Cellular epigenetic stability and cancer Peter Sarkies, Julian E. Sale  Trends in Genetics  Volume 28, Issue 3, Pages 118-127 (March 2012) DOI: 10.1016/j.tig.2011.11.005 Copyright © 2011 Elsevier Ltd Terms and Conditions

Figure 1 Histone post-translational modifications, transcription factors and their interplay in maintaining transcriptional memory through cell division. (a) Restoration of parental histone modifications following DNA replication. A tract of nucleosomes carrying the H3K9me2 mark (small red circles), characteristic of heterochromatin, is replicated. The new [H3/H4]2 tetramers (magenta centres) do not carry the H3K9me2 mark but can have it restored by the histone methyltransferase suppressor of variegation 3-9 (SUV39), which is recruited in complex with heterochromatin protein 1 (HP1), which binds through its chromodomain to an adjacent H3K9me2-modified nucleosome [12,13]. (b) Principle of epigenetic signal carriage by a transcription factor network. Transcription of Gene X (a transcription factor) is activated by an external signal. The product of Gene X, Protein X (orange box), can activate transcription of its own gene, providing positive feedback. In addition, Protein X activates transcription of Gene Y. Thus, as long as sufficient Protein X is carried through mitosis, expression of Gene Y will be maintained even in the absence of the initial signal. (c) A simplified diagram of the genetic circuit that determines the epigenetically stable white and opaque phenotypes of Candida albicans. Green indicates an actively transcribed gene, and red a repressed gene. The default state of the circuit is proposed to be the inactive, white state. Activation of white-opaque regulator 1 (WOR1) results in a series of positive feedback loops that can maintain the opaque state for many generations. For more details, see [78]. (d) Cooperation between trans-factor binding and histone modification in establishment of the H3K27me3 mark in Drosophila melanogaster. The polycomb repressive complex 2 (PRC2; purple), including at its core the methyl-binding protein extra sex combs (ESC) and histone methyltransferase Enhancer of zeste [E(Z)], is recruited to specific DNA sequences termed ‘Polycomb response elements’ (PRE) through the pleiohomeotic–recessive complex (PHO–RC) (light-green oval). This promotes trimethylation of H3K27. PRC2 is also able to bind H3K27me3 through its ESC subunit, providing a second mechanism for the recruitment of the enzymatic activity of E(Z) and the propagation of the H3K27me3 mark [28]. Cooperation between these two mechanisms is illustrated by the loss of silencing seen if either PHO or ESC is knocked out (ko) [26,28]. Trends in Genetics 2012 28, 118-127DOI: (10.1016/j.tig.2011.11.005) Copyright © 2011 Elsevier Ltd Terms and Conditions

Figure 2 Propagation of DNA methylation through replication and its interplay with histone modifications. (a) Restoration of parental DNA methylation patterns after DNA replication. Unmethylated DNA is excluded from the active site of DNA (cytosine-5-)-methyltransferase 1 (DNMT1), the major maintenance mammalian DNA methyltransferase (DNMT1 in purple; the active site in pink). This inhibition is relieved by hemi-methylated DNA, thereby providing a mechanism for restriction of the activity of DNMT1 to maintenance of DNA methylation after replication but preventing it from introducing de novo methylation at new sites [30]. (b) Cooperation between DNA methylation and histone methylation. Key to crosstalk between DNA methylation and H3K9me3 is the multi-domain protein ubiquitin-like with PHD and ring finger domains 1 (UHRF1; also known as Np95 or ICBP90). UHRF1 is able to recognise hemi-methylated DNA [31–33,79] and is also able to bind to the histone methyltransferase G9a [80], suggesting a mechanism by which DNA methylation reinforces histone methylation. Conversely, UHRF1 can bind H3K9me3 through its tandem Tudor domains [37], and it also interacts with DNMT1 [79], suggesting a mechanism by which histone methylation reinforces maintenance DNA methylation. Trends in Genetics 2012 28, 118-127DOI: (10.1016/j.tig.2011.11.005) Copyright © 2011 Elsevier Ltd Terms and Conditions

Figure 3 RNAi-mediated epigenetic memory and its interplay with histone modifications. (a) RNAi-based maintenance of pericentric heterochromatin in Schizosaccharomyces pombe. Bidirectional transcription of the pericentromeric repeats during S phase generates transcripts that pair to form double-stranded RNAs. These are acted on by Dicer to create short interfering RNAs that in turn recruit the RNA-induced transcriptional silencing (RITS) complex comprising Argonaute 2 (Ago2), Tas3 and Chp1. This complex recruits the histone methyltransferase Clr4 to create the H3K9me2 histone mark to reinitiate the formation of pericentromeric heterochromatin. (b) Cooperation between histone methylation and RNAi-mediated epigenetic memory [48]. In normal S. pombe cells, RNAi-driven H3K9 methylation by Clr4 counteracts competing acetylation by Mst2. In the absence of Dicer, however, Mst2 is able to acetylate newly synthesised H3 on K14, preventing Clr4 [recruited by the heterochromatin protein 1 (HP1) homologue, Swi6] from methylating K9, resulting in progressive replication dependent loss of silencing. However, in the absence of competition from Mst2, the Swi6/Clr4-mediated maintenance of H3K9 methylation is sufficient to maintain H3K9me2 and the heterochromatic state [48]. Abbreviation: ko, knockout. Trends in Genetics 2012 28, 118-127DOI: (10.1016/j.tig.2011.11.005) Copyright © 2011 Elsevier Ltd Terms and Conditions

Figure 4 A model for replication arrest-induced epigenetic instability. (a) Histone management during replication. Parental [H3/H4]2 tetramers (cyan centre) carrying, in this example, H3K9me2 marks (red circles) are displaced ahead of the replicative helicase (grey star). The parental and unmarked newly synthesised tetramers (magenta centre) are deposited behind the fork in an approximately 50:50 distribution between the two strands. Mark copying takes place by recognition of the H3K9me2 on the parental nucleosomes by heterochromatin protein 1 (HP1; light blue), which in turn recruits suppressor of variegation 3-9 (SUV39; pink) to methylate H3K9 in newly synthesised tetramers. (b) Stalling of replication by a G-quadruplex structure in REV1 or Fanconi anemia, complementation group J (FANCJ)-deficient cells. A G-quadruplex structure is shown having formed between the helicase and polymerase [orange oval attached to the proliferating cell nuclear antigen (PCNA) in green]. In the absence of REV1 or FANCJ, the fork remains paused at the structure, allowing the helicase to run ahead. DNA synthesis can reinitiate downstream of the stalled polymerase, but leaves an unreplicated gap. The advancing replicative helicase continues to displace parental nucleosomes but because only the lagging strand is available for redeposition, excess [H3/H4]2 tetramers are split and buffered by anti-silencing function 1 (ASF1) [51]. (c) Resolution and chromatinisation of the gap. The G-quadruplex structure is resolved, presumably by specialised helicases, and the gap is filled in. However, because this DNA synthesis takes place remote from the replication fork, there is no supply of marked parental histones. The gap is therefore chromatinised only with newly synthesised histones lacking the marks present at the locus prior to replication [52,53]. Trends in Genetics 2012 28, 118-127DOI: (10.1016/j.tig.2011.11.005) Copyright © 2011 Elsevier Ltd Terms and Conditions