R-loop-dependent rolling-circle replication and a new model for DNA concatemer resolution by mitochondrial plasmid mp1

Backert, Steffen

doi:10.1093/emboj/cdf311

Cited by 28 publications

(26 citation statements)

References 44 publications

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“…In support of this, Flores and Lambert provided evidence that HPV16 and HPV31 undergo a shift in the mode of replication upon differentiation (69). In undifferentiated cells, HPV replicates bidirectionally; however, two-dimensional (2D) gel analysis revealed that productively replicating genomes exhibit branched structures, including X structures and simple Y arcs, consistent with strand invasion (Holliday junction formation) and unidirectional (rolling-circle) replication, respectively (70,71). These studies offer support for our findings that recombination is important for successful viral genome amplification and suggest that productive replication may occur through rolling-circle replication and concatemer formation.…”

Section: Discussionsupporting

confidence: 73%

Homologous Recombination Repair Factors Rad51 and BRCA1 Are Necessary for Productive Replication of Human Papillomavirus 31

Chappell

Gautam

et al. 2016

J Virol

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High-risk human papillomavirus 31 (HPV31)-positive cells exhibit constitutive activation of the ATM-dependent DNA damage response (DDR), which is necessary for productive viral replication. In response to DNA double-strand breaks (DSBs), ATM activation leads to DNA repair through homologous recombination (HR), which requires the principal recombinase protein Rad51, as well as BRCA1. Previous studies from our lab demonstrated that Rad51 and BRCA1 are expressed at high levels in HPV31-positive cells and localize to sites of viral replication. These results suggest that HPV may utilize ATM activity to increase HR activity as a means to facilitate viral replication. In this study, we demonstrate that high-risk HPV E7 expression alone is sufficient for the increase in Rad51 and BRCA1 protein levels. We have found that this increase occurs, at least in part, at the level of transcription. Studies analyzing protein stability indicate that HPV may also protect Rad51 and BRCA1 from turnover, contributing to the overall increase in cellular levels. We also demonstrate that Rad51 is bound to HPV31 genomes, with binding increasing per viral genome upon productive replication. We have found that depletion of Rad51 and BRCA1, as well as inhibition of Rad51's recombinase activity, abrogates productive viral replication upon differentiation. Overall, these results indicate that Rad51 and BRCA1 are required for the process of HPV31 genome amplification and suggest that productive replication occurs in a manner dependent upon recombination. IMPORTANCEProductive replication of HPV31 requires activation of an ATM-dependent DNA damage response, though how ATM activity contributes to replication is unclear. Rad51 and BRCA1 play essential roles in repair of double-strand breaks, as well as the restart of stalled replication forks through homologous recombination (HR). Given that ATM activity is required to initiate HR repair, coupled with the requirement of Rad51 and BRCA1 for productive viral replication, our findings suggest that HPV may utilize ATM activity to ensure localization of recombination factors to productively replicating viral genomes. The finding that E7 increases the levels of Rad51 and BRCA1 suggests that E7 contributes to productive replication by providing DNA repair factors required for viral DNA synthesis. Our studies not only imply a role for recombination in the regulation of productive HPV replication but provide further insight into how HPV manipulates the DDR to facilitate the productive phase of the viral life cycle. H uman papillomaviruses (HPVs) are small double-stranded DNA viruses approximately 8 kb in size that exhibit a preferential tropism for epithelial cells. High-risk mucosal HPV subtypes are the causative agents of cervical cancer and have been increasingly associated with anogenital, oropharyngeal, and head and neck cancers (1). The life cycle of HPV is intimately linked to the differentiation of its host cell, the keratinocyte (2). After exposure through a microwound in the stratified ep...

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Section: Discussionsupporting

confidence: 73%

Homologous Recombination Repair Factors Rad51 and BRCA1 Are Necessary for Productive Replication of Human Papillomavirus 31

Chappell

Gautam

et al. 2016

J Virol

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“…The recombination-based mtDNA replication model has recently gained strong support from studies suggesting that the replicative forks in Candida albicans mtDNA are generated from recombination intermediates (196). The presence of recombination-based replication in human heart and brain (87) and in plant mitochondria (197)(198)(199) has also been proposed. The physiological impact of this specific mechanism vis-à-vis the conventional RNA-primed replication mode is yet to be substantiated.…”

