Telomeres are nucleoprotein complexes at the physical ends of linear eukaryotic chromosomes. They protect the chromosome ends from various external attacks to avoid the loss of genetic information. Telomeres are maintained by cellular activities associated with telomerase and telomere-binding proteins. In addition, epigenetic regulators have pivotal roles in controlling the chromatin state at telomeres and subtelomeric regions, contributing to the maintenance of chromosomal homeostasis in yeast, animals, and plants. Here, we review the recent findings on chromatin modifications possibly associated with the dynamic states of telomeres in
In eukaryotic cells, the chromosome ends are protected by telomeres from inappropriate fusion and degradation, and incomplete DNA synthesis during DNA replication (1). Maintenance of the proper structure and function of telomeres is essential for the conservation of genetic information, chromosomal stability, and thus, cell survival (2). Eukaryotic chromosome ends are mainly divided into telomeres and adjacent subtelomeric regions (3). Telomeres consist of double-stranded repetitive G-rich DNA with single-stranded overhangs. When deletion of DNA sequences at the end of chromosomes is caused by various cellular events, telomerase accesses and adds telomeric repeats to critically short telomeres preferentially with its reverse transcriptase activity using its own internal RNA template, thereby effectively stabilizing telomere length (4). Telomere-binding proteins in mammals, known as shelterin, participate in the formation and maintenance of the specialized telomeric structure (T-loop) and the precise regulation of telomere length (5). Moreover, they interact with several non-telomere-binding proteins involved in DNA repair and recombination, contributing to the integrity and dynamics of the telomeres (6). Subtelomeric regions in humans are composed of degenerated telomeric repeat sequences with a high density of methylated CpG DNA sequences (7). The heterochromatic nature of subtelomeric regions influences the epigenetic silencing of telomere-adjacent genes by the telomere position effect (TPE) and abnormal chromosome recombination in yeast and humans (8). Non-coding RNAs containing telomeric repeats (TERRA) are generated in these regions (9). TERRA expression is regulated by the chromatin state, and in turn, telomere length is regulated by the expression level of TERRA (9, 10). Unique structures of telomeres and characteristics of telomere-related proteins have been observed in yeast, animals, and plants, demonstrating that these are evolutionarily-conserved and essential features of telomeres (5, 11, 12).
Telomeres, like centromeres, are generally defined as heterochromatic regions of the genome, characterized by increased chromatin condensation and decreased access to regulatory proteins (13). Many researchers have tried to understand what kinds of epigenetic marks are enriched in telomeric chromatin, how telomeres are regulated by these epigenetic modifications at the level of chromatin state, and thus, how these telomeric modifications affect their biological function. A repressive chromatin environment, formed by histone modifications and DNA methylation at telomeres and subtelomeric regions, has been shown to control the telomeric structure and function in yeast and mammals (14, 15).
In yeast, NAD+-dependent histone deacetylase Sir2 is a component of the silent information regulator (SIR) complex, which is implicated in the silencing of subtelomeric chromatin (16). Sir2 is recruited to telomeres by Rap1, and its HDAC (histone deacetylase) activity is necessary for its proper localization on telomeres and regulation of the heterochromatin structure in telomeres. Mammalian SIRT6 is specifically associated with telomeric chromatin and the loss of its HDAC activity results in the hyperacetylation of telomeric histone H3K9, telomere dysfunction, premature cellular senescence, and impaired silencing of telomere-proximal genes, indicating that SIRT6 modulates the telomeric chromatin structure (17, 18). Mammalian TPE is HDAC- and telomere length-dependent (19, 20).
In addition, histone methyltransferases involved in the trimethylation of H3K9 and H4K20, which are the main histone marks of telomeric and subtelomeric heterochromatin, contribute to the regulation of telomere length in mammals (21, 22). Moreover, heavily methylated DNA at mammalian subtelomeric regions are associated with the regulation of telomere elongation, TERRA expression, and stability (23, 24).
These studies present strong evidence that epigenetic modifications are involved in the composition of telomeric chromatin and have important roles in its regulation. Despite these outstanding achievements, the precise composition of epigenetic marks at telomeric chromatin and the relationship between telomeres, telomere-binding proteins, and these epigenetic regulators are not fully understood yet. Especially, many chromatin modifying proteins associated with epigenetic modifications in various target loci do not contain DNA-binding domains, thus prompting questions regarding how these proteins find their target loci.
Recent data in
In eukaryotes, the combination of different post-translational modifications (methylation, acetylation, phosphorylation, ubiquitination, sumoylation, and ADP-ribosylation) on the N-terminal tails of histone dictates the rapid change of the chromatin state into a transcriptionally-active euchromatin or silent heterochromatin state (25). Histone acetylation and deacetylation of lysine residues are reversible processes, mediated by histone acetyltransferases (HATs) and histone deacetylases (HDACs), respectively (26).
