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cells

Review The Code of

Harikrishnareddy Paluvai, Eros Di Giorgio and Claudio Brancolini *

Department of Medicine, Università degli Studi di Udine. P.le Kolbe 4, 33100 Udine, Italy; [email protected] (H.P.); [email protected] (E.D.G.) * Correspondence: [email protected]

 Received: 2 February 2020; Accepted: 17 February 2020; Published: 18 February 2020 

Abstract: Senescence is the end point of a complex cellular response that proceeds through a set of highly regulated steps. Initially, the permanent -cycle arrest that characterizes senescence is a pro-survival response to irreparable DNA damage. The maintenance of this prolonged condition requires the adaptation of the cells to an unfavorable, demanding and stressful microenvironment. This adaptation is orchestrated through a deep epigenetic resetting. A first wave of epigenetic changes builds a dam on irreparable DNA damage and sustains the pro-survival response and the cell-cycle arrest. Later on, a second wave of epigenetic modifications allows the genomic reorganization to sustain the of pro-inflammatory genes. The balanced epigenetic dynamism of senescent cells influences physiological processes, such as differentiation, embryogenesis and aging, while its alteration leads to , and premature aging. Here we provide an overview of the most relevant histone modifications, which characterize senescence, aging and the activation of a prolonged DNA damage response.

Keywords: DNA damage; OIS; RS; SIPS; SAHF; SASP

1. Introduction Aging is a physiological condition characterized by the functional deficit of tissues and organs due to the accumulation of senescent cells [1]. The key role of senescence in aging is well-established. Clearance of senescent cells in mouse models delays the appearance of age-related tissue and organ disfunctions [2,3]. Senescent cells are characterized by the permanent cell-cycle arrest sustained by the accumulation of cyclin-dependent kinase inhibitors/CDKi), like , and p27, as well as by the release of , chemokines and soluble factors. This modified microenvironment is known as senescence-associated secretory (SASP) [4]. The senescence state is triggered by different stimuli/stressors. These include the shortening of the (replicative senescence), the -induced replication stress, the oncogene-induced senescence (OIS), the accumulation of misfolded protein and/or (stress-induced premature senescence, SIPS) [5]. The impairment of the non-homologous end joining (NHEJ) and (HR) repair mechanisms are common traits of senescent cells [6–9]. Moreover, a widespread epigenetic resetting characterizes senescent cells and sustains cell-cycle arrest and cellular survival, through the activation of (i) CDKi [10,11], (ii) tumor suppressors [12], and (iii) secretion of chemokines and cytokines, as well as the remodeling of the microenvironment [13]. Macroscopically, senescent cells are characterized by the formation of peculiar areas of , named as SAHF (senescence-associated heterochromatin foci), mainly at E2F loci [14]. However, SAHF do not characterize all senescent cells [15] and are not causally linked to the onset of senescence [16]. Other epigenetic features, like the distension of satellites (senescence-associated distension of satellites, SADS) [17], the re-activation of transposable elements, and of endogenous retroviruses (ERV) [18,19], seem to better qualify different types of senescence. Finally, aging appears

Cells 2020, 9, 466; doi:10.3390/cells9020466 www.mdpi.com/journal/cells Cells 2020, 9, 466 2 of 18 to be marked by substantial re-arrangements of the , with the loss of H3 and H4 [20,21]. During senescence the epigenome undergoes temporal and sequential modifications that are mandatory to accomplish different cellular adaptations. Initially, this epigenetic resetting is mainly due to the accumulation of irreparable DNA damage. After this first wave of epigenetic modifications, the epigenome is remodeled and fixed in order to sustain the permanent cell-cycle arrest and to modulate the microenvironment.

2. The Epigenome of Replicative Senescence (RS) The telomeric TTAGGG repeats at ends protect the genome from degradation and distinguish natural ends from double-strand breaks (DSBs) [5,22,23]. Histone and non-histone (Shelterin) proteins sustain the folding of telomeric repeats in high-order structures that acquire a G-quadruplex shape as a consequence of Hoogsteen base pairing between consecutive guanines [24]. The loss of active telomerase complexes in somatic human cells blocks the lengthening of the telomeric ends. As a consequence, for each successful , telomeres get shorter and is restricted. This phenomenon is defined as replicative senescence (RS) [25]. The accumulation of irreparable DNA damage triggered during RS leads to permanent cell-cycle arrest and is considered among the main driving forces of aging [22].

