Histones are fundamental proteins that play a crucial role in the organisation and regulation of DNA within eukaryotic cells. These highly conserved proteins are abundant in lysine and arginine residues, giving them a positive charge that allows them to bind tightly to the negatively charged DNA molecule. This electrostatic attraction is essential for the compact packaging of DNA within the nucleus.
The primary function of histones is to act as spools around which DNA winds, creating structural units called nucleosomes. A nucleosome, the fundamental unit of chromatin, consists of approximately 146 base pairs of DNA wrapped around a core octamer of histones. This octamer is composed of two copies each of four core histone proteins: H2A, H2B, H3, and H4. Additionally, a linker histone, H1, binds to the DNA as it enters and exits the nucleosome, further stabilising the structure.
The structure of histones is noteworthy. Core histones possess a globular domain and N-terminal tails that exhibit high flexibility. These tails protrude from the DNA-wrapped nucleosome and are particularly important as they can undergo various chemical modifications, which have significant implications for gene regulation.
Histones serve several critical functions in the cell:
- DNA packaging: Without histones, the DNA in each human cell would stretch to about 1.8 metres when fully extended. However, when wound around histones, this length is reduced to approximately 90 micrometres of 30 nm diameter chromatin fibres. This remarkable compaction allows a full genome to fit within the confines of the nucleus.
- DNA protection: By tightly wrapping DNA, histones help protect it from damage. This protective function is crucial for maintaining the integrity of the genetic material.
- Gene regulation: Histones play a vital role in gene regulation. DNA accessibility to transcription factors and other regulatory proteins is influenced by how tightly it is wound around histones. Moreover, chemical modifications to histone tails can significantly impact gene expression.
- DNA replication: Histones are also important in the process of DNA replication. The expression of histone genes is tightly regulated and coupled with DNA replication to ensure proper packaging of newly synthesised DNA into chromosomes.
Histone modifications are a key aspect of epigenetic regulation. These modifications can include methylation of arginine or lysine residues, or acetylation of lysine. Such changes can affect how other proteins, such as transcription factors, interact with the nucleosomes. For instance, lysine acetylation removes a positive charge from the lysine, reducing the electrostatic attraction between DNA and histone. This results in partial unravelling of the DNA, making it more easily accessible and thus increasing gene expression.
In conclusion, histones are not merely structural proteins but are dynamic regulators of genomic function. Their ability to compact DNA, protect genetic material, and influence gene expression through various modifications makes them central players in the complex world of molecular biology and genetics. Understanding histones and their modifications is crucial for unravelling the intricacies of gene regulation and epigenetic inheritance.
Types of histone modifications
Histone modifications are post-translational modifications (PTMs) of histone proteins that make up the nucleosome, the fundamental unit of chromatin in eukaryotic cells. These modifications play a crucial role in regulating chromatin structure and function, ultimately impacting gene expression and various cellular processes. The most well-studied histone modifications include acetylation, methylation, phosphorylation, ubiquitination, sumoylation, and ADP-ribosylation.
Acetylation
Histone acetylation, first reported by Allfrey et al. in 1964, is a highly dynamic process regulated by two main enzyme families: histone acetyltransferases (HATs) and histone deacetylases (HDACs). HATs transfer an acetyl group from acetyl CoA to the ε-amino group of lysine side chains, neutralising the lysine’s positive charge and potentially weakening histone-DNA interactions.
Acetylation is generally associated with transcriptional activation, as it makes chromatin more accessible to transcriptional machinery. This modification primarily occurs on lysine residues of histones H3 and H4, with acetylation of K9 and K27 on histone H3 (H3K9ac and H3K27ac) typically associated with enhancers and promoters of active genes.
Methylation
Histone methylation occurs mainly on the side chains of lysines and arginines. Unlike acetylation, methylation does not alter the charge of the histone protein. Lysines can be mono-, di-, or tri-methylated, while arginines can be mono-, symmetrically, or asymmetrically di-methylated.
