Cloning and characterization of the Neurospora crassa cyt-5 gene. A nuclear-coded mitochondrial RNA polymerase with a...

The Neurospora crassa mutants, cyt-5-1 and cyt-5-4 have a cytochrome b- and aa3-deficient phenotype, suggesting that they result from a deficiency in a nuclear-coded component of the mitochondrial gene expression apparatus (Bertrand, H., Nargang, F. E., Colllins, R. A., and Zagozeski, C. A. (1977) Mol. Gen. Genet. 153,247-257). The complementing wild-type gene has been cloned and and shown to encode a protein with significant sequence similarity to Saccharomyces cerevisiae mitochondrial RNA polymerase and bacteriophage RNA polymerases. There are remarkable differences between the N. crassa protein and its yeast homologue, including a region of very little homology near the N termini of the two gene products. The cyt-5 gene encodes a stretch of polyglutamine in this region of uniquesequence. In addition, an acidic insertion (86 amino acids, of which 24 are Asp or Glu and 10 are Arg or Lys) is present near the C terminus of the cyt-5 gene product. Transcript levels of the cytochrome b and cytochrome oxidase subunit III genes are severely reduced in cyt-5 mutants, suggesting a likely mechanism for the cytochrome-deficient phenotype. In contrast, mitochondrial rRNAs accumulate to nearly normal levels in cyt-5 mutants. However, mitochondrial rRNA levels are not indicative of the rate of transcription of the corresponding genes, since crude lysates of mitochondria from cyt-5 mutants exhibit greatly reduced transcriptional activity with a 19 S rRNA promoter. The cyt-5 gene is flanked by at least one gene whose product also may be involved in mitochondrial function.