Mitochondrial RNase H1 activity regulates R-loop homeostasis to maintain genome integrity and enable early embryogenesis in Arabidopsis

Plant mitochondrial genomes undergo frequent homologous recombination (HR). Ectopic HR activity is inhibited by the HR surveillance pathway, but the underlying regulatory mechanism is unclear. Here, we show that the mitochondrial RNase H1 AtRNH1B impairs the formation of RNA:DNA hybrids (R-loops) and participates in the HR surveillance pathway in Arabidopsis thaliana. AtRNH1B suppresses ectopic HR at intermediate-sized repeats (IRs) and thus maintains mitochondrial DNA (mtDNA) replication. The RNase H1 AtRNH1C is restricted to the chloroplast; however, when cells lack AtRNH1B, transport of chloroplast AtRNH1C into the mitochondria secures HR surveillance, thus ensuring the integrity of the mitochondrial genome and allowing embryogenesis to proceed. HR surveillance is further regulated by the single-stranded DNA-binding protein ORGANELLAR SINGLE-STRANDED DNA BINDING PROTEIN1 (OSB1), which decreases the formation of R-loops. This study uncovers a facultative dual targeting mechanism between organelles and sheds light on the roles of RNase H1 in organellar genome maintenance and embryogenesis.

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CKS Proteins Protect Mitochondrial Genome Integrity by Interacting with Mitochondrial Single-stranded DNA-binding Protein

Molecular & Cellular Proteomics ◽

10.1074/mcp.m900078-mcp200 ◽

2009 ◽

Vol 9 (1) ◽

pp. 145-152 ◽

Cited By ~ 14

Author(s):

Marko Radulovic ◽

Eleanor Crane ◽

Mark Crawford ◽

Jasminka Godovac-Zimmermann ◽

Veronica P. C. C. Yu

Keyword(s):

Dna Binding ◽

Mitochondrial Genome ◽

Binding Protein ◽

Dna Binding Protein ◽

Genome Integrity ◽

Single Stranded Dna ◽

Cks Proteins

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The mitochondrial single-stranded DNA binding protein is essential for initiation of mtDNA replication

Science Advances ◽

10.1126/sciadv.abf8631 ◽

2021 ◽

Vol 7 (27) ◽

pp. eabf8631

Author(s):

Min Jiang ◽

Xie Xie ◽

Xuefeng Zhu ◽

Shan Jiang ◽

Dusanka Milenkovic ◽

...

Keyword(s):

Dna Binding ◽

Transcription Initiation ◽

Binding Protein ◽

Dna Binding Protein ◽

Replication Initiation ◽

Single Stranded Dna ◽

Mammalian Mitochondria ◽

Mtdna Replication ◽

Knockout Model ◽

Light Strand

We report a role for the mitochondrial single-stranded DNA binding protein (mtSSB) in regulating mitochondrial DNA (mtDNA) replication initiation in mammalian mitochondria. Transcription from the light-strand promoter (LSP) is required both for gene expression and for generating the RNA primers needed for initiation of mtDNA synthesis. In the absence of mtSSB, transcription from LSP is strongly up-regulated, but no replication primers are formed. Using deep sequencing in a mouse knockout model and biochemical reconstitution experiments with pure proteins, we find that mtSSB is necessary to restrict transcription initiation to optimize RNA primer formation at both origins of mtDNA replication. Last, we show that human pathological versions of mtSSB causing severe mitochondrial disease cannot efficiently support primer formation and initiation of mtDNA replication.

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Mitochondrial Single-stranded DNA-binding Protein Is Required for Mitochondrial DNA Replication and Development in Drosophila melanogaster

Molecular Biology of the Cell ◽

10.1091/mbc.12.4.821 ◽

2001 ◽

Vol 12 (4) ◽

pp. 821-830 ◽

Cited By ~ 62

Author(s):

Dieter Maier ◽

Carol L. Farr ◽

Burkhard Poeck ◽

Anuradha Alahari ◽

Marion Vogel ◽

...

Keyword(s):

Dna Binding ◽

Binding Protein ◽

Dna Binding Protein ◽

Single Stranded Dna ◽

Immunocytochemical Analysis ◽

The Third ◽

Mtdna Replication ◽

Mitochondrial Dna Replication ◽

Essential Function ◽

Higher Eukaryotes

The discovery that several inherited human diseases are caused by mtDNA depletion has led to an increased interest in the replication and maintenance of mtDNA. We have isolated a new mutant in thelopo (low power) gene fromDrosophila melanogaster affecting the mitochondrial single-stranded DNA-binding protein (mtSSB), which is one of the key components in mtDNA replication and maintenance.lopo 1 mutants die late in the third instar before completion of metamorphosis because of a failure in cell proliferation. Molecular, histochemical, and physiological experiments show a drastic decrease in mtDNA content that is coupled with the loss of respiration in these mutants. However, the number and morphology of mitochondria are not greatly affected. Immunocytochemical analysis shows that mtSSB is expressed in all tissues but is highly enriched in proliferating tissues and in the developing oocyte.lopo 1 is the first mtSSB mutant in higher eukaryotes, and its analysis demonstrates the essential function of this gene in development, providing an excellent model to study mitochondrial biogenesis in animals.

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