2.1. Histone Variants The progressive accumulation of double-strand breaks (DSBs) at the chromosome ends is coupled with a deep epigenetic resetting that can be observed in pre-senescent cells, even distal from telomeres. This epigenetic repertoire builds up an epigenetic clock that dictates the replicative potential of human cells [26]. Late passage IMR90 and WI38 human fibroblasts are characterized by a reduced expression of core and H4 [21], of the linker histone H1 [27] and of the histone chaperons ASF1A/B and CAF1-p150/p60 [28]. While the decreased levels of H3 and H4 are due to reduced neosynthesis and increased mRNA degradation [21,29], H1 is post-translationally regulated [27]. Moreover, alternative spliced histone mRNAs belonging to the HIST1 cluster are reported to be accumulated in quiescent and RS-arrested human fibroblasts [30]. The epigenome of RS cells is also characterized by the deposition, at certain genomic loci, of the histone variants H3.3 [31], H2A.J [32] and by the release of genomic DNA from H2A.Z [33–35] (Table1). This redistribution results in and promotes the transcription of (i) tumor suppressors [30,31], (ii) inflammatory genes marking the SASP, [32] and iii) the cleavage of H3.3, which mediates the repression of E2F/RB target genes [31]. While in senescence, the HIRA-mediated deposition of H3.3 sustains cell-cycle arrest [31], and in embryonic stem cells ATRX and DAXX recruit H3.3 to repress the transcription of endogenous retroviruses (ERVs) [36]. It is possible that the redistribution of H3.3 between the proliferating and senescent cells, which depends on the detachment from ATRX/DAXX and the complexing to HIRA, is at the basis of the activation of ERVs observed in senescence [37] and aging [38]. The regulated deposition of all these histone variants is necessary [30] and sufficient to sustain cell-cycle arrest [31]. Interestingly, genes encoding these histone variants are frequently mutated in cancer, in confirmation of their tumor-suppressive properties [30,39].

2.2. SAHF an Open Question? The HIRA chaperone complex (HIRA/UBN1/CABIN1) controls both the deposition of H3.3 [30] and SAHF formation [40]. SAHF accumulation of //macroH2A blocks on E2F loci characterizes OIS and cells undergoing oncogene-induced replication stress [15]. However, RS is not uniformly characterized by the formation of SAHF [41]. In fact, focused heterochromatinization of E2F loci maintains cell-cycle arrest, also in cells described as SAHF-negative (e.g., BJ and MEFs) [41,42]. SAHF are defined as DAPI-dense nuclear regions characterized by the presence of a central core of Cells 2020, 9, 466 3 of 18 condensed chromatin, enriched for H3K9me3 and macroH2A. This core is surrounded by a peripheral ring of H3K27me3 [43,44]. SAHF formation requires p16/INK4 and consists of a deep and focused heterochromatin re-organization [45]. This reorganization is HMGA1/ASF1/HIRA-dependent [40,46] and is triggered by the GSK3β-mediated HIRA re-localization at PML bodies [47]. Even though SAHF dismantling, achieved through HMGA1 [46], ASF1 [40] or GSK3β knockdown [48], allows senescence escape, BJ fibroblasts and Hutchinson–Gilford syndrome (HGPS) cells enter senescence with minimal or no signs of SAHF formation. On the opposite, the SAHF formation in HMEC and MCF10A mammary cells in response to H-RAS/G12V over-expression fails to bring the cells to senescence [15]. Whether SAHF formation is only due to the arising of replication stress and could act as a barrier to DNA double-strand breaks spreading [15], or could mark chromatin regions stitched between remodeled LAD domains [45], needs further investigation. It is possible that a better definition of the SAHF, achieved through the improvement of the resolution of confocal nanoscopy and of ChIP-seq histone mapping, will clarify the debated role played by SAHF in senescence.

2.3. Histone Modifications A global decrease in /3 and but increase in H3K9me1 levels in gene bodies characterize RS in human fibroblasts [49,50]. By contrast, the heterochromatin marker H3K27me3 and the euchromatin marker are mostly redistributed with respect to proliferating cells (Table2). This redistribution correlates well with the expression profile of senescent cells [51–53]. A different behavior was reported for the repressive mark H4K20me3 and the activating mark . Although they are enriched in the regulative elements of genes modulated during RS, they do not correlate with changes observed in senescent cells [30,54]. This apparent paradox could stem from the masking effect imposed by the senescence-specific activation of super-enhancers (marked by /) and the activation of neighbor genes involved in SASP and metabolism [55]. A detailed comparison of H3K4me3 and H3K27me3 levels in proliferating and senescent human evidenced large-scale chromatin modifications during RS [56]. In senescent cells the augmented levels of H3K4me3 and H3K27me3 frequently co-localize in areas defined “mesas” that extend for hundreds of kilobases. Larger domains of RS genome (up to 10Mb) and defined “canyons” are characterized by decreased levels of H3K27me3 [56]. H3K4me3 and H3K27me3 mesas colocalize in LMNB1-associated domains and overlap DNA hypomethylation. Instead, canyons are enriched in gene bodies and enhancers and H3K27me3 loss correlates with the up-regulation of senescent transcriptional programs [56].

