Stoichiometric shifts in the common bean mitochondrial genome leading to male sterility and spontaneous reversion to fertility

The plant mitochondrial genome is characterized by a complex, multipartite structure. In cytoplasmic male-sterile (CMS) common bean, the sterility-inducing mitochondrial configuration maps as three autonomous DNA molecules, one containing the sterility-associated sequence pvs-orf239. We constructed a physical map of the mitochondrial genome from the direct progenitors to the CMS cytoplasm and have shown that it maps as a single, circular master configuration. With long-exposure autoradiography of DNA gel blots and polymerase chain reaction analysis, we demonstrate that the three-molecule CMS-associated configuration was present at unusually low copy number within the progenitor genome and that the progenitor form was present substoichiometrically within the genome of the CMS line. Furthermore, upon spontaneous reversion to fertility, the progenitor genomic configuration as well as the molecule containing the pvs-orf239 sterility-associated sequence were both maintained at substoichiometric levels within the revertant genome. In vitro mitochondrial incubation results demonstrated that the genomic shift of the pvs-orf239-containing molecule to substoichiometric levels upon spontaneous reversion was a reversible phenomenon. Moreover, we demonstrate that substoichiometric forms, apparently silent with regard to gene expression, are transcriptionally and translationally active once amplified. Thus, copy number suppression may serve as an effective means of regulating gene expression in plant mitochondria.