A novel pea mitochondrial in vitro transcription system recognizes homologous and heterologous mRNA and tRNA promoters.

To elucidate the mechanism involved in the transcription initiation process in mitochondria of dicotyledonous plants, an in vitro transcription system was established for pea (Pisum sativum L.). The partially purified mitochondrial protein extract initiates transcription on homologous pea templates as well as on heterologous mitochondrial DNA from other dicot plant species. In vitro transcription begins within the nonanucleotide 5'-(-7)CRTAAGAGA(+2)-3' (transcription start site is underlined) conserved at most of the identified transcription initiation sites in dicot plant mitochondria. The in vitro initiation at promoters of protein as well as of tRNA coding genes indicates a common mode of transcription initiation for different types of RNA. The competent recognition of different heterologous templates supports a general functional role of the conserved nonanucleotide within mitochondrial promoters of dicotyledonous plants. Initial studies of the promoter structure by deletion analysis in the 5' region of the pea atp9 promoter show that in addition to the conserved nonanucleotide, which is essential for transcription initiation in vitro, sequences up to 25 nucleotides upstream of the transcription start site are necessary for an efficient initiation event.