2.4. Nuclear Lamins The loss of LMNB1 typifies all senescence conditions [57] and triggers a deep re-modelling of lamina-associated domains (LADs) [44,56]. LADs re-modelling contributes to re-organizing not only heterochromatin and SAHF [44], but also euchromatin (eLADs) [58]. Moreover, the knockdown of LMNB1 in proliferating cells promotes the premature senescence and gives rise to a H3K4me3 /H3K27me3 re-organization similarly to what observed in RS, OIS and HGPS [56]. However, LMNB1 re-expression in RIS senescent cells does not overcome the proliferation of arrest and does not repair nuclear membrane blebbing [59]. It is still unknown if the re-expression of LMNB1 in senescent cells is strong enough to re-establish normal LAD domains or if the co-expression of an epigenetic regulator is needed. Similarly, the ectopic expression of lamina-associated polypeptide 2α restores the proliferation of HGPS cells [60], characterized by the expression of the form of LMNA [61,62]. Unfortunately, the impact of LAP2α expression on LAD domains has not been explored yet. The re-localized LAD domains in RS are characterized also by a general DNA hypomethylation that affects LINE and SINE repetitive elements and pericentromeric satellites [17]. This hypomethylation triggers the general distension of the chromatin associated to these genomic regions (senescence-associated distension of satellites or SADS) and their de-repression [17]. The activation of centromeric and Cells 2020, 9, 466 4 of 18 pericentromeric satellite repeats is associated with their exclusion from nucleoli compartments, while all the other nucleoli-associated domains (NADs) are significantly altered in senescent cells [63]. Despite LADs redistribution, 3D chromatin organization of human dermal fibroblasts (HDFs) is only partially altered when proliferating, quiescent and senescent cells are compared. A modest gain of short-range and the loss of long-range intra-chromosome interactions in permanently arrested cells was observed [64]. More evident in quiescent and senescent cells is the switch of topologically associated domains (TADs) from euchromatin areas (Hi-C compartment A) to heterochromatin (Hi-C compartment B) and vice versa. This remodeling reflects the transcriptional status of cell-cycle-associated genes [64]. Most of the heterochromatinization is due to condensin mobilization [65] and PRC2 (EZH2) deposition [66], while MLL1 plays a role in mediating euchromatinization [67,68]. Moreover, RS cells lose the TADs associated to the telomeres [69].

2.5. The CpG Clock In general, DNA methylation during aging progressively involve both hypomethylation and hypermethylation events. Importantly, the methylation status of a limited number of well-defined CpG islands associate well with human aging [26]. The quantification of the methylation rate of these CpG island allows a good prediction of human biological age [26,70–72]. According to these estimations, the methylome of men ages 4% faster than women [70]. Moreover, the methylation clock is accelerated in patients affected by neurodegenerative diseases [73–75], chronic stress and insomnia [76], as well as in two premature aging disorders like Werner’s syndromes and Hutchinson–Gilford Progeria syndrome [76,77], while it is subverted in cancer [78,79]. Aging is characterized by hypomethylation [41] and this correlates with the loss-of-function of stem cells [80]. Similar changes in the methylation prolife also characterize RS [81], and global hypomethylation characterizes both genomic and mitochondrial DNA [82]. CpG hypermethylated regions are associated with H3K27me3, H3K4me3 and H3K4me1, whereas hypomethylation is observed in the constitutive heterochromatin and lamina-associated domains (LADs) [83].

3. The Epigenome of Oncogene-Induced Senescence (OIS) The expression of a certain number of (K-RAS and H-RAS, BRAFV600E, myrAKT1, STAT5, nuclear HDAC4, N1ICD, ERBB2 and β-catenin) [84–92] or ablation of tumor suppressors (PTEN, APC and AXIN) in normal cells triggers a permanent and premature cell-cycle arrest defined as OIS [93,94]. OIS can occur also in the presence of ectopic TERT expression [6]. The most studied model of OIS is the RAS-induced senescence (RIS), obtained by over-expressing oncogenic mutants of RAS in fibroblasts, melanocytes and retinal pigment epithelial cells. The consistency of the RIS model has been validated in mice, after conditional induction of the monoallelic expression of K-RAS/G12V. Mice develop lung adenomas characterized by the accumulation of senescent cells [95]. Similarly, premature senescence blocks the spreading of oncogenic lesions in BRAF/V600E expressing melanocytic nevi [89]. Curiously, RAS fails to induce senescence in immortalized human mammary epithelial cells (HMECs), even after the ectopically expression of p16/INKa [96,97]. This failure has been associated to defects in the TGFβ signaling pathway [97]. It is generally accepted that the premature cell-cycle arrest characterizing OIS is elicited by the accumulation of irreparable DNA damage. In fact, the abrogation of ATM signaling allows senescence escape [6]. It is important to note that the DDR dependence was observed for RAS but its involvement in the case of other oncogenes, such as NOTCH and AKT1, needs further validations [90,98]. As explained above, RIS is characterized by the accumulation of SAHF [14]. However, the oncogenic activation of AKT1 and NOTCH or the knockdown of PTEN trigger OIS without SAHF formation [85,86,98]. In the case of NOTCH, this is due to the repression of HMGA1 [90]. Differently, in the case of AKT1 it was hypothesized a mechanism operating through GSK3β inhibition. In fact GSK3β controls the -dependent HIRA sub-compartmentalization [99]. In this respect, a similar defect in HIRA signaling prevents SAHF formation in senescent murine embryonic and fibroblasts expressing Cells 2020, 9, 466 5 of 18

