Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress

Verona, Italy - 24/27 September, 2003

ISBN 88-900622-4-X

Poster Abstract - 5.29

MITOCHONDRIAL DNA VARIABILITY IN RPL5-RPS14 GENOME REGION OF SOLANUM SPP. INVOLVES MULTIPLE RECOMBINATION EVENTS

N. SCOTTI*, L. MARECHAL-DROUARD**, T. CARDI*

*) CNR-IGV, Institute of Plant Genetics Res. Div. Portici, Via Università 133, 80055 Portici (NA), Italy

**) IBMP, Institut de Biologie Moleculaire des Plantes du CNRS, 12 rue du General Zimmer, 67084 Strasbourg cedex, France

potato, somatic hybrids, mitochondrial DNA, homologous recombination

The mitochondrial genome of higher plants shows much more variation in size and structure than any other genome type investigated. This variability is associated with a high frequency of DNA recombination across homologous repeated sequences of variable size. Somatic hybridisation is a powerful method to produce plants with new combinations of nuclei, plastids and mitochondria. Mitochondrial DNA (mtDNA) in somatic or cytoplasmic hybrids (cybrids) often contains regions contributed by both fusion partners as well as novel fragments not seen in parents.

Southern analysis of Solanum commersonii, S. tuberosum and their somatic hybrids (SH) with rpl5 and rps14 gene probes detected polymorphisms between parental genotypes, and a novel fragment in eight somatic hybrids. This novel fragment could be generated by inter-parental recombination and/or by a selective amplification of pre-existing substoichiometric sequence arrangements. In order to understand the origin of this new fragment, several molecular analyses (cloning, sequencing and PCR) were carried out on mtDNA of parents and two somatic hybrids SH9A and SH9B, regenerated from the same callus, and male fertile and male sterile, respectively. PCR analysis showed that the S. commersonii rpl5-rps14 arrangement, is present at substoichiometric level on S. tuberosum mtDNA. Furthermore, in addition to the two stoichiometric rpl5-rps14 arrangements, SH9A mtDNA contains the third new substoichiometric rpl5-rps14 region.

The stoichiometric and substoichiometric rpl5-rps14 arrangements detected in the mitochondrial genome of the SH9A somatic hybrid and a computer-assisted sequence similarity analysis strengthened our hypothesis that the recombinant molecules we observed could be the result of inter-parental recombination. The sequence similarity searches detected upstream to rpl5 gene a short repeat of 7bp (R1) involved in a rare recombination event found in a single wheat regenerant (Hartmann et al. 1994), whereas revealed a break point of homology 161 bp (R2) downstream of the rps14 gene (rps14-cob intergenic region) in S. commersonii.

From the above findings, we propose two models that explain the mechanism responsible of the generation of the different rpl5-rps14 arrangements showed in SH9A. The models and the experimental results demonstrate how somatic hybridisation and resultant mitochondrial fusion may enhance mitochondrial recombinations between short repeats or activate recombination between non-recombinogenic sequence. In addition, we showed that in our somatic hybrids the rearrangements observed are not random; in fact the breakpoints resulting from fusion, upstream and downstream rpl5 and rps14 genes, respectively, were located near the evolutionary breakpoints of the Angiosperms (Quiñones et al. 1996) suggesting that the rpl5-rps14 can be defined as a “hot spot” of recombination in the mitochondrial genome of higher plants.

References

Hartmann C., Récipon H., Jubier M.F., Valon C., Delcher-Besin, E., Henry Y., De Buyser J., Lejeune B., Rode A. (1994) Curr Genet, 25, 456-464

Quiñones V., Zanlungo S., Moenne A., Gómez I., Holuigue L., Litvak S. , Jordana X. (1996) Plant Mol Biol, 31, 937-943