Protein methylation, like other post-translational modifications, has a complex mechanism and plays a critical role in life regulation. Protein methylation modification occurs under the catalysis of methyltransferase, where the amines on the side chains of lysines or arginines undergo methylation. Moreover, methylation can also form methyl esters on the carboxyl side chains of aspartic acid or glutamic acid, although the focus here is primarily on the former methylation form. Methylation increases steric hindrance and replaces the hydrogen in amino groups, thereby influencing the formation of hydrogen bonds. As such, methylation can regulate the interactions between molecules themselves and the interaction of these molecules with their target proteins.
The classification of Arginine Methylation
In 1967, Paik and Kim discovered arginine methylation and its roles in life processes like signal transmission, transcription activation, and protein sorting. Many proteins undergo arginine methylation. In eukaryotic cells, there are three types of methylated arginine: NG-monomethylarginine (MMA), NGNG-asymmetric dimethylarginine (aDMA), and NGN'G-symmetric dimethylarginine (sDMA). Different proteins utilize differing methylation patterns for arginine residues, with some proteins adopting a variety of methylation forms. The methylation of heterogeneous nuclear ribonucleoproteins (hnRNPs) and other RNA-binding proteins often occurs on RGG tripeptide segments. Moreover, all arginine methylation in RGG segments is monomethylation and asymmetric dimethylation rather than symmetric dimethylation; this pattern is also true for RXR and RG segments. Conversely, the arginine residues of myelin basic protein (MBP) not only undergo monomethylation but also symmetric dimethylation; Similarly, the arginine residues in SmD1 and SmD3 proteins undergo symmetric dimethylation. Unlike hnRNPs, the methylated arginine in MBP, SmD1, and SmD3 proteins lies in GRG tripeptide segments, indicating that different types of amino acids in the adjacent positions of arginine residues may impact its methylation form.
Histone Methylation Modification
Histones, essential for transcription and other processes, take part in cellular nuclear activity through the chemical modification of their termini, such as phosphorylation, acetylation, and methylation. The methylation of histones lysine and arginine is associated with transcription regulation and the formation of heterochromatin. In summary, the increase in histone acetylation level relates positively to transcription activity, while the results of histone methylation modification are more complex, ranging from transcription enhancement to transcription suppression.
Methylation of Histone Lysine Residues
Methylation of histone lysine residues occurs on H3-K4, H3-K9, H3-K27, H3-K36, H3-K79, and H4-K20, and can also occur on the N-terminus of H1. The methylation of H3-K9, H3-K27, and H4-K20 is related to chromosome condensation, while methylation of H4-K9 may be associated with large-scale chromatin level repression. The methylation of H3-K4, H3-K36, and H3-K79 is related to chromosomal transcriptional activation, with monomethylation modifications on H3-K4 capable of negating gene repression prompted by H4-K9 methylation.
In 2001, Hisashi et al. were the first to confirm a correlation between histone H3-K9 methylation and DNA methylation. Through genomic scanning of Neurospora crassa strains with DNA methylation defects, a gene containing the SET domain, dim-5, was identified. Dim-5 is a H3 histone methyltransferase (HMTase) that can directly or indirectly bind DNA methyltransferase by acting on a methylated H3-K9 site via a mediator. In 2002, subsequent functional analyses of the kryptonite mutant strain in Arabidopsis thaliana by Jackson et al. further corroborated this relationship.
Shi et al. discovered that the nucleosome homolog of amine oxidase, LSD1 (KIAA0601), can act as a histone demethylase and a transcriptional corepressor. LSD1 specifically demethylates histone H3 Lys4. Lysine demethylation reactions occur via oxidation, producing formaldehyde. Significantly, upon repression of LSD1 through RNA interference, there was an increase in H3 Lys4 methylation and corresponding target gene repression, suggesting that LSD1 inhibits transcription via histone demethylation. This research reveals that histone methylation is dynamically regulated by histone methyltransferases and demethylases.