RAS, but not to the accumulation of nuclear macroH2A.1 [42]. The deposition of the histone variant macroH2A.1 is not only required for SAHF formation. It also sustains a chromatin re-organization that allows the focused histone and the expression of the typical SASP cytokines and chemokines [100]. Moreover, the knock-down of macroH2A.1 in H-RAS/V12 IMR90-expressing cells not only blunts SASP, but also decreases the phosphorylation of γH2AX [100]. A common property of OIS cells is the loss of LMNB1 [57]. This causes a deep re-organization of LAD domains [44], similarly to what was observed during RS [56]. As a consequence, OIS is characterized by the re-localization and re-organization of LAD-associated heterochromatin domains [43,101]. This re-organization is sculptured in the distribution of H3K4me3 and H3K27me3 “mesas” and H3K27me3 “canyons”, as described above for RS cells [56]. Interestingly, the expansion of H3K27me3 “canyons” achieved though EZH2 repression sustains OIS by promoting the expression of cytokines [56] and of CDKi [66,102,103]. Similarly to RS, the histone variant H3.3 [31,104] and its Cathepsin L-mediated cleavage product H3.3cs1 [104] are deposited in RIS cells in the regulative elements of RB/E2F target allowing the permanent removal of H3K4me3 [31] and the increase in H3K9me3 levels [14]. Contemporary, the removal of H2A.Z and the de-methylation of H3K27me3 at tumor suppressor loci sustains cell-cycle arrest [33,66]. The inflammatory response is achieved through the deposition of the histone variant H2A.J [32] and the binding of HMGB2, which excludes these loci from SAHF [41]. HMGB2 also allows the H3K4 trimethylation mediated by the methyltransferase MLL1 [53]. Moreover, SASP loci are localized in newly formed super-enhancers that require the binding of BRD4 to promote their transcription [105,106]. Finally, OIS in human fibroblasts is characterized by the spatial rearrangement of pre-existing heterochromatin that give rise to SAHF [45]. Differently, HGPS cells are refractory to SAHF formation [45,107,108] and to heterochromatin focusing [45], probably because the huge alterations in the lamina compartment of these cells prevents the proper heterochromatin 3D-structure organization [109]. A similar displacement of H3K9me3 from LADs is observed in OIS, but it is followed by an increase in local interactions between H3K9me3 domains and sharp heterochromatinization [45]. Curiously and differently from RS and aging, OIS cells do not display any changes in CpG island methylation levels [110].

Different Types of OIS? While studies are increasingly describing RIS , detailed data about the epigenetic modifications in other types of OIS are unavailable. Additional data are desirable since increasing evidences highlight the key roles played by epigenetic regulators in maintaining OIS and counteracting oncogenic transformation [84,111–116]. For example, in melanomas the activation of H3K9me3 demethylases (LSD1 and JMJD2C) selectively de-represses E2F target genes, allowing senescence escape and tumorigenesis [117]. This result reinforces the idea that a better investigation of the epigenetic mechanism that sustains the early step of tumorigenesis is desperately needed.

4. The Epigenome of Stress Induced Premature Senescence (SIPS) SIPS is characterized by the accumulation of ROS (), due to mitochondrial disfunctions, ER stress or the exogenous administration of oxidative compounds [118–120]. Even though SIPS onset is independent from attrition, ROS accumulation can cause telomere disfunction and fusion in primary cells, thus sustaining cell-cycle arrest [120,121]. Accordingly, murine embryonic fibroblasts (MEFs) cultured in normoxia undergo premature senescence due to the accumulation of ROS-induced DNA damage, while the same cells cultured in hypoxia tend to indefinitely proliferate in virtue of the long telomeres [122]. When human cells are exposed to oxidative stress in vitro, they undergo stochastic transcriptional changes which resemble aged tissue. Recently, it has been demonstrated that oxidative stress contributes not only to aging but also to age-related diseases [123]. Moreover, the accumulation of 8-oxo-7,8-dihydro- Cells 2020, 9, 466 6 of 18

