Post-transcriptional and developmental regulation of a CMS-associated mitochondrial gene region by a nuclear restorer...

Transcripts of the mitochondrial gene region orf224/atp6, which is associated with the Polima or pol cytoplasmic male sterility (CMS) of Brassica napus, differ among fertile, sterile and nuclear-restored plants. We show here that the effects of the restorer gene Rfp on orf224/atp6 transcripts varies among different floral organs. Relative to monocistronic atp6 transcripts, levels of the dicistronic transcripts spanning orf224 and atp6 are dramatically reduced in petals, stamens and carpels, but not sepals, of restored flowers. In pol CMS plants, the relative levels of different orf224/atp6 transcripts are similar among the floral organs. Analysis of guanylyltransferase-labeled mtRNA indicates that only the dicistronic 2.2 and 1.9 kb orf224/atp6 transcripts carry an initiator 5' terminus; hence the 1.4 and 1.3 kb transcripts of restored plants, as well as the 1.1 kb atp6 transcript common to all genotypes, are generated by RNA processing and not de novo initiation. Although steady-state levels of dicistronic transcripts in flower buds are lower in restored than in sterile plants, run-on transcription experiments show that these transcripts are synthesized at the same rate in both types of flowers. These findings imply that the restorer gene acts by conditioning the removal of sequences from the 5' end of dicistronic transcripts in a developmentally regulated manner. Run-on transcription experiments indicate that the single 1.1 kb atp6 transcript of nap cytoplasm is also generated by removal of sequences from the 5' end of a precursor. We suggest that specific endonucleolytic cleavage of a precursor RNA, followed by non-specific 3' to 5' exonuclease action, may represent a common mechanism for tailoring transcripts in plant mitochondria.