The effects of histone methylation on transcription depend on the specific residue modified and the degree of methylation. For instance, both mono- and tri-methylation on K4 of histone H3 (H3K4me1 and H3K4me3) are activation markers, with H3K4me1 typically marking transcriptional enhancers and H3K4me3 marking gene promoters. In contrast, tri-methylation on K9 and K27 of histone H3 (H3K9me3 and H3K27me3) are repressive signals.
The first histone lysine methyltransferase (HKMT) identified was SUV39H1, which targets H3K9. Most HKMTs that methylate N-terminal lysines contain a SET domain harbouring enzymatic activity.
Phosphorylation
Histone phosphorylation is a highly dynamic modification that occurs on serines, threonines, and tyrosines, predominantly in the N-terminal histone tails. This modification is controlled by kinases and phosphatases that add and remove phosphate groups, respectively.
Phosphorylation plays a critical role in chromosome condensation during cell division, transcriptional regulation, and DNA damage repair. For example, phosphorylation of histone H3 at serine 10 and 28, and histone H2A on T120, are involved in chromatin compaction and regulation of chromatin structure during mitosis. Phosphorylation of H2AX at S139 (resulting in γH2AX) serves as a recruiting point for DNA damage repair proteins.
Other modifications
In addition to acetylation, methylation, and phosphorylation, histones can undergo other modifications such as ubiquitination, sumoylation, and ADP-ribosylation. These modifications also play important roles in regulating chromatin structure and function.
Ubiquitination, for instance, occurs on all histone core proteins, with H2A and H2B being the most common targets. Monoubiquitylation of histones H2A, H2B, and H2AX is observed at DNA double-strand break sites, playing a central role in the response to DNA damage.
These various histone modifications work together in complex patterns to regulate gene expression and other cellular processes, forming what is often referred to as the ‘histone code’. Understanding these modifications and their interactions is crucial for unravelling the intricacies of epigenetic regulation and its impact on cellular function.
Histone acetylation and gene activation
Histone acetylation, a highly conserved process across eukaryotes, has been linked to gene activation since its discovery nearly 60 years ago. This post-translational modification plays a crucial role in regulating chromatin structure and transcriptional activity. Generally, hyper-acetylation of histone N-terminal tails results in an open chromatin state and is correlated with gene activation, while hypo/de-acetylation leads to condensed chromatin structure and gene repression.
HATs and HDACs
The level of histone acetylation is regulated by two families of enzymes: histone acetyltransferases (HATs) and histone deacetylases (HDACs). HATs catalyse the addition of acetyl groups from acetyl-CoA to the ε-amino group of lysine residues in histone tails, neutralising the positive charge and increasing hydrophobicity. Conversely, HDACs remove acetyl groups, resetting the physicochemical properties of histone tails.
The HAT-HDAC system plays a pivotal role in modulating overall cellular fate by regulating gene expression and repression on both temporal and spatial bases. This balance is crucial for cellular homeostasis, as evidenced by studies in various cancer forms where somatic mutations of HATs or altered HDAC activity due to chromosomal translocation can lead to oncogenic orientations.
Effects on chromatin structure
Acetylation has a profound impact on chromatin structure and accessibility. By decreasing the affinity of DNA for histones and releasing histone tails from linker DNA, acetylation opens up the usually condensed chromatin structure. This relaxed, open conformation allows transcription factors, co-factors, and RNA polymerase II complexes to access the DNA.
Studies have shown that acetylation of histones H3 and H4 counteracts the tendency of nucleosomal fibres to fold into highly compact structures in vitro. In vivo, acetylated chromatin exhibits increased sensitivity to DNase I, indicating greater accessibility to interacting proteins.
The effects of histone acetylation on chromatin dynamics can be further shaped by nucleosome stability. Acetylations on the core domain of histones can directly affect this stability, influencing overall chromatin structure.