20-deoxyguanosine (8-oxodG) has been observed in the liver of aged mice [124]. Global histone of H3K4, K27 and K9 are increased when BEAS-2B cells are exposed to H2O2, and preincubation with ascorbate reverse these changes [125]. These evidences are confirmed also in other contexts [126]. The general heterochromatinization observed after H2O2 treatment is achieved in two steps. Firstly, by reducing acetylation (, , H4K16ac) [125,127] in a HDAC-dependent manner [127,128]. Secondly, by recruiting histone methyltransferases (HMTs) and inducing H3K27me3, H3K9me3 and H4K20me3 [127,129]. Chromatin condensation is an attempt to preserve the DNA from genotoxicity [130]. The downstream pathways that lead to cell-cycle arrest in cells exposed to oxidative stress imply the up-regulation of CDKi, the DDR response and SASP production, similarly to cells undergoing RS [131,132]. In addition, mitochondrial dysfunctions in IMR90 human fibroblasts lead to the ROS–JNK retrograde signaling pathways, which promote SASP and drive cytoplasmic chromatin fragments (CCFs) [133]. Importantly, the epigenetic homeostasis perturbation, achieved through HMTs or HDACs inhibition [129], is reported to expose cells to endogenous ROS production [134] or to trigger and sustain the senescence induced by the treatment with oxidative compounds [135]. In particular, some ROS generators inhibit PRC2 methyltransferases to allow the focused demethylation and transcriptional activation of CDKi [132]. Similarly, the senescence entry of PAK2 knocked-down MEFs cultured in normoxia is delayed because of the decreased deposition of H3.3 on CDKi loci [136]. At the DNA level, SIPS is characterized by a global DNA hypomethylation that only partially overlaps with the one observed during RS [81]. 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ChromatinHistone VariantsIncreasedH2A2/H2A1 ratio H2A ChromatinSenescence ndensation Chromatin Increasedduring ndensation ratioRS SenescenceIncreasedduringNI OISratio duringduringNI SIPS RS during[15,40 OIS] during SIPS H3, H4 ChromatinH3, condensationH4 co ndensationChromatin H2A2 condensation/ H2A1Chromatin condensationNI NI [15,40][21][27]NI [15,40][21][27]NI [21][27] H3,H2AH1 H4 ChromatinH3,H2AH1 H4 co ndensationChromatinH3, H1 H4 H2A2/H2A1 co ndensation Chromatin H2A2/H2A1 condensation [21] [21] [21] H3, H4 ChromatinH3, H4Chromatin co ndensationH3, H4 [21] H3, H4H3, H4 Chromatin condensation Chromatin condensation NI [27][21 ] NI [27] H1 condensation H1Increased ratio Increased ratio Increased ratio H2A.X ChromatinSensorH2A.X of DNAco ndensation damageChromatinSensor of DNAcondensation damageChromatinSensor of DNAcondensationNI damage NI [15,40][21]NI [15,40][21]NI [15,40][21] H3,H2A.XH2A H4 SensorH3,H2A.XH2ASensor H4 of DNA of damage DNASensorH2A.X H2A of H2A2/H2A1 DNA damage Sensor ofH2A2/H2A1 DNA damage H2A2/H2A1 [21] [21] [21] H2A.XH2A.X SensorH2A.X of DNA damage H2A.XIncreased ratio Increased ratio [21][21 ] H2A.X Chromatindamage condensation Increased ratioChromatin condensationNI NI [15,40]NI NI [15,40] H2A H2A H2A2/H2A1 H2A2/H2A1 H2A.J ChromatinSensorH2A.JPromote of DNA condensation SASP damage ChromatinSensor Promote ofH2A2/H2A1 DNAcondensation SASP damage Chromatin Promote condensation SASP [21][32] [21][32] [32][21] H3,H2A.XH2A.J H4H2A.J H3,H2A.XH2A.JPromotePromote H4 SASP SASP H3,H2A.JPromote H4 SASP Promote SASP [32][32 ] [32] [32] H2A.J H2A.JPromote SASP H2A.J [32] H2A.J Chromatin condensation Chromatin condensation [21] [21] H3, H4 Regulation ofH3, H4 H2A.ZH2A.Z SensorH2A.ZRegulationPromote of DNA ofSASP damage CDKi Sensor RegulationPromote of DNA ofSASP damage CDKi Sensor Regulation of DNA of damage CDKi NINI [33,34][32][21][33NI , 34 ] [33,34][32][21]NI [33,34][21] H2A.ZH2A.XH2A.J H2A.ZH2A.XH2A.JRegulation CDKi of CDKi H2A.ZH2A.XRegulation of CDKi Regulation of CDKi NI [33,34]NI [33,34]NI [33,34] H2A.Z H2A.ZRegulation of CDKi H2A.Z NI [33,34] H2A.Z SensorGene of DNA activation, damage Sensor of DNA damage [21] [21] H2A.X Gene activation, silencing,GeneH2A.X activation, silencing, H3.3, H3.3cs1 H3.3,GeneRegulation activation,PromoteH3.3cs1silencing, ofSASP silencing, CDKi Gene Regulation activation,Promote ofSASP silencing, CDKi Gene activation,Promote SASP silencing, NI [31,36,104][33,34][32]NI [31,36,104][33,34][32] [31,36,104][32] H3.3,H2A.ZH2A.J H3.3cs1H3.3, H3.3cs1 H3.3,GenechromosomeH2A.ZH2A.J H3.3cs1activation, segregation