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Methylation of histone proteins plays a crucial role controlling genome activity. To understand how the responsible histone methyltransferases are regulated it is important to know their fundamental biochemical properties within the cell and relate these to cellular methylation dynamics. Repeated experiment-modeling cycles have led to insights into the in vivo dynamics of methylation of lysine 79 on histone H3 (H3K79) by the methyltransferase Disruptor of Telomeric Silencing 1 (Dot1). Genetic perturbation in yeast, quantitative measurements, and computational modeling were combined to show that Dot1 employs an uncommon, distributive methylation mechanism. A steady-state in vivo methylation model using this information has provided validated explanations for methylation defects in mutants. Subsequent single-cell models have provided insights into the dynamics of H3K79 methylation throughout the cell cycle and uncovered a role for histone protein aging. These integrated modeling approaches will aid in understanding how regulatory mechanisms influence Dot1’s role in gene expression, cell cycle progression, and cancer and can be applied to other methylation systems.

Publisher Summary Silencing in Saccharomyces cerevisiae is a chromatin-mediated repression of genes located within specific chromosomal domains. Each time chromosomal DNA replicates, silent chromatin must be recreated by an orchestrated series of assembly processes and events. Mutations affecting DNA replication, cell cycle progression, DNA repair, chromatin assembly, histone modification, telomere length, protein turnover, or cellular metabolism modulate silencing, demonstrating that many cellular pathways are connected to the process. A loss of silencing, particularly at the silent mating loci, also has its consequences. Absence of silencing in a haploid cell triggers a cascade of events that cause the cell to act as if it were a diploid. As a result of its intimate ties to overall cellular physiology and processes, and the fact that silent chromatin is found in all eukaryotes, silencing has become a well-studied aspect of yeast biology. This chapter focuses on genetic and cellular assays to monitor gene silencing in yeast. In addition, strains and vectors most commonly used in the assays are described with their advantages and limitations. The chapter provides an overview is given of assays that can facilitate identification of new silencing components.