Recruitment of transcription factors
Histone acetylation plays a crucial role in facilitating the recruitment of transcription factors and other regulatory proteins. Transcriptional activation within a permissive domain frequently correlates with additional, targeted acetylation of histones at promoter nucleosomes. Many cases have been documented where activating transcription factors recruit HAT-containing co-activators to specific promoters.
The genome-wide distribution of histone acetylation shows a positive correlation between enrichment at transcriptional start sites (TSS) and gene expression. This relationship underscores the importance of acetylation in creating a favourable environment for transcription initiation.
In Arabidopsis, for example, histone acetylation is generally deposited near transcription start sites. Mutation of AtGCN5, a histone acetyltransferase, significantly reduces H3K9/14ac levels in promoter regions, leading to inhibition of coding or non-coding RNA synthesis. Additionally, the BRAT1 protein in Arabidopsis binds directly to acetylated H4 to facilitate transcription activation, preventing gene silencing at methylated genomic regions.
In conclusion, histone acetylation serves as a critical regulatory mechanism for gene activation by modifying chromatin structure and facilitating the recruitment of transcriptional machinery. The interplay between HATs and HDACs maintains a delicate balance that is essential for proper cellular function and homeostasis.
Histone methylation and gene silencing
Histone methylation plays a crucial role in gene silencing and chromatin regulation. This post-translational modification occurs on lysine and arginine residues of histone proteins, influencing gene expression and chromatin structure. The effects of histone methylation on transcription depend on the specific residue modified and the degree of methylation.
Lysine and arginine methylation
Lysine methylation can occur as mono-, di-, or trimethylation, while arginine can be mono- or dimethylated (either symmetrically or asymmetrically). These modifications are catalysed by protein arginine methyltransferases (PRMTs) and histone lysine methyltransferases (HKMTs).
Methylation of histone H3 at lysine 9 (H3K9) and lysine 27 (H3K27) is associated with heterochromatin formation and transcriptional repression. For instance, trimethylation of H3K9 (H3K9me3) is a hallmark of constitutive heterochromatin, while H3K27 trimethylation (H3K27me3) is associated with facultative heterochromatin.
The first identified histone lysine methyltransferase, SUV39H1, targets H3K9. Most HKMTs that methylate N-terminal lysines contain a SET domain harbouring the enzymatic activity. In contrast, arginine methylation often occurs in intrinsically disordered regions of proteins, impacting biological processes like protein-protein interactions and phase separation.
Polycomb group proteins
Polycomb group (PcG) proteins are well-studied chromatin regulators that play a crucial role in gene silencing. They form two distinct complexes: Polycomb Repressive Complex 1 (PRC1) and Polycomb Repressive Complex 2 (PRC2).
PRC2 is responsible for methylating H3K27, a key repressive mark. The catalytic subunit of PRC2 is EZH1 or EZH2, which also contains RBAP46/48, SUZ12, and EED. PRC1, on the other hand, catalyses monoubiquitylation of histone H2A on lysine 119.
These complexes work together to maintain transcriptional repression through various mechanisms:
- Chromatin compaction: PRC1 can induce compaction of nucleosomal arrays, with each Polycomb complex compacting three nucleosomes.
- Inhibition of transcription activation-associated histone modifications: For example, CBX interacts directly with the histone acetyltransferase CBP to inhibit acetylation of histone H3.
- Maintenance of chromatin interaction: PRC1 and PRC2 can form chromatin loops to keep target genes silenced.
Heterochromatin formation
Heterochromatin is a key architectural feature of eukaryotic chromosomes, crucial for maintaining genomic stability. It can be categorised into two types: constitutive and facultative heterochromatin.
Constitutive heterochromatin (CH) is consistently silenced in all cell types and is molecularly defined by the presence of H3K9me3. This modification is carried out by the histone methyltransferases Suv39h in mammals, Su(var)3-9 in Drosophila, and Clr4 in yeast.