silencing,H3.3,GenechromosomeH2A.J H3.3cs1activation, segregation silencing,Gene activation, silencing, [31,36,104][31,36 ,104 ] [31,36,104] [31,36,104] H3.3, H3.3cs1 H3.3,Genechromosome activation, H3.3cs1chromosome segregation silencing, H3.3,chromosome H3.3cs1 segregation chromosome segregation [31,36,104] H3.3, H3.3cs1 chromosomePromote segregation SASP chromosome segregation chromosome Promote segregation SASP [32] [32] H2A.J chromosomeGene activation,segregation segregation silencing,GeneH2A.J activation, silencing, GeneRegulation activation, of silencing, CDKiGene Regulation activation, of silencing, CDKiGene Regulation activation, ofNI silencing, CDKi NI [31,36,104][33,34][21]NI [31,36,104][33,34][21]NI [33,34][21] H3.3,H2A.ZH3.1 H3.3cs1 H3.3,GenechromosomeH2A.ZH3.1 H3.3cs1activation,Gene activation, segregation silencing,GenechromosomeH2A.ZH3.1 activation, segregation silencing,Gene activation, silencing, H3.1 GenechromosomeH3.1 activation, segregation silencing,chromosome H3.1 segregation chromosome segregationNI NI [21]NI [21]NI [21] chromosomeRegulationsilencing, segregation of CDKichromosome segregation chromosomeRegulation segregation ofNI CDKi NINI [33,34][21] NI [33,34] H2A.ZH3.1 H3.1 chromosomeRegulation segregation of CDKi H2A.Z NI NINI [33,34][21 ] Gene activation,chromosome silencing,Gene activation, silencing,Gene activation, silencing, H3.3,H3.1 H3.3cs1 H3.3,H3.1 H3.3cs1 H3.3, H3.3cs1 NI NI [31,36,104][21]NI [31,36,104][21] [31,36,104] Table 2. HistonechromosomeTable post-translation segregation2. Histone segregation chromosome modificationspost-translation segregation (PMTs) chromosomemodifications observed segregation (PMTs)in senescence. observed RS: in Replicativesenescence. RS: Replicative Table 2. HistoneGeneTable post-translation activation, 2. Histone silencing, Table modificationspost-translation 2. Histone (PMTs)Gene modificationspost-translation activation, observed silencing, (PMTs)in modifications senescence. observed RS: (PMTs)in Replicativesenescence. observed RS: in Replicativesenescence. RS: Replicative senescence;Table 2. Histone OIS: oncogenesenescence;Table post-translation 2. Histoneinduced OIS: oncogeneTable modificationspost-translationsenescence; 2. Histoneinduced SIPS: (PMTs) modifications post-translationsenescence;Stress observed induced SIPS: (PMTs)in prematuremodifications senescence.Stress observed induced senescence; RS: (PMTs)in premature Replicativesenescence. [31,36,104]SASP:observed senescence; RS: in Replicativesenescence. SASP: RS: Replicative[31,36,104] H3.3,Tablesenescence; H3.3cs1 2. Histone OIS: Genechromosome oncogenesenescence;post-translation activation, induced segregation OIS: silencing,H3.3, Geneoncogenesenescence;modifications senescence; activation,H3.3cs1 induced OIS: silencing, SIPS:(PMTs) Genechromosomeoncogene senescence;Stress activation, observed induced induced segregation silencing,SIPS: in premature senescence.senescence;Stress induced senescence; SIPS:RS: prematureReplicative Stress SASP: induced senescence; premature SASP: senescence; SASP: senescence; OIS: chromosomeoncogenesenescence; inducedsegregation OIS: oncogenesenescence; senescence; ↑ induced OIS: SIPS: oncogene senescence;Stress ↑induced inducedNI SIPS:↓ premature senescence;Stress induced senescence;NI SIPS:↓ premature Stress →SASP: induced[21] senescence;NI premature →SASP:[21] senescence;NI SASP:[21] senescence;SenescenceH3.1 associatedOIS: oncogeneSenescenceH3.1 secretory induced associated phenotype; senescence;H3.1 secretory ↑: Increased SIPS: phenotype; Stress expression; induced↑: Increased ↓ :premature Decreased expression;↑ senescence;expression; ↓: Decreased →SASP:: No expression; ↓ →: No → TableSenescence 2. Histone associatedchromosomeTableSenescence post-translation secretory 2. Histone segregation associated phenotype; chromosomeSenescence modificationspost-translation secretory ↑: segregation Increasedassociated phenotype;(PMTs) chromosomemodifications expression; secretory observed ↑: segregation Increased phenotype; (PMTs)↓in: Decreased senescence. expression; observed ↑: Increasedexpression; RS: ↓in: Decreased Replicativesenescence. expression; →: No expression; RS: ↓: DecreasedReplicative →: No expression; →: No change;Senescence NI: associatedNot investigated.Genechange;Senescence activation, secretory NI: associatedNot silencing, phenotype; investigated.Senescence