Leveraging dominant-negative histone H3 K-to-M mutations to study chromatin during differentiation and development

Abstract

Histone modifications are associated with regulation of gene expression that controls a vast array of biological processes. Often, these associations are drawn by correlating the genomic location of a particular histone modification with gene expression or phenotype; however, establishing a causal relationship between histone marks and biological processes remains challenging. Consequently, there is a strong need for experimental approaches to directly manipulate histone modifications. A class of mutations on the N-terminal tail of histone H3, lysine-to-methionine (K-to-M) mutations, was identified as dominant-negative inhibitors of histone methylation at their respective and specific residues. The dominant-negative nature of K-to-M mutants makes them a valuable tool for studying the function of specific methylation marks on histone H3. Here, we review recent applications of K-to-M mutations to understand the role of histone methylation during development and homeostasis. We highlight important advantages and limitations that require consideration when using K-to-M mutants, particularly in a developmental context.

Keywords: Chromatin; Differentiation; Epigenetics; Histone modification; Stem cell.

Fig. 1.

H3K4, H3K9, H3K27 and H3K36 are modified by multiple enzymes and are associated with transcriptional regulation. Multiple methyltransferases (light blue) and demethylases (black) deposit and erase methylation, respectively. H3K4 and H3K36 methylation are typically associated with transcriptional activation (green). H3K9 and H3K27 methylation are associated with transcriptional repression (red). Asterisks indicate putative methyltransferases based on biochemistry data. Jarid1a, also known as Kdm5a; Jarid2b, also known as Jarid2; Jarid1c, also known as Kdm5c; Jarid1d, also known as Kdm5d; Jhdm1d, also known as Kdm7a; No66, also known as Riox1.

Fig. 2.

Histone H3 K-to-M mutants bind cognate methyltransferases or demethylases. (A) Human EZH2 methyltransferase (cyan) and H3K27M peptide (magenta, mutant M residue shown in green) with S-adenosyl-l-homocysteine (SAH; yellow). Structure deposited with the Protein Data Bank (PDB) under accession code 5HYN (Justin et al., 2016). (B) Human SETD2 methyltransferase (cyan) and H3K36M peptide (magenta, mutant M residue shown in green) with SAH (yellow). Structure deposited with the PDB under accession code 5V22 (Zhang et al., 2017). (C) Human EHMT2 methyltransferase (cyan) and H3K9M peptide (magenta, mutant M residue shown in green) with S-adenosyl-l-methionine (SAM; yellow). Structure deposited with the PDB under accession code 5T0K (Shan et al., 2016). (D) Human LSD1 (cyan) and H3K4M mutant peptide (magenta, mutant M residue shown in green) with flavin adenine dinucleotide (FAD; yellow). Structure deposited with the PDB under accession code 2V1D (Forneris et al., 2007).