Section: Linking Mtdna Recombination Repair and Replicationmentioning

confidence: 99%

Mechanism of Homologous Recombination and Implications for Aging-Related Deletions in Mitochondrial DNA

Chen

2013

Microbiol Mol Biol Rev

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SUMMARY Homologous recombination is a universal process, conserved from bacteriophage to human, which is important for the repair of double-strand DNA breaks. Recombination in mitochondrial DNA (mtDNA) was documented more than 4 decades ago, but the underlying molecular mechanism has remained elusive. Recent studies have revealed the presence of a Rad52-type recombination system of bacteriophage origin in mitochondria, which operates by a single-strand annealing mechanism independent of the canonical RecA/Rad51-type recombinases. Increasing evidence supports the notion that, like in bacteriophages, mtDNA inheritance is a coordinated interplay between recombination, repair, and replication. These findings could have profound implications for understanding the mechanism of mtDNA inheritance and the generation of mtDNA deletions in aging cells.

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“…The DNA in the gel was transferred to a Hybond-N ϩ membrane (Amersham Biosciences, Piscataway, NJ) in 10ϫ SSC (1.5 M NaCl, 0.15M sodium citrate) by capillary blotting and was analyzed by a Southern hybridization with 32 Plabeled pUC119 DNA, which shares some sequence with pMK2 DNA and thus can be used as a specific probe. The DNA species on the two-dimensional gel were assigned as described previously (Backert, 2002).…”

Section: Analyses Of Mt-pmk2 Dna Isolated From Cells By Standard One mentioning

confidence: 99%

“…The mt-pMK2 DNA was detected by Southern hybridization by using the [ 32 P]pMK2 DNA-probe, after separation by size in the first dimensional electrophoresis and by shape in the second dimensional run. (C) Schematic diagram; 1, linear monomer; m (area), nicked circular monomer; 2, circular multimers; c (area), linear concatemers (see Backert, 2002 for the assignments of DNA species); 1D, the first dimensional run; 2D, the second dimensional run. (D) The twodimensional gel profile of the mt-pMK2 DNA isolated from MHR1 cells grown at 34°C.…”

Section: The Effects Of Mhr1p Overproduction On the Vegetative Segregmentioning

confidence: 99%

Mhr1p-dependent Concatemeric Mitochondrial DNA Formation for Generating Yeast Mitochondrial Homoplasmic Cells

Ling

Shibata

2004

MBoC

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Mitochondria carry many copies of mitochondrial DNA (mtDNA), but mt-alleles quickly segregate during mitotic growth through unknown mechanisms. Consequently, all mtDNA copies are often genetically homogeneous within each individual ("homoplasmic"). Our previous study suggested that tandem multimers ("concatemers") formed mainly by the Mhr1p (a yeast nuclear gene-encoded mtDNA-recombination protein)-dependent pathway are required for mtDNA partitioning into buds with concomitant monomerization. The transmission of a few randomly selected clones (as concatemers) of mtDNA into buds is a possible mechanism to establish homoplasmy. The current study provides evidence for this hypothesis as follows: the overexpression of MHR1 accelerates mt-allele-segregation in growing heteroplasmic zygotes, and mhr1-1 (recombination-deficient) causes its delay. The mt-allele-segregation rate correlates with the abundance of concatemers, which depends on Mhr1p. In G1-arrested cells, concatemeric mtDNA was labeled by [ 14 C]thymidine at a much higher density than monomers, indicating concatemers as the immediate products of mtDNA replication, most likely in a rolling circle mode. After releasing the G1 arrest in the absence of [ 14 C]thymidine, the monomers as the major species in growing buds of dividing cells bear a similar density of 14 C as the concatemers in the mother cells, indicating that the concatemers in mother cells are the precursors of the monomers in buds.

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R-loop-dependent rolling-circle replication and a new model for DNA concatemer resolution by mitochondrial plasmid mp1

Cited by 28 publications

References 44 publications

Homologous Recombination Repair Factors Rad51 and BRCA1 Are Necessary for Productive Replication of Human Papillomavirus 31

Homologous Recombination Repair Factors Rad51 and BRCA1 Are Necessary for Productive Replication of Human Papillomavirus 31

Mechanism of Homologous Recombination and Implications for Aging-Related Deletions in Mitochondrial DNA

Mhr1p-dependent Concatemeric Mitochondrial DNA Formation for Generating Yeast Mitochondrial Homoplasmic Cells

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