טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentGuttmann-Raviv Noga
SubjectThe Role of IME2 in Meiosis in the Yeast Saccharomyces
Cerevisiae
DepartmentDepartment of Biology
Supervisor Professor Emeritus Yona Kassir


Abstract

   Successful entry and completion of meiosis in the yeast Saccharomyces cerevisiae depends on three main regulators: the transcription factor - Ime1, a serine threonine protein kinase, Ime2, whose transcription is regulated by Ime1, and the cyclin dependent kinase - Cdc28.

   In the first part of this work we reexamined the role of Cdc28 and Ime2 in the meiotic pathway. Our results indicate that cdc28-degron cells initiate and progress through both premeiotic DNA replication and meiotic recombination, suggesting that Cdc28 is not essential for these events. We show that the delayed premeiotic DNA replication observed in the absence of Ime2 depends on Cdc28. We conclude that Ime2 and Cdc28 possess redundant functions in promoting premeiotic DNA replication. We demonstrate that the kinase activity of Ime2 and not only its physical existence is required for its role in premeiotic DNA replication. Using the two-hybrid system we show that Ime2 and Cdc6 - a key regulator of DNA replication, associate. These results suggest the Ime2 is directly involved with initiation of premeiotic DNA replication, and that in its absence, Cdc28 may substitute.

   In the second part of this work we demonstrate that Ime2 reduces the stability of Ime1, and that the kinase activity of Ime2 is required for this effect. We further show that Ime2 associates with Ime1, and that the ubiquitin/proteasome pathway degrades Ime1. We also show that Ime2 itself is an extremely unstable protein, whose expression in vegetative cultures is toxic. We propose that a negative feedback loop ensures that the activity of Ime1 will be confined to a narrow window, and that following activation of the meiotic transcriptional cascade, it will be degraded.