Showing papers by "Jeremy N. Timmis published in 2012"

Environmental stress increases the entry of cytoplasmic organellar DNA into the nucleus in plants

Abstract: Mitochondria and chloroplasts (photosynthetic members of the plastid family of cytoplasmic organelles) in eukaryotic cells originated more than a billion years ago when an ancestor of the nucleated cell engulfed two different prokaryotes in separate sequential events. Extant cytoplasmic organellar genomes contain very few genes compared with their candidate free-living ancestors, as most have functionally relocated to the nucleus. The first step in functional relocation involves the integration of inactive DNA fragments into nuclear chromosomes, and this process continues at high frequency with attendant genetic, genomic, and evolutionary consequences. Using two different transplastomic tobacco lines, we show that DNA migration from chloroplasts to the nucleus is markedly increased by mild heat stress. In addition, we show that insertion of mitochondrial DNA fragments during the repair of induced double-strand breaks is increased by heat stress. The experiments demonstrate that the nuclear influx of organellar DNA is a potentially a source of mutation for nuclear genomes that is highly susceptible to temperature fluctuations that are well within the range experienced naturally.

Single molecule PCR reveals similar patterns of non-homologous DSB repair in tobacco and Arabidopsis.

Abstract: DNA double strand breaks (DSBs) occur constantly in eukaryotes. These potentially lethal DNA lesions are repaired efficiently by two major DSB repair pathways: homologous recombination and non-homologous end joining (NHEJ). We investigated NHEJ in Arabidopsis thaliana and tobacco (Nicotiana tabacum) by introducing DNA double-strand breaks through inducible expression of I-SceI, followed by amplification of individual repair junction sequences by single-molecule PCR. Using this process over 300 NHEJ repair junctions were analysed in each species. In contrast to previously published variation in DSB repair between Arabidopsis and tobacco, the two species displayed similar DSB repair profiles in our experiments. The majority of repair events resulted in no loss of sequence and small (1–20 bp) deletions occurred at a minority (25–45%) of repair junctions. Approximately ∼1.5% of the observed repair events contained larger deletions (>20 bp) and a similar percentage contained insertions. Strikingly, insertion events in tobacco were associated with large genomic deletions at the site of the DSB that resulted in increased micro-homology at the sequence junctions suggesting the involvement of a non-classical NHEJ repair pathway. The generation of DSBs through inducible expression of I-SceI, in combination with single molecule PCR, provides an effective and efficient method for analysis of individual repair junctions and will prove a useful tool in the analysis of NHEJ.

Plastid Sequences Contribute to Some Plant Mitochondrial Genes

Abstract: DNA of plastid (chloroplast) origin comprises between 1% and 10% of the mitochondrial genomes of higher plants, but functions are currently considered to be limited to rare instances where plastid tRNA genes have replaced their mitochondrial counterparts, where short patches of mitochondrial genes evolved using their homologous plastidic copies by gene conversion or where a new promoter region is created. Here, we show that, in some angiosperms, plastid-derived DNA in mitochondrial genomes (also called mtpt for mitochondrial plastid DNA) contributes codons to unrelated mitochondrial protein-coding sequences and may also have a role in posttranscriptional RNA processing. We determined that these transfers of plastid DNA occurred a few to 150 Ma and that mtpts can sometimes remain dormant many millions of years before contributing to the mitochondrial proteome.

Promiscuous Organellar DNA

Abstract: Endosymbiotic transfer of DNA from the cytoplasmic organelles (mitochondria and chloroplasts) to the nucleus has been a major factor driving the origin of new nuclear genes, a process central to eukaryote evolution. Typically, transfer of organelle DNA to the nucleus is quickly followed by decay, deletion and rearrangement. However, in rare instances these new sequences lead to functional relocation of organelle genes to the nucleus or the generation of genes with novel function. Similar transfer of chloroplast DNA has also added to the complexity of plant mitochondrial genomes. Significantly, these processes are ongoing, making promiscuous organellar DNA an important contributor to the continued evolution of plant genomes.

Plastid DNA in the nucleus: new genes for old.

Abstract: Nuclear genomes of eukaryotes are bombarded by a continuous deluge of organellar DNA which contributes significantly to eukaryote evolution. Here, we present a new PCR-based method that allows the specific amplification of nuclear integrants of organellar DNA (norgs) by exploiting recent deletions present in organellar genome sequences. We have used this method to amplify nuclear integrants of plastid DNA (nupts) from the nuclear genomes of several nicotiana species and to study the evolutionary forces acting upon these sequences. The role of nupts in endosymbiotic evolution and the different genetic factors influencing the time available for a chloroplastic gene to be functionally relocated in the nucleus are discussed.

Nuclear genome diversity in somatic cells is accelerated by environmental stress

Abstract: DNA transfer to the nucleus from prokaryotic ancestors of the cytoplasmic organelles (mitochondria and plastids) has occurred during endosymbiotic evolution in eukaryotes. In most eukaryotes, organelle DNA transfer to nucleus is a continuing process. The frequency of DNA transposition from plastid (chloroplast) to nucleus has been measured in tobacco plants (Nicotiana tabacum) experimentally. We have monitored the effects of environmental stress on the rate of DNA transfer from plastid to nucleus by exploiting nucleus-specific reporter genes in two transplastomic tobacco lines. DNA migration from plastids to the nucleus is markedly increased by mild heat stress. In addition, insertions of mitochondrial DNA into induced double-strand breaks are observed after heat treatment. These results show that movement of organelle DNA to the nucleus is remarkably increased by heat stress.

Gene Transfer to the Nucleus

Abstract: The endosymbiotic evolution of mitochondrial and plastid genomes from ancestral prokaryotes is outlined stressing the net passage of genes from the organelle precursor genomes to the nucleus. We highlight recent experimental analyses of the process of nucleic acid transfer between extant organelle genomes and the nucleus. The molecular events that have accompanied functional gene transfers are described and specific cases of the relocation and nuclear expression of former organellar genes are summarised. The mechanisms underpinning gene transfer, including the movement of extant cytoplasmic organellar nucleic acids are described together with emphasis on the genetic consequences for the nuclear size and genetic complexity. We discuss the major contribution of endosymbiosis to the evolution of eukaryotes and we mention the selective forces that may be responsible for making the nucleus the preferred location for eukaryotic genes.

2012 Landes Bioscience. Do not distribute. Nuclear genome diversity in somatic cells is accelerated by environmental stress

Abstract: Keywords: environmental stress,DNA transfer, chloroplast, mitochondria,endosymbiotic evolution, cell divisionAbbreviations: nupt, nuclear integrant ofplastid DNA; numt, nuclear integrant ofmitochondrial DNA; norg, nuclearintegrant organelle DNA; cp, chloroplast;ptDNA, plastid DNA; DSB, double-strandbreak; orgDNA, organelle DNA; orgV,organellar value; EGT, endosymbiotic genetransfer; ORF, open reading frameSubmitted: 02/29/12Accepted: 03/01/12http://dx.doi.org/10.4161/psb.19871