Facultative heterochromatin (FH) consists of cell-type-specific heterochromatic regions that can switch to euchromatin under certain conditions. It is marked by the presence of PRC1 and PRC2, with the latter depositing H3K27me3.
The formation and maintenance of heterochromatin involve complex mechanisms:
- Reader-modifier coupling: For example, HP1 proteins selectively bind methylated H3K9 through their chromodomains.
- RNA-mediated silencing: Non-coding RNAs transcribed from heterochromatin can be processed into small RNAs, which are important for heterochromatin formation in several organisms.
- Spreading from nucleation sites: Heterochromatin can spread along the chromatin from specific nucleation points, a process counteracted by inhibitory factors.
Understanding these mechanisms of histone methylation and heterochromatin formation is crucial for unravelling the complexities of gene regulation and epigenetic inheritance.
Histone phosphorylation and cell signaling
Histone phosphorylation plays a crucial role in regulating chromatin structure and function in eukaryotic cells. This post-translational modification, which occurs primarily on serine and threonine residues, has an influence on various cellular processes, including mitosis, DNA damage response, and transcriptional regulation.
Mitosis and chromosome condensation
During mitosis, histone phosphorylation is essential for chromosome condensation and segregation. The phosphorylation of histone H3 at serine 10 (H3S10) is particularly significant in this process. This modification appears early in the G2 phase within pericentromeric heterochromatin and spreads, coinciding with mitotic chromosome condensation.
The Aurora kinase family, specifically Aurora-A and Aurora-B, has a significant impact on H3 phosphorylation during the G2/M transition. Aurora-B colocalises with phosphorylated histone H3 during the G2 phase and is present in the centromeric region concurrent with H3 phosphorylation during prophase and metaphase. Both Aurora-A and Aurora-B can physically interact with the H3 tail and efficiently phosphorylate Ser10 both in vitro and in vivo.
Interestingly, mutation of Ser-10 in Tetrahymena results in abnormal chromosome segregation and extensive chromosome loss during mitosis and meiosis, establishing a strong link between signalling and chromosome dynamics.
DNA damage response
Histone phosphorylation plays a crucial role in the DNA damage response, particularly in the repair of double-strand breaks (DSBs). The phosphorylation of histone H2AX at serine 139 (γH2AX) serves as a signal for the recruitment of DNA repair proteins to the sites of DNA lesions.
Several phosphoinositide 3-kinase-related protein kinases (PIKKs) are responsible for H2AX phosphorylation, including ATM (ataxia telangiectasia mutated), ATR (ATM and Rad3-related), and DNA-dependent protein kinase (DNA-PK). ATM is considered the major physiological mediator of H2AX phosphorylation in response to DSB formation.
The MRN complex (MRE11-RAD50-NBS1) recognises DNA damage, recruits ATM to the site of damage, and targets ATM to initiate phosphorylation of its substrates. This process leads to the formation of γH2AX foci, which attract repair factors and lead to a higher concentration of repair proteins surrounding a DSB site.
Transcriptional regulation
Histone phosphorylation also has a significant role in transcriptional regulation. Phosphorylation of histones, such as H3S10 and H3 serine 28 (H3S28), can modulate chromatin structure and influence the binding of transcription factors and chromatin-remodelling complexes, thereby regulating gene expression.
Specific phosphorylation events have been linked to the regulation of particular genes:
- H3S10ph and H2B serine 32 phosphorylation (H2BS32ph) have been associated with the expression of proto-oncogenes such as c-fos, c-jun, and c-myc.
- Phosphorylation of H3 threonine 11 (H3T11) and H3 threonine 6 (H3T6) in response to androgen receptor-dependent gene activation has been shown to regulate transcription by controlling H3 methylation.
- Phosphorylation of H3 tyrosine 41 (H3Y41) by JAK2 disrupts chromatin binding by HP1α, leading to transcriptional activation of JAK2-regulated genes, including the oncogene imo2.
