Biology subjects

Gualberto, J. M.

Publications and source records attributed to Gualberto, J. M..

3 recordsLinked to original sources

bioRxiv2024

R-Loop control and mitochondria genome stability requires the 5'-3' exonuclease/flap-endonuclease OEX1

Maintenance of the plant organelle genomes involves factors mostly inherited from their bacterial symbiotic ancestors. In bacteria, a major player in genome maintenance is DNA Polymerase I (Pol I), which provides a 5-3-exonuclease/flap-endonuclease activity required for multiple replication and repair functions. In plant organelles, DNA polymerases POL1A and POL1B are evolutionarily derived from DNA Pol I but lack this domain. In Arabidopsis, OEX1 and OEX2 (Organellar Exonucleases 1 and 2) represent this missing domain and are targeted to mitochondria and chloroplasts, respectively. An oex1 mutant allele shows developmental and fertility defects that correlate with the differential segregation of mitochondrial DNA (mtDNA) subgenomes generated by recombination, suggesting that OEX1 processes replication and recombination intermediates whose accumulation results in genome instability. Alternative splicing generates two OEX1 isoforms that can differentially interact with POL1A and POL1B and variably affect mtDNA repair. Recombinant OEX1 has 5-3-exonuclease and flap endonuclease activities, the latter being a key function in replication and repair. Furthermore, OEX1 has high affinity for RNA:DNA hybrids, rapidly degrading RNA in Okazaki-like structures and R-loops. Consistent with a role in suppressing R-loops, oex1 plants accumulate R-loops in highly transcribed mtDNA regions. Taken together, our results show that OEX1 plays multiple important roles in the processes required to maintain mtDNA stability.

plant biology↗

bioRxiv2019

RADA is the main branch migration factor in plant mitochondrial recombination and its defect leads to mtDNA instability and cell cycle arrest

The mitochondria of flowering plants have large and complex genomes whose structure and segregation are modulated by recombination activities. The late steps of mitochondrial recombination are still poorly characterized: while the loss of mitochondrial recombination is not viable, a deficiency in RECG1-dependent branch migration has little impact on plant development, implying the existence of alternative pathways. Here we present RADA, an ortholog of bacterial RadA/Sms, which is required for the processing of organellar recombination intermediates. While bacterial RadA is dispensable, RADA-deficient plants are severely impacted in their development and fertility, correlating with increased mtDNA ectopic recombination and replication of recombination-generated subgenomes. The radA mutation is epistatic to recG1, indicating that RADA drives the main branch migration pathway of plant mitochondria. In contrast, the double mutation radA recA3 is lethal, underlining the importance of an alternative RECA3-dependent pathway. Although RADA is dually targeted to mitochondria and chloroplasts, we found little to no effects of radA on the stability of the plastidial genome. The stunted growth of radA mutants could not be correlated with obvious defects in mitochondrial gene expression. Rather, it seems that is partially caused by a retrograde signal that activates nuclear genes repressing cell cycle progression.

plant biology↗

bioRxiv2018

Reciprocal cybrids reveal how organellar genomes affect plant phenotypes

Assessing the impact of variation in chloroplast and mitochondrial DNA (collectively termed the plasmotype) on plant phenotypes is challenging due to the difficulty in separating their effect from nuclear derived variation (the nucleotype). Haploid inducer lines can be used as efficient plasmotype donors to generate new plasmotype-nucleotype combinations (cybrids)(Ravi et al., 2014). We generated a panel comprising all possible cybrids of seven Arabidopsis thaliana accessions and extensively phenotyped these lines for 1859 phenotypes under stable and fluctuating conditions. We show that natural variation in the plasmotype results in additive as well as epistatic effects across all phenotypic categories. Plasmotypes which induce more additive phenotypic changes also cause more significant epistatic effects, suggesting a possible common basis for both additive and epistatic effects. On average epistatic interactions explained twice as much of the variance in phenotypes as additive plasmotype effects. The impact of plasmotypic variation was also more pronounced under fluctuating and stressful environmental conditions. Thus, the phenotypic impact of variation in plasmotypes is the outcome of multilevel Nucleotype X Plasmotype X Environment interactions and, as such, the plasmotype is likely to serve as a reservoir of variation which is only exposed under certain conditions. The production of cybrids using haploid inducers is a quick and precise method for assessing the phenotypic effects of natural variation in organellar genomes. It will facilitate efficient screening of unique nucleotype-plasmotype combinations to both improve our understanding of natural variation in nucleotype plasmotype interactions and identify favourable combinations to improve plant performance.

genetics↗