Epigenetic Machinery

DNA Methyltransferases

DNA methylation represents a principal and common epigenetic modification. DNA methyltransferases (DNMTs), including DNMT1, DNMT3a and DNMT3b, catalyze the methylation of CpG dinucleotides by addition of a methyl group from S-adenosyl-L-methionine to the 5’ carbon position of cytosine. DNMTs are responsible for setting and maintaining DNA methylation patterns. DNMT3a and DNMT3b methylate previously unmethylated CpG sequences and are classified as de novo methyltransferases. In contrast, DNMT1, the most prevalent methyltransferase in mammals, ensures the maintenance of methylation tags onto new DNA strands during replication.

DNMT1 is predominantly localized to the nuclei of cells undergoing growth phases (G₁ and G₂) and is highly expressed during chromosome duplication (S phase) when it targets replication foci. DMAP1 binding site (DMAP); PCNA binding motif (PCAN); Replication foci targeting sequence (RTFS); zinc binding domain (CxxC); Bromo-adjacent homology domain (BAH).

Writers and erasers help determine chromatin’s transcriptional state through a dynamic process of post-translational covalent tags-addition and -removal. Specificity for the addition or removal of marks varies among family members, however HMT and HDM members generally target specific histones and residues.

Readers are chromatin-associated proteins which recognize specific epigenetic modifications and influence transactivation via different mechanisms including:

• Induction of nucleosome cross-linking
• Modulation of transcription factor and RNA polymerase complex access
• Promotion of additional chromatin post-translational modifications

Different families of readers have been identified by their unique modified-histone binding domains or “binding modules”. Readers interact specifically with modified histone sequences as part of multiprotein complexes and help modulate transactivation, either activating or repressing expression.

Chromatin-associated Proteins 

Histone Code

The sum of DNA-associated histones and histone marks, often referred to as the “histone code”, influence chromatin’s structure and function. Structurally, the histone code plays a key role in DNA folding and the extent of chromatin condensation. For example, specific histone tags are associated with euchromatin or heterochromatin which are transcriptionally active or inactive chromatin states respectively.

Antibodies to Histone Targets Validated for Single Cell Western

Beyond transcriptional regulation, the histone code, by virtue of its effects on chromatin’s structure influences various DNA-related functions including:

• DNA repair
• DNA replication
• Recombination events

Chromatin Remodelers

The position of the nucleosome influences the access of transcription factors to a gene’s regulatory region and therefore is a key determinant of gene expression. Nucleosome positioning is regulated by four subfamilies of chromatin remodeling protein complexes which depend on ATP hydrolysis to mediate various functions:

1. change the position of histones within nucleosomes
2. removal of histones
3. introduction of histone variants

Histone Variants

In addition to the four major histones that conform the nucleosome, histone variants have been identified in different species including mammals. Histone variants are incorporated into the chromatin independently of DNA-replication and play key roles in epigenetic mechanisms regulating gene expression. The sequence and domains of histone variants are not overtly different from that of canonic nucleosome histones. The mechanisms by which histone variants alter nucleosome structure or chromatin organization are not fully resolved.

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