H3K9 methylation (H3K9me) specifies the establishment and maintenance of transcriptionally silent epigenetic states or heterochromatin. The enzymatic erasure of histone modifications is widely assumed to be the primary mechanism that reverses epigenetic silencing. Here, we reveal an inversion of this paradigm where a putative histone demethylase Epe1 in fission yeast, has a non-enzymatic function that opposes heterochromatin assembly. Mutations within the putative catalytic JmjC domain of Epe1 disrupt its interaction with Swi6HP1 suggesting that this domain might have other functions besides enzymatic activity. The C-terminus of Epe1 directly interacts with Swi6HP1, and H3K9 methylation stimulates this protein-protein interaction in vitro and in vivo. Expressing the Epe1 C-terminus is sufficient to disrupt heterochromatin by outcompeting the histone deacetylase, Clr3 from sites of heterochromatin formation. Our results underscore how histone modifying proteins that resemble enzymes have non-catalytic functions that regulate the assembly of epigenetic complexes in cells.

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H3K9 methylation (H3K9me) marks transcriptionally silent genomic regions called heterochromatin. A conserved class of HP1 proteins are critically required to establish and maintain heterochromatin. HP1 proteins bind to H3K9me, recruit factors that promote heterochromatin formation, and oligomerize to form phase-separated condensates. We do not understand how HP1 protein binding to heterochromatin establishes and maintains transcriptional silencing. Here, we demonstrate that the S.pombe HP1 homolog, Swi6, can be completely bypassed to establish silencing at ectopic and endogenous loci when an H3K4 methyltransferase, Set1 and an H3K14 acetyltransferase, Mst2 are deleted. Deleting Set1 and Mst2 enhances Clr4 enzymatic activity, leading to higher H3K9me levels and increased spreading. In contrast, Swi6 and its capacity to oligomerize were indispensable during epigenetic maintenance. Our results demonstrate the role of HP1 proteins in regulating histone modification crosstalk during establishment and identifies a genetically separable function in maintaining epigenetic memory.

The covalent modification of histones establishes distinct phenotypic or epigenetic states without altering the underlying genetic code. This process allows cells to respond to acute changes in cellular homeostasis resulting from alterations in intracellular, environmental, or metabolic conditions. The consequences of post‐translational modifications of histones are multifaceted, leading to the emergence of chemotherapy resistance in cancers, drug resistance in fungal pathogens and temperature‐dependent initiation of flowering in plants. One modification, histone H3 lysine 9 methylation (H3K9me), is associated with heterochromatin formation, and under conditions of acute stress, heterochromatin relocates to silence novel genes that enhance cellular fitness. It is still a mystery how cells activate this adaptive pathway and mark specific genes with H3K9me to successfully adapt to adversity. We find that, in S. pombe cells, adaptive H3K9me is preceded by activation of a global stress response and downregulation of ribosome biogenesis machinery. Our results indicate that aberrant silencing of the ribosomal DNA repeats triggers adaptation via H3K9me and allows cells to respond to complex stresses by developing novel heritable phenotypes. My completed work provides me unprecedented temporal resolution into H3K9me adaptation and sets the foundation for investigation of more complex questions involving the H3K9me adaptive process.