MBT510/GENET551 Berg

Paper for 6 Dec 2001

Stevenson, B. J., Rhodes, N., Errede, B. and Sprague, G. F., jr. 1992. Constitutive mutants of the protein kinase STE11 activate the yeast pheromone response pathway in the absence of the G protein. Genes Devel. 6: 1293-1304.

This paper employs a powerful method for identifying new genes in a pathway. Many interesting but sometimes subtle processes are evident from these studies. Try your best to understand their results. We will emphasize the logic of the approach, its strengths and weaknesses. Homework: Write brief answers to the questions in bold.

Questions for Thought

1) What was known at the start of this work? Summarize the known pathway. What is the evidence that STE5, STE11, STE7, FUS3, KSS1 and STE12 act downstream of SCG1/STE4/STE18? What other ways could you test these relationships?

2) What is the goal of these experiments? How do they approach the problem? (i.e., give method and logic.)

3) Table 1: How do they characterize their mutants? Why do they perform these tests?

4) The authors use a two-step protocol to identify the genes mutated in their screen. What is the logic of their approach? THINK THROUGH THIS SECTION CAREFULLY. We will emphasize these studies in our discussion. What is their result? How do they interpret it? How do they confirm it?

5) Analyses of the STE11-1 and STE11-4 alleles in a defined background reveal that these two mutations produce different phenotypes. Why would one activating mutation produce a stronger phentoype than another? What is different about the two alleles?

6) The authors propose two hypotheses to explain the quantitative differences in suppression of ste4 D and ste5 D by STE11-1 and STE11-4. What experiments would you do to distinguiush these possibilities? Why would a gain of function mutation require an UPSTREAM gene for activity? Is there any other evidence to support one or the other hypothesis?

7) Summarize the evidence that Ste11p acts as a kinase upstream of Ste7p, Fus3p, Kss1p, and Ste12p. If STE7, KSS1 and FUS3 encode kinases also, why didn’t the authors obtain suppressor mutations in these genes during their screen?