Expression studies and promoter analysis of the nuclear gene for mitochondrial transcription factor 1 (MTF1) in yeast

The basal mitochondrial transcription apparatus of Saccharomyces cerevisiae consists of the core enzyme for mitochondrial RNA polymerase and the speci ficity factor. The core enzyme is homologous to those of bacteriophages T3, T7 and SP6 whereas the specificity factor shows similarities with bacteria l sigma factors. Recently it was shown that the bacteriophage-type core enz yme is widespread among the eukaryotic lineage and a common picture for the mitochondrial transcription apparatus in eukaryotic cells is now emerging. In contrast to the situation for the core enzyme, the gene for the specifi city factor has only been identified from S. cerevisiae and more recently f rom two other yeast species. As the specificity factor is the key component for initiation of transcription at the mitochondrial promoter we wanted to study in more detail gene expression, regulation, and the function of the promoter of the nuclear MTF1 gene. For this purpose the messenger RNA level for scMTF1 was investigated under a large number of different growth condi tions and thereby exhibited a very low, but regulated and carbon source-dep endent, expression. Deletion experiments identify the minimal promoter for functional complementation in yeast. To evaluate the functional conservatio n of the promoter elements the homologous MTF1 gene from the closely relate d yeast Saccharomyces douglasii was isolated and tested in heterologous com plementation experiments. In spite of a highly conserved protein sequence t hese studies demonstrate that at lowcopy number sdMTF1 is not able to subst itute for scMTF1 in S. cerevisiae. Promoter exchange experiments with MTF1 from S. cerevisiae and S. douglasii demonstrate that differences in gene ex pression are responsible for the failure in heterologous complementation. T his finding prompted us to compare the promoter regions of MTF1 from four d ifferent yeast species. For this purpose the sequences of the 5' regions fr om S. douglasii, S. kluyveri and Kluyveromyces lactis were determined. A co mparison of these sequences identifies significant differences and rapid ch anges in the intergenic regions, even between closely related yeast species .