Abstract
HP1 proteins are essential for establishing and maintaining transcriptionally silent heterochromatin. They dimerize, forming a binding interface to recruit diverse chromatin-associated factors. HP1 proteins are specialized and rapidly evolve, but the extent of variation required to achieve functional specialization is unknown. The fission yeast S. pombe has two HP1 homologs, Swi6 and Chp2, that have essential, nonredundant functions in heterochromatin regulation. Here, we investigate how changes in amino acid sequence impacts epigenetic inheritance by performing a targeted mutagenesis screen of the S. pombe HP1 homolog, Swi6. We identify substitutions within an auxiliary surface adjacent to the HP1 dimerization interface produce Swi6 variants with divergent maintenance properties. Remarkably, substitutions at a single amino acid position led to the persistent gain or loss of epigenetic inheritance. We find these substitutions increase Swi6 chromatin occupancy in vivo and altered Swi6-protein interactions that reprogram H3K9me maintenance. We present several biochemical approaches to characterize Swi6 and Chp2 biophysical properties in vitro. We observe differences in how these proteins bind to chromatin in vitro and in vivo. We show relatively minor changes in Swi6 amino acid composition can lead to profound changes in epigenetic inheritance which provides a redundant mechanism to evolve effector specificity. The findings in this work lay the groundwork for future studies in HP1 proteins and epigenetic inheritance.