These various phosphorylation events demonstrate the complex interplay between histone modifications and gene regulation, highlighting the importance of histone phosphorylation in cellular signalling pathways.
Crosstalk between histone modifications
Histone modifications play a crucial role in regulating gene expression and chromatin structure. These post-translational modifications (PTMs) do not function in isolation but interact with one another in complex ways, creating a dynamic and intricate system of gene regulation. This interplay between different histone modifications is often referred to as ‘crosstalk’.
Combinatorial effects
The combinatorial effects of histone modifications are a key aspect of the ‘histone code’, which dictates the transcriptional state of genomic regions. Multiple effector modules in a protein or complex can interact with histone modifications on the same or across histones and nucleosomes. These interactions can be categorised as intranucleosomal (binding to the same nucleosome) or internucleosomal (binding to different nucleosomes).
The diversity of these interactions allows for precise control of gene expression. For instance, a single epigenetic mark, such as H3K4me3, may activate gene transcription in one context but repress it in another, depending on the surrounding marks. This multivalent engagement of histone modifications is crucial for recognising discrete marking patterns with composite specificity and enhanced affinity.
Sequential modifications
Histone modifications often occur in a sequential manner, with one modification influencing the occurrence of another. This sequential process is evident in the activation of gene expression. During the activation state, DNA-bound activators first recruit histone acetyltransferases (HATs) to the promoter, while DNA-bound RNA polymerase recruits histone methyltransferases to the open reading frame (ORF).
An example of this sequential process is the interplay between H3K4 methylation and histone acetylation. H3K4me3 has been shown to promote H3 or H4 acetylation. This pattern is consistent with notable patterns of coexisting histone marks in human cells, such as ‘H3K4me2/3 + H4K16ac’ and ‘H3K4me2/3 + H3K9/14/18/23ac’, which have been supported by chromatin immunoprecipitation (ChIP) and mass spectrometry (MS) methods.
Antagonistic modifications
While some histone modifications work together, others can have antagonistic effects. For instance, H3S10 phosphorylation prevents heterochromatin protein 1 (HP1) from binding to H3K9me3, a marker for mammalian heterochromatin. This antagonistic relationship demonstrates how different modifications can regulate each other’s effects on chromatin structure and gene expression.
Another example of antagonistic modifications involves the Polycomb Repressive Complex 2 (PRC2). H3K36me3, when present in a symmetric configuration (on both H3 molecules within a nucleosome), inhibits PRC2’s ability to methylate H3K27 on oligonucleosome substrates. This interaction highlights the complex interplay between different histone modifications and their effects on enzyme activity.
The crosstalk between histone modifications is not limited to interactions within the same histone tail. Trans-histone interactions, where modifications on different histone tails influence each other, have also been observed. For example, the lysine demethylase KDM5A exhibits enhanced demethylase activity for H3K4me3 when its PHD finger binds to unmodified H3K4 peptide in trans.
Understanding the intricate crosstalk between histone modifications is crucial for unravelling the complexities of gene regulation and epigenetic inheritance. As research in this field progresses, it becomes increasingly clear that these modifications do not act in isolation but form a complex, interconnected network that fine-tunes gene expression and chromatin structure.
Conclusion
To wrap up, histone modifications play a crucial role in regulating gene expression and chromatin structure. These modifications, including acetylation, methylation, phosphorylation, and others, work together in complex patterns to fine-tune cellular processes. The interplay between different histone modifications, often referred to as ‘crosstalk’, creates a dynamic system of gene regulation that impacts various aspects of cell biology.
Understanding the intricate relationships between histone modifications has far-reaching implications for our knowledge of epigenetics and gene regulation. This insight could lead to breakthroughs in fields such as cancer research and developmental biology. As research in this area continues to advance, it’s likely to uncover even more complex interactions, further expanding our understanding of how cells control their genetic information.
