Quantitative Proteomics Reveals Histone Modifications in Crosstalk with H3 Lysine 27 Methylation

Abstract

Methylation at histone H3 lysine 27 (H3K27me) is an evolutionarily conserved epigenetic mark associated with transcriptional repression and replication elongation. We have previously shown that in Tetrahymena thermophila, a unicellular eukaryote, the histone methyltransferases (HMTs) TXR1 and EZL2 are primarily responsible for H3K27 mono-methylation (H3K27me1) and di-/tri-methylation (H3K27me2/3), respectively. In eukaryotic cells, nuclear DNA wraps around an octamer of two copies each of the four core histones H2A, H2B, H3, and H4, to form the nucleosome, the basic unit of chromatin. Many post-translational modifications (PTMs) 1 , including methylation, acetylation, phosphorylation, biotinylation, citrullination, ADP-ribosylation, and ubiquitylation, occur at numerous sites on histones (1). Dynamic changes in the chromatin structure and architecture, including the switch between condensed and decondensed states as well as interactions with a wide range of protein complexes, are modulated by these PTMs, deposited by histone modifying enzymes in a combinatorial pattern that is still being actively deciphered (2, 3). Reflecting its crucial role in DNA-mediated transactions, the functionality of chromatin is relatively robust, tolerating substantial mutations in histone residues carrying PTMs as well as histone modifying enyzmes (4). The robustness is generally attributed to redundant or parallel roles played by many histone modifications, which often adapt compensatory changes to these perturbations. However, details of the crosstalk among histone PTMs and their physiological impacts are still incomplete (5). Deposited by histone methyltransferases (HMTs) and removed by demethylases, histone lysine methylation (mono-, di-, and tri-methylated forms) is a dynamic epigenetic mark (6 -8). Boosted by the identification and characterization of the enzymatic machinery, histone lysine methylation has been the target of considerable research efforts, reflecting its key role in modulating histone functions. Unlike acetylation, methylation does not directly affect histone/DNA interactions by altering the net charge of histones, and its physiological impacts depend on the exact form and site of modification, as well as the effector proteins recognizing them (9).The evolutionarily conserved H3K27 methylation has been traditionally associated with heterochromatin formation and transcriptional repression (10). Enhancer of zeste (E(z)), initially identified by its role in Polycomb repression, is the HMT required for H3K27 methylation in Drosophila (11-13). E(z) homologs are found in protozoa, metazoa, and plants, though they are conspicuously missing in fungi. All three forms of H3K27 methylation (H3K27me1/2/3) are catalyzed by E(z) homologs in metazoa (14). However, in protozoa and plants, a different family of HMTs are specific for H3K27me1, examplified by Arabidopsis ATXR5/ATXR6 (15).As a protozoan model organism, Tetrahymena thermophila features high levels of H3K27 methylation and several H3K27-

Orchestration of H3K27 methylation: mechanisms and therapeutic implication

“…In addition, these proteins have been reported to suppress gene expression by increasing H3K27me3 [ 78 , 79 ]. In Tetrahymena thermophila, three genes EZL1 , EZL2 and EZL3 are found to be the EZ homologs [ 80 , 81 ]. Among them, EZL2 is expressed at higher level and is required for the introduction of me2 and me3 of H3K27 [ 82 ].…”

Quantitative proteomic analysis of histone modifications in decitabine sensitive and resistant leukemia cell lines

“…(4) The use of Q Exactive mass spectrometers that offer high-resolution option on both MS1 and MS2, allows for high confidence identification of peptides and PTMs. All together, these efforts greatly improve the number of PTMs identified and quantified by LC–MS/MS [ 63 , 68 , 70 , 71 ]. Our optimized method allowed profiling of histones and their PTMs from as few as 10 5 leukemia cells.…”

Rbbp7 Is Required for Uterine Stromal Decidualization in Mice1

“…As generally accepted, post-translational modification of histones converts unique regions of chromatin into transcriptionally active or inactive status. There are four kinds of core histones, H2A, H2B, H3 and H4, the amino-terminal tails of which provide the sites for acetylation, methylation, phosphorylation and ADP-ribosylation [11]. Among these modifications of histones, the most well studied is the acetylation on lysine residues in the amino-terminal tail of the histones [12].…”