secretory↑ : Increased associated phenotype;Gene activation,expression; secretory : IncreasedNI silencing, phenotype;↓ : Decreased expression; : Increasedexpression;NI : Decreased expression;→ : [21]No expression;NI : Decreased : No expression;NI : [21]No Senescencesenescence;change;H3.1 NI: associated NotOIS: investigated. oncogenesenescence;change; secretory NI: induced NotOIS: phenotype; investigated. oncogenechange; senescence;H3.1 NI: :induced Increased Not SIPS: investigated. senescence;Stress expression; induced NI SIPS: : prematureDecreased Stress induced expression;senescence;NI premature SASP:: No[21] senescence; SASP: change; NI: Not investigated.chromosomechange; NI: segregation Not investigated.change; NI: Not investigated.chromosome segregation change; NI: Not investigated. ↑ ↑ ↓ ↓ → → SenescenceTable 2. Histone associatedSenescenceTable post-translation secretory 2. Histone associated phenotype; Table modificationspost-translation secretory 2. Histone: Increased phenotype;(PMTs) modificationspost-translation expression; observed : Increased (PMTs)in : modificationsDecreased senescence. expression; observed expression; RS: (PMTs)in: Decreased Replicativesenescence. observed: No expression; RS: in Replicativesenescence. : No RS: Replicative Histonechange;senescence; NI: NotOIS: investigated.Enzymes Histoneoncogenechange;senescence; NI: induced NotOIS: investigated.Enzymes HistoneRoleoncogenesenescence; senescence; during induced OIS: SIPS:Enzymes Roleoncogene Changessenescence;Stress during induced induced SIPS:Changes Rolepremature Changessenescence;Stress during induced senescence; Changes SIPS:Changes premature ChangesStress SASP: References induced senescence;Changes Changes premature SASP: References senescence;Changes SASP:References HistoneTable 2. HistoneEnzymesHistone post-translation Enzymes HistoneRoleTablemodifications during 2. Histone (PMTs)EnzymesRole post-translationChanges duringobserved Changesin Rolemodifications Changessenescence. during Changes RS:(PMTs)Changes ReplicativeChanges observed References Changes Changesin senescence. ReferencesChanges RS: Replicative References HistoneSenescencePMTs associatedEnzymesinvolvedSenescencePMTs secretory associated phenotype;involvedRoleSenescencesenescencePMTs duringsecretory ↑: Increasedassociated phenotype;involvedsenescenceduringChanges expression; secretory RS ↑ : Increased duringphenotype; Changes↓:senescence duringDecreased expression;OIS RS ↑:during Increasedexpression;Changesduring ↓: SIPSduringDecreased expression;OIS →References :RS Noduring expression; during ↓: SIPSDecreased OIS →: Noduring expression; SIPS →: No senescence;PMTs OIS: involvedoncogenePMTs inducedinvolvedsenescence;senescencePMTs senescence; OIS: SIPS: involvedoncogenesenescenceduring Stress RSinduced induced during prematuresenescence duringsenescence; OIS RS during senescence; SIPS:during SIPSduring Stress OIS SASP: RS inducedduring during prematureSIPS OIS duringsenescence; SIPS SASP: PMTschange; NI: Not involvedinvestigated.change; NI: Not investigated.senescencechange; NI: Not investigated.↓during RS ↑during↓ OIS during↑ SIPS HistoneSenescence associatedEnzymesHistone secretory phenotype;EnzymesRoleSenescence during ↑ : Increasedassociated ↓RoleRelocalizedChanges expression; duringsecretory in ↑ Relocalizedphenotype; Changes↓↓: RelocalizedDecreasedChanges in ↑ : inIncreasedexpression;Changes ↑RelocalizedChanges↓ expression; → inReferences: No Changes ↑ ↓: Decreased References expression; →: No KDM4A KDM4B KDM4AHeterochromatin KDM4B KDM4AHeterochromatin↓Relocalized KDM4B in ↑RelocalizedHeterochromatin↓Relocalized in in ↑Relocalized ↓Relocalized in[43,45,128,129] in ↑Relocalized in[43,45,128,129] H3K9me3PMTs H3K9me3involvedPMTs H3K9me3involvedsenescence senescencefocusedRelocalizedduring areaRS in duringfocusedRelocalizedfocusedRelocalizedduring OISarea areain RS during in duringfocusedRelocalized SIPSRelocalized OISarea [43,45,128,129]in during in Relocalized SIPS [43,45,128,129]in [43,45,128,129] H3K9me3change; NI: NotKDM4A H3K9me3investigated. KDM4B KDM4AHeterochromatinH3K9me3change; KDM4B NI: Not KDM4A Heterochromatin↓investigated.Relocalizedfocused KDM4B area in ↑ RelocalizedfocusedHeterochromatinfocused area inarea focused focused area area[43,45,128,129] focused area[43,45,128,129] [43,45,128,129] H3K9me3 KDM4A KDM4B Heterochromatin focused area focusedfocused area area