DNA repair in plastids

DNA replication, recombination, and repair are interrelated processes and the homologous recombination pathway in plastids is likely to play a role in DNA repair (Cerutti et al. 1995). The estimated mutation rate of plastid genes is approximately two-fold lower than that of nuclear genes (Wolfe et al. 1987). Within plastids the synonymous substitution rate of genes located in the large inverted repeat is about two-fold lower than that for genes located in the single copy regions (Perry and Wolfe 2002). This has been interpreted to be the result of the two-fold higher dosage of inverted repeat sequences and biased gene conversion in favour of WT plastid DNA sequences (Birky and Walsh 1992; Perry and Wolfe 2002). Non-biased repair will either correct the mutation to WT or fix the mutation (convert WT to mutant) and give rise to both outcomes in equal proportions. Biased repair favours one of these outcomes to give 100% of only one product (Fig. 14). Direct experimental confirmation for biased gene conversion has been obtained by monitoring correction of mutations tightly linked (31 bp distance) to an aadA insertion in transgenic N. tabacum plastids (Khakhlova and Bock 2006). Whilst the aadA gene was retained by spectinomycin selection the mutations were repaired to WT with a bias for reversing AT to GC changes more efficiently than GC to AT mutations. It has been suggested that this bias towards AT might underlie the high overall AT content (>70% AT) of plastid genomes (Khakhlova and Bock 2006). Multiple copies of plastid DNA and biased gene conversion in favour of WT would reduce the rate at which mutations are fixed.

Alternatives to RecA-based recombination repair include photoreactivation, base excision repair, nucleotide excision repair, and mismatch repair (Kimura and Sakaguchi 2006). Little is known on these alternative repair pathways in plastids. A putative plastid-localised uracil-DNA glycosylase activity probably involved in base excision repair was partially purified from Z. mays chloroplasts (Bensen and Warner 1987). UV-induced lesions in the plastid psbA gene of G. max suspension culture cells were repaired in the light (but not in the dark) with kinetics that were considerably slower than expected for photoreactivation by photolyases (Cannon et al. 1995). Experiments on purified S. oleracea chloroplasts (Hada et al. 2000) and the lack of identification of plastid transit peptides in the products of plant genes encoding photolyases (Draper and Hays 2000) led to the possibility that plastids might be deficient in photolyase-mediated photoreactivation. However, tolerance of plastid DNA replication to UV-B lesions in A. thaliana plants grown in blue (photorepair-compatible) light might suggest the presence of as yet unidentified photolyases in plastids (Draper and Hays 2000). Under gold light where light-dependent photorepair does not take place, a UV-B fluence rate of 5 kJ m-2 inhibits replication of plastid DNA but not nuclear and mitochondrial DNA indicating a deficiency in light-independent (dark) repair pathways in chloroplasts (Draper and Hays 2000).

Endonuclease activities that could act on apurinic sites following base removal were purified from H. vulgare chloroplasts (Veleminskiy et al. 1980). A singlestrand specific nuclease activity from T. aestivum chloroplasts cleaves single stranded DNA or RNA regions including 5' flaps, 5' overhangs, and 3' pseudoflaps and has been suggested to be involved in DNA repair (Przykorska et al. 2004). The multi-subunit replication protein A (RPA) binds to single-stranded DNA and is involved in pathways including nucleotide excision repair. RPA sub-units appear to be targeted to plastids (Kimura and Sakaguchi 2006). Plastid-localised homologues of RecQ have been implicated in DNA repair (see Sec-tion13.5 below). Nitroso-methy-urea and nitroso-guanidine are particularly effective for inducing mutations in flowering plant plastid genomes (Hagemann 1976). Methyl transferases reverse the damage to bases caused by these alkylating agents. The presence of methyl transferases in nuclei but their absence in plastids might explain the utility of these mutagens for inducing plastid mutations (Sears 1998).

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