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rRNA transcription initiation is decreased by inhibitors of the yeast cell cycle control step "start"
L M Veinot-Drebot1, R A Singer, G C Johnston
1Department of Microbiology, Dalhousie University, Halifax, Nova Scotia, Canada.
Abstract:
Inhibitors of the "start" regulatory step in the cell cycle of the yeast Saccharomyces cerevisiae are known by indirect studies to perturb RNA metabolism. We have investigated these effects further and show here by a pulse-labeling and quantitative hybridization procedure that pre-rRNA transcription was substantially decreased by five inhibitors of start but was transiently stimulated by the mating pheromone alpha-factor. Thus in contrast to the effects of the other start inhibitors, the inhibition of start by alpha-factor is unrelated to this aspect of biosynthetic activity. Mating factor treatment also stimulated the synthesis rate of poly(A)+ RNA. The start inhibitors o-phenanthroline and L-ethionine inhibited pre-rRNA transcription with little effect on poly(A)+ RNA synthesis rates. Northern analysis showed that all inhibitors of start also inhibited pre-rRNA transcript cleavage, a process that has been dissociated from the inhibition of start. Most inhibitors also affected ATP pool size. One inhibitor, o-phenanthroline, markedly induced the general control response.
Insights
Start inhibitors in yeast cell cycle impact RNA metabolism. While most decrease pre-ribosomal RNA (rRNA) transcription, alpha-factor transiently stimulates it, affecting poly(A)+ RNA synthesis differently.
Area of Science:
- Molecular Biology
- Yeast Genetics
- Cell Cycle Regulation
Background:
- The
- start
- regulatory step in the yeast Saccharomyces cerevisiae cell cycle is known to influence RNA metabolism.
- Previous indirect studies suggested a link between start inhibition and RNA processing.
- This study provides direct experimental evidence to elucidate these effects.
Purpose of the Study:
- To investigate the direct effects of start inhibitors on RNA metabolism in Saccharomyces cerevisiae.
- To differentiate the impact of various start inhibitors, including alpha-factor, on pre-ribosomal RNA (rRNA) and poly(A)+ RNA synthesis.
- To explore the relationship between start inhibition, pre-rRNA transcript cleavage, and ATP pool size.
Main Methods:
- Pulse-labeling and quantitative hybridization techniques were employed to assess RNA synthesis rates.
- Northern blot analysis was used to examine pre-rRNA transcript cleavage.
- ATP pool size was measured, and the general control response was monitored.
Main Results:
- Five inhibitors of the start regulatory step substantially decreased pre-rRNA transcription.
- Mating pheromone alpha-factor transiently stimulated pre-rRNA transcription and also increased poly(A)+ RNA synthesis rate.
- o-phenanthroline and L-ethionine inhibited pre-rRNA transcription with minimal impact on poly(A)+ RNA synthesis.
- All tested start inhibitors reduced pre-rRNA transcript cleavage.
- Most inhibitors altered ATP pool size, and o-phenanthroline induced the general control response.
Conclusions:
- The effects of start inhibitors on RNA metabolism are diverse, with alpha-factor exhibiting a distinct response compared to other inhibitors.
- Inhibition of the start step is linked to reduced pre-rRNA transcription and cleavage, but alpha-factor's effect is not solely tied to this.
- Start inhibitors can impact cellular energy levels (ATP) and activate stress responses like the general control pathway.