Saccharomyces Cerevisiae RNA Research Articles

Purified Saccharomyces cerevisiae RNA polymerase II interacts homologously with two different promoters as revealed by P1 endonuclease analysis.

The intergenic region of the Saccharomyces cerevisiae GAL1-GAL10 divergent promoters has been circularized in vitro in different topological states. In defined conditions, purified homologous RNA polymerase II forms two stable complexes (half-life approximately equal to 5 h) with this DNA in the presence of the four ribonucleotides, as determined by measurement (Gamper and Hearst 1983) of the amount and stability of the resulting unwinding. Each stable complex induces in the closed DNA domain a region of hypersensitivity to P1 endonuclease. The two induced hypersensitive regions are very similar: each maps on one promoter, spans over the 100 bp DNA sequence that encompasses the RNA Initiation Sites (RIS) and the TATA box, is composed by three subregions (one on the RIS, one proximal or overlapping the TATA sequence, one intermediate). We show that this promoter-localized interaction is supercoil-dependent.

Electron-microscopic examination of the binding of a large RNA polymerase III transcription factor to a tRNA gene

A Saccharomyces cerevisiae RNA polymerase III transcription factor was previously shown to bind stably to tRNA genes. This transcription factor has been further purified on the basis of its large size and its binding to a S. cerevisiae tRNA 3 Leu gene has been examined by electron microscopy. Site-specific binding of the factor to the tRNA 3 Leu gene sharply bends the DNA.

Synthesis of two DNA-dependent RNA polymerases in yeast

Specific activities of Saccharomyces cerevisiae RNA polymerases I and II were measured in cells growing under different nutrient conditions and throughout the mitotic cell cycle. The specific activity of RNA polymerase I (possibly the ribosomal polymerase) does not vary during the yeast cell cycle. In contrast the specific activity of RNA polymerase II (messenger polymerase) increases during the first third of the cycle and thereafter declines. The independent regulation of synthesis of these two enzymes is further emphasised by observations on the response to different nutrient conditions. Shifting cells from minimal to rich medium led to enhanced RNA polymerase I activity but very little change in activity of RNA polymerase II. Furthermore the activity of RNA polymerase I varies directly with change in growth rate whereas the activity of RNA polymerase II is approximately constant over a range of growth rates. From this data it is suggested: ( i) The synthesis of these two enzymes is independently regulated; ( ii) RNA polymerase I is synthesised continuously throughout the cycle whereas RNA polymerase II is synthesised periodically early in the cell cycle.

Autonomy of mitochondria in Saccharomyces cerevisiae in their production of messenger RNA.

In ts(-)136, a temperature-sensitive mutant of Saccharomyces cerevisiae, nuclear and mitochondrial RNA production can be inhibited selectively by exposure to 36 degrees and ethidium bromide, respectively. Using the programming of mitochondrial polysomes, as measured by their ability to form nascent polypeptide chains, as an assay for functional messenger RNA, we have determined its response to temperature shifts and ethidium bromide. Only ethidium bromide produced a measurable effect; in contrast the cell-sap system responded exclusively to temperature shifts. We conclude that transcription of mitochondrial DNA is sufficient and that import of messenger RNA transcribed from nuclear chromosomes makes no measurable contribution to intramitochondrial protein synthesis.

Characterization of rapidly labelled rat liver ribonucleic acid showing high affinity for columns of methylated albumin on kieselguhr.

Most of the rapidly labelled RNA from rat liver submitted to column chromatography on methylated albumin on kieselguhr remains tightly bound to the column and can only be recovered by elution with m-ammonia. The tightly bound RNA is composed mainly of DNA-like RNA. The binding capacity is dependent not only on base composition but also on molecular size: the heavier RNA molecules show a greater affinity to the column than do the lower-molecular-weight components. Rapidly labelled mouse liver and Saccharomyces cerevisiae RNA show similar behaviour to rat liver RNA on columns of methylated albumin on kieselguhr.