. The importance of this process is highlighted in a patient with mitochondrial disease caused by biallelic pathogenic variants in TOP3A, characterized by muscle-restricted mtDNA deletions and chronic progressive external ophthalmoplegia (CPEO) plus syndrome. Our work establishes Top3α as an essential component of the mtDNA replication machinery and as the first component of the mtDNA separation machinery. Nicholls et al. identify a role for topoisomerase 3α in the separation of mtDNA following replication. Loss of Top3α activity impairs mtDNA segregation and, consequently, segregation of the mtDNA nucleoid within the mitochondrial network. Mutations in TOP3A cause human mitochondrial disease associated with mtDNA deletions and impaired mtDNA separation. © 2017 Elsevier Inc.
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| 4. |
- Zhu, Xuefeng, et al.
(författare)
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Genome-wide occupancy profile of mediator and the Srb8-11 module reveals interactions with coding regions
- 2006
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765 .- 1097-4164. ; 22:2, s. 169-178
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Tidskriftsartikel (refereegranskat)abstract
- Mediator exists in a free form containing the Med12, Med13, CDK8, and CycC subunits (the Srb8-11 module) and a smaller form, which lacks these four subunits and associates with RNA polymerase II (Pol II), forming a holoenzyme. We use chromatin immunoprecipitation (ChIP) and DNA microarrays to investigate genome-wide localization of Mediator and the Srb8-11 module in fission yeast. Mediator and the Srb8-11 module display similar binding patterns, and interactions with promoters and upstream activating sequences correlate with increased transcription activity. Unexpectedly, Mediator also interacts with the downstream coding region of many genes. These interactions display a negative bias for positions closer to the 5' ends of open reading frames (ORFs) and appear functionally important, because downregulation of transcription in a temperature-sensitive med17 mutant strain correlates with increased Mediator occupancy in the coding region. We propose that Mediator coordinates transcription initiation with transcriptional events in the coding region of eukaryotic genes.
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