Based on the sequence homology to yeast HDACs, plant HDACs are classified into three major families: RPD3/HDA1, SIR2, and HD2 (27). Studies have reported that several histone modifiers, as partners of telomere-binding proteins, are required for regulation of the chromatin state of
Through Southern blot analysis using mutants, it was shown that
These reports also suggest the possible regulation of histone modification via combinatorial composition and competition between the different kinds of HDACs. This implies that the coordination between the various HDACs at telomeres is a universal regulatory mechanism in yeast and plants (32). In budding yeast, reports have shown that different types of HDAC proteins competed with each other for appropriate adjustment of the boundary element on telomeric chromatin. Rpd3 HDAC protein is necessary to restrict the SIR complex to telomeres and thus, modulates a barrier to prevent the spread of the SIR-dependent telomere position effect (32–34). Additionally, the competition of Sir2 HDAC and Sas2 HAT creates flexible boundaries at telomeres in yeast (35, 36).
Moreover, it has been noted that histone modifications at lysine 9 and lysine 27 on histone H3 were significant components of the telomeric chromatin in
DNA methylation is conserved in many eukaryotic organisms. Once established, DNA methylation is inherited through mitosis, and often through meiosis, and this provides an effective epigenetic mark (40). In
The chromatin state is regulated by interplay between the epigenetic modifications in eukaryotes (42). In some cases, the modifications on histone tails provide the binding site for effector proteins. It has been proposed that the interconnections between the epigenetic modifications act as signals to each other for establishing and maintaining stable epigenetic states. In
In contrast to the effect of deacetylated H3K9 on telomeric DNA methylation, an
In addition to DNA methyltransferases and histone modifiers, chromatin structural proteins also control the composition and level of epigenetic marks. Chromatin remodeling factor DDM1, a SWI2/SNF2 orthologue, has been previously reported to facilitate heterochromatin formation by promoting the access of DNA methyltransferase to the heterochromatin (51, 52). Southern blot analysis using a
Most recent study on DDM1 reported that telomere shortening in a late generation
Heterochromatin is stably-inherited and thus, must contain one or more epigenetic marks to direct its maintenance during cell division (54). Heterochromatin is generally characterized by H3K9me1,2, H3K27me1,2, H4K20me1, and methyl cytosine, whereas euchromatin is characterized by H3K4me1,2,3, H3K36me1,2,3, H4K20me2,3, and histone acetylation in
In mammals, telomeric nucleosomes have a more compact structure with heterochromatic features (15). There is a higher density of H3K9me3, H4K20me3, and HP1 (heterochromatin protein 1) in mammalian telomeres and subtelomeric regions, as well as higher levels of methyl cytosine by DNMT1 and DNMT3a/b in subtelomeric regions. Heterochromatic marks at telomeres have been proposed to act as negative regulators of telomere elongation. Interestingly, loss of heterochromatic marks at telomeres did not seem to affect TRF1 and TRF2 binding, indicating that shelterin recruitment was uncoupled from telomeric chromatin regulation (21, 22, 56, 57). Most recently, a study on the epigenetic characteristics of human telomeres revealed that telomeres had lower levels of H3K9me3 and enriched levels of H4K20me1 and H3K27Ac marks compared to certain heterochromatic loci in different human cell lines (58). In addition, several cancer cell lines that maintain their telomeres through ALT exhibited heterochromatic levels of H3K9me3. This suggested that telomeres in ALT cells became ‘subtelomeric’ according to their heterogeneous length and sequence composition containing degenerated telomeric repeats via recombination with subtelomeric regions (59, 60). It also implies that the mechanism of telomere maintenance by recombination in ALT is considerably different from that in canonical conditions whereby telomeres are elongated by telomerase. These results highlight the differences in several previous reports. It appears that the effect of the epigenetic features of telomeres and subtelomeric regions on their functions in humans is still an open question.
Analysis of epigenetic marks on the chromatin structure of
Although the epigenetic characteristics of
The principal functions of the heterochromatic state of telomeres are the protection of chromosome ends, the regulation of telomere length, and the suppression of recombination events at the telomeres. Recent findings noted that DNA methylation and histone modifications were involved in the regulation of chromatin status and the elongation of telomeres in many species, and suggested the possibility that cooperation and/or competition of these epigenetic modifications are required for the subtle and elaborate regulation of telomeric and subtelomeric chromatin state, thus maintaining the homeostasis of chromosomes.
However, the biological meaning of the formation of chromosome ends and the regulation of the chromatin state at the chromosome ends is considerably unrevealed and disputable. Especially, there are many unidentified pieces in the puzzle of the epigenetic regulation of
Moreover, the unrevealed functions of telomere-binding proteins are still remained. In humans, hTRF1 and hTRF2 are associated with ITSs, as well as telomeres, contributing to the stability of chromosomes.
This work was supported by the Basic Science Research Program from the Korean National Research Foundation (Grant no. NRF-2016R1D1A1A09919983 to M.H.C.), and in part by the Brain Korea 21(BK21) PLUS program.