Mutations defective in ribonucleotide reductase activity interfere with pollen plastid DNA degradation mediated by DPD1 exonuclease

Abstract

SUMMARYOrganellar DNAs in mitochondria and plastids are present in multiple copies and make up a substantial proportion of total cellular DNA despite their limited genetic capacity. We recently demonstrated that organellar DNA degradation occurs during pollen maturation, mediated by the Mg 2+ -dependent organelle exonuclease DPD1. To further understand organellar DNA degradation, we characterized a distinct mutant (dpd2). In contrast to the dpd1 mutant, which retains both plastid and mitochondrial DNAs, dpd2 showed specific accumulation of plastid DNAs. Multiple abnormalities in vegetative and reproductive tissues of dpd2 were also detected. DPD2 encodes the large subunit of ribonucleotide reductase, an enzyme that functions at the rate-limiting step of de novo nucleotide biosynthesis. We demonstrated that the defects in ribonucleotide reductase indirectly compromise the activity of DPD1 nuclease in plastids, thus supporting a different regulation of organellar DNA degradation in pollen. Several lines of evidence provided here reinforce our previous conclusion that the DPD1 exonuclease plays a central role in organellar DNA degradation, functioning in DNA salvage rather than maternal inheritance during pollen development.

Elevation of Pollen Mitochondrial DNA Copy Number by WHIRLY2: Altered Respiration and Pollen Tube Growth in Arabidopsis

“…However, as a nonenzyme DNA-binding factor without sequence specificity (Maréchal et al, 2008;Cappadocia et al, 2010), AtWHY2 may affect mtDNA copy number in a way similar to TFAM, by impeding mtDNA degradation by pollen mitochondrial DNases such as DPD1 (Matsushima et al, 2011;Tang et al, 2012). This agrees with a previous study showing that AtWHY2 protects DNA against nuclease digestion in vitro (Cappadocia et al, 2010).…”

“…1, A and B) and other, earlier developmental stages (Supplemental Fig. S1), indicating a decrease of mtDNA levels during pollen development, as indicated previously (Nagata et al, 1999;Matsushima et al, 2011;Tang et al, 2012). Using a competitive PCR technique developed for determining the copy number of mtDNA in single cells (Wang et al, 2010), we measured 133.3 6 11.5 copies of the mtDNA per BCP and 10.3 6 2.1 per mature pollen grain ( Fig.…”

“…In addition, our investigation reports, to our knowledge, the first nonenzyme protein factor that affects mtDNA levels in plants. In fact, except for the core enzymes of mtDNA replication, PolIA and PolIB, which increase the amount of mtDNA (Parent et al, 2011), and an exonuclease, DPD1, which degrades mtDNA (Matsushima et al, 2011;Tang et al, 2012), few plant factors have been experimentally shown to affect mtDNA quantity. The DNA-binding factors so far identified in plants, such as OSB, RecA, and MSH, likely affect mtDNA via its recombination and repair rather than regulate mtDNA levels (for review, see Maréchal and Brisson, 2010).…”

“…Mutation of Arabidopsis PolIA or PolIB (homologs of bacterial DNA polymerase I) causes a reduction in mtDNA copy number, and double mutation of these proteins is lethal (Parent et al, 2011). Also, an Mg 2+ -dependent exonuclease, DEFECTIVE IN POLLEN ORGANELLE DNA DEGRADATION1 (DPD1), degrades organelle DNA, helping to produce the proper amounts of mtDNA in pollen cells (Matsushima et al, 2011;Tang et al, 2012). These results provide insights into the molecular control of mtDNA levels in plants, via both mtDNA replication and mtDNA degradation.…”

Cellular and molecular aspects of quinoa leaf senescence

“…Furthermore, DPD1 is the only example among nucleases in which DNA degradation has been confirmed in vivo [14][15][16][17]. However, DNA degradation and the action of these nucleases remain limited because nucleases have high affinity to ssDNA and RNA and are localised in specific cellular compartments (namely, mitochondria and chloroplasts), raising the possibility that they do not act specifically on genomic DNA degradation.…”