Abstract
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.
Highlights
The covalent and reversible modification of histones allows cells to establish stable and heritable patterns of gene expression without any changes to their genetic blueprint
Through a series of titration measurements, we found that using an MBP antibody and a chemiluminescence based readout produces a very limited linear response, which precludes us from reporting an apparent Kd
Cells maintain high H3K9me2 levels in the presence of Gal4-Clr3 before and after tetracycline (Figure 7C-D). These results suggest that constitutively tethering Clr3 to sites of heterochromatin formation is sufficient to oppose Epe1 activity resulting in maintenance of H3K9 methylation even after +tetracycline addition
Summary
The covalent and reversible modification of histones allows cells to establish stable and heritable patterns of gene expression without any changes to their genetic blueprint. Point mutations of amino acid residues involved in Fe (II) or a-ketoglutarate binding affect Epe activity in cells and lead to heterochromatin spreading beyond normal boundary sequences (Trewick et al, 2007) These conflicting lines of biochemical and genetic data have prompted several alternative explanations for how Epe might fulfill its anti-silencing role. These models include the possibility that Epe acts as a protein hydroxylase which targets non-histone proteins such as Swi6HP1, regulates the activity of the multi-subunit H3K9 methyltransferase CLRC complex, or functions as an H3K9 demethylase when in complex with Swi6HP1 (Aygun et al, 2013; Iglesias et al, 2018; Trewick et al, 2007; Zofall and Grewal, 2006). Our work highlights the versatile, non-canonical ways in which histone demethylases can oppose establishment and maintenance of epigenetic states
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