Histone Arginine Methylation
Histone arginine methylation at the H3-R2, H3-R4, H3-R17, and H3-R26 sites can enhance transcription. Although histone methyltransferases have long been studied, demethylases have remained elusive. In 2004, Cuthbert et al. proposed the process of "deimination," transforming histone arginine into citrulline to counteract methylation on arginine. Subsequent research by Wang et al. showed that PAD4 (peptidyl arginine deiminase 4) regulated histone arginine methylation through converting methyl arginine to citrulline, simultaneously generating methylamine. They found that PADI4 (peptidyl arginine deiminase 4, also abbreviated by Wang et al. as PAD4) specifically acts on the arginine residues at the terminal end of H3, R2, R8, R17, and R26, converting them to guanidino alanine. The deimination process mediated by PAD4 would then inhibit arginine methylation mediated by CARM1. While mono-methylation could still undergo deimination, arginine di-methylation impedes PAD4's deimination process. In vivo targeting experiments on endogenous promoters revealed that PAD4 could inhibit gene activation mediated by hormone receptors. Moreover, PAD4 was "recruited" by the pS2 promoter when this hormone-mediated gene transcription activity was curtailed, suggesting PAD4 regulates gene expression by modulating histone arginine methylation and citrullination.
Trojer et al.'s study identified three unique protein arginine methyltransferases (PRMTs) in the filamentous fungus Aspergillus nidulans. With specific antibodies, they demonstrated in vivo methylation of histone arginine, and high levels of PRMTs showed specificity to specific core histones in A. nidulans, suggesting these enzymes play a crucial role in chromatin activity regulation. It was found that when expressed as GST fusion proteins, they all manifested inherent histone methyltransferase activity. Protein arginine methyltransferases A (RmtA) and C (RmtC) respectively showed significant sequence homology to human proteins PRMT1 and PRMT5, while protein arginine methyltransferase B (RmtB) was remotely related to human or mouse PRMT3. Both native and recombinant RmtA were found to catalyze the methylation of histone H4-R3 specifically, with recombinant RmtA induced histone H4 methylation affecting acetylation catalyzed by the p300/CBP complex. The RmtB-GST fusion protein was found to specifically catalyze the methylation of histones H3, H4, and H2A.
Protein Methylation in living Organisms
Methylation of histones does not merely occupy a central position in the epigenetic modifications of eukaryotic chromatin, but it also exerts profound effects on cellular differentiation, development, gene expression, genome stability, and cancer research. Other forms of methylation and methyltransferases also play crucial roles in living organisms. Aberrations in protein methylation or mutations in methyltransferases are often implicated in disease onset. For example, monomethylated arginine and asymmetrical dimethylated arginine inhibit nitric oxide synthase (NOS), and an aberrant presence of these amino acids has been discovered in many people with cardiovascular diseases. The irreversible nature of arginine methylation might influence NOS activity in proteins undergoing methylation. Several methylated proteins have also been identified in autoimmune diseases, such as the discovery of myelin basic protein antibodies in patients with multiple sclerosis. Chuikov et al. have discovered a new mechanism that regulates p53 function, accomplished via lysine methylation catalyzed by Set9. Set9 specifically methylates a residue in the C-terminal regulatory region of p53. Methylated p53 is sequestered within the cell nucleus where its stability is heightened. Set9 regulates the expression of p53 target genes contingent on the methylation sites of p53. The crystal structure of the composite formed by Set9, p53, and accessory factor product provides the molecular foundation for the recognition of p53 through lysine methyltransferases.
References
- Trievel R C. Structure and function of histone methyltransferases. Critical Review 48 s in Eukaryotic Gene Expression, 2004, 14(3): 147~169
- Sims III R J, Nishioka K, Reinberg D. Histone lysine methylation: a signature for chroma 49 tin function. Trends in Genetics, 2003, 19(11): 629~639
- Tamaru H, Selker E U. A histone H3 methyltransferase controls DNA methylatio 50 n in Neurospora crassa. Nature, 2001, 414(6861): 277~283
- Jackson J P, Lindroth A M, Cao X F, et al. Control of CpNpG methylation by the KRYPTONITE 51 histone H3 methyltransferase. Nature, 2002, 416: 556~560
- Shi Y, Lan F, Matson C, et al. Histone demethylation mediated by the nuclear amine oxida 52 se homolog LSD1. Cell, 2004, 119(7): 941~953
- Cuthbert G L, Daujat S, Snowden A W, et al. Histone deimination antagonizes arginine methylation. Cell, 2004, 118(5): 545~553
- Wang Y, Wysocka J, Sayegh J, et al. Human PAD4 regulates histone arginine methylation levels 54 via demethylimination. Science, 2004, 306(5694): 279~283
- Trojer P, Dangl M, Bauer I, et al. Histone methyltransferases in Aspergillus nidulans: evidence for a novel enzyme with a unique 55 substrate specificity. Biochemistry, 2004, 43: 10834~10843
- Chuikov S, Kurash J K, Wilson J R, et al. Regulation of p53 activity through lysine methylation. Nature, 2004, 432: 353~360
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