Related Experiment Videos
G1 cyclins CLN1 and CLN2 repress the mating factor response pathway at Start in the yeast cell cycle
1Rockefeller University, New York, New York 10021.
Abstract:
Transcriptional induction by the mating pheromone alpha-factor was monitored at different stages of the yeast cell cycle. G2/M-phase and pre-Start cells showed strong FUS1 mRNA induction, whereas in post-Start cells the signaling was reduced significantly. This reduction in signaling activity in post-Start cells was correlated with the presence of CLN1 or CLN2 transcripts and was not observed in synchronized cells lacking functional CLN1 and CLN2 genes. Activation of the Cln-Cdc28p kinase by overexpression of CLN2 from the GAL1 promoter strongly reduced FUS1 mRNA induction. CLN1 overexpression had a similar effect when the FAR1 gene, encoding a negative regulator of CLN1/2 function, was deleted. This reduction of pheromone signaling was specific for CLN1 and CLN2, as it was not observed when CLN3 was overexpressed. Inactivation of the Cln-Cdc28p kinase complex by thermal inactivation of temperature-sensitive Cdc28p prevented repression of FUS1 signaling. CLN2 overexpression suppressed the constitutive signaling and division-arrest phenotypes of cells with a disrupted gpa1 gene, indicating that the site of action for repression is downstream of the alpha-subunit (Gpa1p) of the heterotrimeric G protein. The repression at Start of pheromone signaling by Cln1-Cdc28p or Cln2-Cdc28p kinase complexes may contribute to the acquisition of pheromone resistance as cells execute Start.
Insights
Yeast cells reduce mating pheromone signaling after cell division begins, specifically when CLN1 or CLN2 genes are active. This cell cycle regulation helps yeast become resistant to pheromones during mating.
Area of Science:
- Molecular Biology
- Cell Biology
- Yeast Genetics
Background:
- Yeast mating pheromones trigger cellular responses essential for sexual reproduction.
- Cell cycle progression influences the sensitivity of yeast cells to external signals like mating pheromones.
Purpose of the Study:
- To investigate how yeast cell cycle progression affects transcriptional induction by the mating pheromone alpha-factor.
- To identify specific cell cycle regulators involved in modulating pheromone signaling pathways.
Main Methods:
- Monitoring FUS1 mRNA induction at different yeast cell cycle stages.
- Utilizing gene overexpression (CLN1, CLN2, CLN3) and deletion (FAR1, gpa1) strategies.
- Employing temperature-sensitive mutants (Cdc28p) to inactivate cell cycle kinases.
Main Results:
- FUS1 mRNA induction by alpha-factor was high in G2/M-phase and pre-Start cells but significantly reduced in post-Start cells.
- This reduction correlated with CLN1/CLN2 transcript presence and was dependent on functional CLN1 and CLN2 genes.
- Overexpression of CLN2 or CLN1 (in FAR1 deletion mutants) reduced pheromone signaling, while CLN3 overexpression had no effect.
- Inactivation of Cln-Cdc28p kinase prevented repression, and CLN2 overexpression suppressed phenotypes in gpa1 mutants, indicating repression occurs downstream of G-protein alpha-subunit.
Conclusions:
- The Cln1-Cdc28p and Cln2-Cdc28p kinase complexes actively repress mating pheromone signaling at the Start of the cell cycle in yeast.
- This repression mechanism likely contributes to the development of pheromone resistance as cells prepare for division.
- Cell cycle-dependent regulation of signaling pathways is crucial for coordinating cellular processes like mating and division.