Ubiquitin and ubiquitine-like systems in Saccharomyces cerevisiae

Abstract

Ubiquitin and ubiquitin-like modifiers are small proteins that exist in all eukaryotes, from yeast to humans. Ubiquitin(-like) modifiers can be coupled to other proteins, which is mediated specific combinations of enzymes. The attachment of a ubiquitin(-like) modifier to a protein is important for the regulation of a large variety of cellular processes, such as cell division, intracellular transport, metabolism, transcription and DNA repair. However, for many of these enzymes it is still unclear how they cooperate and which cellular processes they regulate. In this thesis we describe how we use microarray expression profiling of yeast mutants (S. cerevisiae) to gain insight into the functional relationships between all the ubiquitin(-like) system components. We have performed a large-scale microarray study of 224 yeast mutants, which are defective for specific components of the ubiquitin(-like) systems. For each individual mutant we established the mRNA expression changes on genome-wide scale. Half of the mutants display a highly specific expression phenotype, which is representive for at least twenty different cellular processes. One of these processes is the degradation of ubiquitinated proteins by the proteasome, also known as the ubiquitin-proteasome system (UPS). We have identified a highly specific mRNA expression signature for mutants with a UPS defect. This expression signature is used to screen a large collection a yeast mutants and identifies many novel mutants with a potential UPS defect. The expression changes of all the mutants is assembled into a large collection of expression phenotypes that is used to identify known and novel functional relationships between all the different ubiquitin(-like) system components. We are able to uncover various signaling pathways between different ubiquitin(-like) system components and many protein complexes. One unknown protein complex, consisting of the proteins Slx5 and Slx8, was further studied in detail. We reveal a new function for Slx5/8 in maintaining genome stability by contributing to accurate chromosome segregation in mitosis. Moreover, this function is evolutionarily conserved in humans. This shows that gene expression analysis is a powerful method to uncover new functions for ubiquitin(-like) system components.