Replication origins are associated with transcription initiation sequences in the mitochondrial genome of yeast

Mitochondrial transcripts have been investigated in a series of spontaneous petite mutants of Saccharomyces cerevisiae endowed with mitochondrial genomes formed by short repeat units containing no genes, but either: (a) one of the seven ori sequences, the canonical origins of DNA replication (ori+ mutants); or (b) partially deleted ori sequences, lacking GC-rich clusters A or C (ori- mutants); or (c) no canonical ori sequence, but only oris sequences, the surrogate origins of replication ( orio mutants). The results indicate that some ori sequences play a role in transcription initiation, and that the presence of cluster C and, more specifically, of an AT-rich sequence next to it, are essential for transcription to take place. Hybridization experiments with separated DNA strands have identified the template strand used in transcription as the strand containing the oligopyrimidine stretch of cluster C. S1 degradation of RNA-DNA hybrids indicated that transcription initiates at a TATTACTTATATATTT sequence next to the oligopyrimidine stretch of cluster C and proceeds in the cluster C----cluster A direction. The relevance of these results for the transcription of the wild-type mitochondrial genome is discussed.