Transcription and RNA-processing in fission yeast mitochondria

Abstract

We systematically examined transcription and RNA-processing in mitochondria of the petite-negative fission yeast Schizosaccharomyces pombe. Two presumptive transcription initiation sites at opposite positions on the circular-mapping mtDNA were confirmed by in vitro capping of primary transcripts with guanylyl-transferase. The major promoter (P ma ) is located adjacent to the 5-end of the rnl gene, and a second, minor promoter (P mi ) upstream from cox3. The primary 5-termini of the mature rnl and cox3 transcripts remain unmodified. A third predicted accessory transcription initiation site is within the group IIA1 intron of the cob gene (cobI1). The consensus promoter motif of S. pombe closely resembles the nonanucleotide promoter motifs of various yeast mtDNAs. We further characterized all mRNAs and the two ribosomal RNAs by Northern hybridization, and precisely mapped their 5-and 3-ends. The mRNAs have leader sequences with a length of 38 up to 220 nt and, in most instances, are created by removal of tRNAs from large precursor RNAs. Like cox2 and rnl, cox1 and cox3 are not separated by tRNA genes; instead, transcription initiation from the promoters upstream from rnl and cox3 compensates for the lack of tRNA-mediated 5-processing. The 3-termini of mRNAs and of SSU rRNA are processed at distinct, C-rich motifs that are located at a variable distance (1-15 nt) downstream from mRNA and SSU-rRNA coding regions. The accuracy of RNAprocessing at these sites is sequence-dependent. Similar 3-RNA-processing motifs are present in species of the genus Schizosaccharomyces, but not in budding yeasts that have functionally analogous A+T-rich dodecamer processing signals.

The conserved Wobble uridine tRNA thiolase Ctu1–Ctu2 is required to maintain genome integrity

“…In addition to plasmids harboring the wild-type gene, we identified plasmids that partially or completely suppressed the phenotype and allowed growth at 36°C in both strains. They all contained fragments of the mitochondrial genome, which was surprising because mitochondrial DNA transcription depends on a T3/T7 phage-like polymerase unrelated to the nuclear RNA polymerase II (17). Moreover, the suppressing fragments overlapped only at a severely truncated version of subunit 6 of the ATPase (Fig.…”

Transcription of the Dictyostelium discoideum mitochondrial genome occurs from a single initiation site

“…Considering the position of the tRNA genes, which are located in the vicinity of the maturation sites at the RNA level, it can be assumed that the processing of the precursor is, at least in part, achieved by the release of tRNA molecules. The excision of tRNAs has been described as an RNA maturation mechanism in a variety of organisms, including animals, fungi, and green algae (Ojala et al 1980(Ojala et al , 1981Burger et al 1985;Dyson et al 1989;Schäfer et al 2005;Wolff and Kück 1996).…”

“…These include a high number of genes (33 protein-encoding genes, six open reading frames, two ribosomal RNA genes, and 18 transfer RNA genes), a high A + T content (72.6%), a small number of noncoding regions, and the presence of large and small subunit ribosomal protein genes, which are arranged in a bacterial-like gene cluster (Ogawa et al 2000;Barth et al 2007). Compared with this, mitochondrial genomes in all multicellular organisms, especially in animals (<20 kb) (Wolstenholme 1992) but also in some fungi (Schizosaccharomyces pombe, 19.4 kb) (Paquin et al 1997;Schäfer et al 2005) and some green (Chlamydomonas reinhardii, 15.8 kb) (Michaelis et al 1990) and red algae (Chondrus crispus, 25.8 kb) (Leblanc et al 1995) appear to be much smaller and more compact mitochondrial DNAs. These smaller mitochondrial genomes are commonly transcribed from either single, bidirectional promoters or from two or more unidirectional promoters into at least two polycistronic transcription units (Bogenhagen and Romanelli 1988;L'Abbe et al 1991;Shadel and Clayton 1993).…”

“…Less-derived organisms tend to have larger mitochondrial genomes, which, due to their size require multiple, if not many promoters for the expression of the more complex genomes. In fungi, the size of the mitochondrial genomes varies from 19.4 kb in Schizosaccharomyces pombe to more than 80 kb in Saccharomyces cerevisiae (Schäfer et al 2005). Depending on the genome size, the number of promoters ranges from two to multiple and a simple consensus sequence consisting of a nine-nucleotide sequence ATA-TAAGTA has been shown to be sufficient for accurate transcription initiation (Schinkel et al 1986;Costanzo and Fox 1990).…”

Functional and Regulatory Profiling of Energy Metabolism in Fission Yeast

“…Mitochondria of fission yeast form a dynamic network along microtubules which mediate their inheritance, as is the case in multicellular eukaryotes [19]. The fission yeast mitochondrial genome is compact (~20 kb, 11 proteincoding genes) and mitochondrial RNA processing is similar to in animal cells [20]. Fission yeast can grow using either respiration or fermentation but, in contrast to budding yeast, does not thrive in strictly anaerobic conditions [21].…”