Edward P. Winter

Signal transduction and MAPK pathways in yeast.

A large amount of signal transduction in eukaryotic cells depends on conserved kinase cascades that activate a family of enzymes called mitogen activated protein kinases (MAPKs). MAPK cascades are required for adaptive responses to changes in the environment, proliferative responses to mitogens, and differentiative responses that occur during embryogenesis. A basic understanding of MAPK signaling is essential for understanding the molecular basis of a wide range of diseases. There are multiple MAPK pathways in the yeast Saccharomyces cerevisiae that allow cells to respond to a variety of distinct environmental and developmental stimuli (see Ballard et al. 1991, EMBO J. 10:3752; Brewster et al. 1993, Science 259:1760; Krisak et al. 1994, Genes and Dev. 8:2151 for work from this laboratory describing different yeast MAPK signaling pathways). This laboratory is studying the basic molecular mechanisms that transduce signals in these pathways, as well as cross-regulatory mechanisms that enable one MAPK pathway to control a parallel pathway (see Hall et al. 1996, Mol. Cell Bio. 16:6715, for a further discussion). We are particularly interested in using yeast as a model system to study the role of MAPKs in regulating developmental programs.


The SMK1 MAPK pathway and meiotic development.

Meiotic development (sporulation) in yeast represents an excellent model system with which to study developmental processes. Similar to differentiation programs in higher eukaryotes, induction is controlled by a combination of cell-type and environmental signals. Once initiated, a transcriptional program is induced that ultimately leads to a cell (or spore) that is both genetically and biochemically distinctfrom its precursor. We have previously identified the SMK1 MAPK that is expressed specifically during sporulation and that is required for the differentiation program (spore morphogenesis) that immediately follows meiotic chromosome segregation (Krisak et al., 1994 Genes and Dev., 8:2151). Analysis of a collection of mutants in the pathway shows that the SMK1 MAPK plays a central role in regulating multiple steps in spore morphogenesis (Wagner et al., 1999 Genetics 151:1327). Genetic approaches with the smk1 mutants have shown that the Cak1 protein kinase activates the SMK1 pathway (Wagner et al., 1997 EMBO J., 16:1305). CAK1 encodes the major cyclin dependent kinase activating kinase and is required for mitotic cell cycle progression and for meiotic chromosome segregation. These observations suggest that Cak1 plays a key role in coordinating meiotic chromosome segregation with the SMK1 pathway. Current work is focused on testing this hypothesis. We are also using genetic, molecular and biochemicalapproaches to identify additional signal transducers as well as downstream targets of the SMK1 MAP kinase pathway.


The Pachytene checkpoint and transcriptional regulators.

A key event in meiotic development in all organisms is genetic recombination. The initiating event in genetic recombination is the formation of double stranded breaks (DSBs) in DNA. Certain yeast mutants that are unable to complete recombination accumulate DSBs. In these mutants a checkpoint signaling pathway blocks meioticprogression (the pachytene checkpoint). Analysis of the SMK1 promoter has identified a transcriptional regulatory pathway that represses meiosis-specific genes in mitotic cells (Pierce et al. 1998, Mol. Cell. Biol., 18:5970). Genetic experiments have identified key components of this pathway including the Sum1 DNA binding protein and Hst1, proteins previously implicated in chromatin silencing (Xie et al. 1999, EMBO J., 18:6448). Our results show that the Sum1 transcriptional repressor is required for the pachytene checkpoint (Lindgren et al. in press). Thus, cells lacking Sum1 progress through meiotic development in the presence of broken chromosomes (recombination intermediates). Furthermore, Sum1 is proteolytically destroyed during prophase in wild-type cells but not in checkpoint arrested cells. These results indicate that Sum1 is a key target of the pachytene checkpoint required to block meiotic progression in the presence of broken chromosomes. These results also link checkpoint pathways withtranscriptional programs and suggest that developmentally regulated proteolysis plays a key role in these processes. Current work is focused on elucidating the molecular mechanisms that link Sum1 proteolysis with checkpoint signaling. The evolutionaryconservation of mechanisms used to regulate cell cycle progression and differentiation programs suggest that the insights from these projects will be applicable to higher eukaryotes including humans.