focused focused area[43,45,128,129] area focused area Histone EnzymesHistone EnzymesHistoneRole during Enzymes↓RolefocusedChanges during area ↑focusedChanges↓RoleChanges area during Changes↑ ChangesChanges [43–References ChangesChanges [43–ReferencesChanges References ↓Relocalized in ↓Relocalized↓Relocalized in in ↓Relocalized↓ in[43– ↓ [43– [43– KDM4AKDM6B (JMJD3)KDM4B KDM4AKDM6BHeterochromatin (JMJD3)KDM4B HeterochromatinRelocalized in RelocalizedRelocalized in in Relocalized Relocalized in[43,45,128,129][43– in Relocalized in[43,45,128,129][43– [43– H3K27me3H3K9me3PMTs KDM6BH3K27me3H3K9me3involvedPMTs (JMJD3) KDM6BHeterochromatininvolvedsenescencePMTs (JMJD3) KDM6BHeterochromatininvolved↓senescencefocusedRelocalizedduring (JMJD3) area RS in ↓duringfocusedRelocalizedHeterochromatin↓senescencefocusedRelocalizedduring OISarea areain RS during in ↓duringfocusedRelocalized↓ SIPSRelocalizedduring OISarea 45,102,103,127–in RS during in ↓duringRelocalized SIPS OIS 45,102,103,127–in during SIPS H3K27me3Histone KDM6BH3K27me3Enzymes (JMJD3) KDM6BH3K27me3HeterochromatinHistoneRole (JMJD3) during KDM6BHeterochromatin↓EnzymesRelocalizedfocusedChanges (JMJD3) area in ↓ RelocalizedfocusedChangesHeterochromatinRolefocused areaduring inarea Changes focusedfocusedChanges area [43–45,102,103,127–areaReferences focusedChanges area 45,102,103,127– Changes 45,102,103,127–References H3K27me3 KDM6BH3K27me3 (JMJD3) HeterochromatinH3K27me3 focused area focusedfocused area area focusedfocused area 129]45,102,103,127–area focused area129]45,102,103,127– 45,102,103,127– H3K27me3PMTs involved senescencePMTs involved↓focusedRelocalizedduring area RS in ↑focusedduringRelocalized↓senescenceRelocalized areaOIS in during in ↑Relocalized↓ RelocalizedSIPSduring 45,102,103,127–[43– 129]inRS in ↑duringRelocalized OIS [43–129]in during SIPS 129] KDM4A KDM4B KDM4AHeterochromatin KDM4B KDM4AHeterochromatin↓Relocalized KDM4B in ↓ RelocalizedHeterochromatin↓Relocalized in in ↓ Relocalized in[43,45,128,129]129] [43,45,128,129]129] [43,45,128,129]129] H3K9me3 IncreasedH3K9me3 activity IncreasedHeterochromatinH3K9me3 activity geneHeterochromatin gene 129] IncreasedKDM6B (JMJD3) activity IncreasedHeterochromatinKDM6BHeterochromatin (JMJD3) activity geneIncreased HeterochromatinHeterochromatin focused activity area gene Heterochromatinfocused focused area area gene focused focused area 45,102,103,127–area [54,127] focused area45,102,103,127– [54,127] H3K27me3 H3K27me3 ↓Relocalizedfocused area in ↑Relocalizedfocusedfocused area inarea focused↓ Relocalized area [54,127] in ↑Relocalized in [54,127] [54,127] H4K20me3 KDM4AIncreasedH4K20me3of Suv420h2 KDM4B activity IncreasedHeterochromatinH4K20me3Heterochromatinof Suv420h2repression activity gene KDM4AIncreasedHeterochromatin Relocalizedrepression KDM4B activity in gene HeterochromatinRelocalizedHeterochromatin in gene [43,45,128,129] [43,45,128,129] H4K20me3H3K9me3 IncreasedKDM4AofH4K20me3 Suv420h2 KDM4Bactivity HeterochromatinofHeterochromatinH4K20me3H3K9me3 Suv420h2repression gene of Suv420h2repression repression 129][43,45,128,129][54,127] 129][54,127] [54,127] H4K20me3 of Suv420h2 of Suv420h2repression of↓ RelocalizedfocusedSuv420h2repression area in ↓Relocalizedfocused↓Relocalizedrepression area in in ↓Relocalized ↓Relocalizedfocused [43–areain [54,127] in ↓Relocalizedfocused area [43–in [43– IncreasedKDM6BKMT2/KDM5D/of Suv420h2 (JMJD3) activity IncreasedHeterochromatinKDM6BKMT2/KDM5D/Euchromatin,Heterochromatinrepression (JMJD3) activity gene geneHeterochromatin KDM6B Euchromatin, Heterochromatin (JMJD3) gene gene Heterochromatin H3K27me3 KMT2/KDM5D/H3K27me3 KMT2/KDM5D/Euchromatin,H3K27me3 gene KMT2/KDM5D/ Euchromatin,focusedRelocalized area gene focusedEuchromatin,RelocalizedfocusedRelocalized area area gene focusedRelocalized focused area 45,102,103,127– area [54,127][56,125] focused area45,102,103,127– [54,127][56,125] 45,102,103,127– H4K20me3H3K4me3 KMT2/KDM5D/H4K20me3H3K4me3 KMT2/KDM5D/Euchromatin, geneKMT2/KDM5D/ Euchromatin,↓RelocalizedRelocalized gene in ↓ RelocalizedRelocalizedEuchromatin,Relocalized in gene Relocalized↓ RelocalizedRelocalized[43– [56,125] in ↓RelocalizedRelocalized in [56,125] [43–[56,125] H3K4me3 KMT2/KDM5D/ofH3K4me3 KDM2BSuv420h2 Euchromatin,ofH3K4me3 KDM2BSuv420h2repressionactivation gene KDM2BRelocalizedrepressionRelocalizedactivation in RelocalizedRelocalizedRelocalizedactivation in Relocalized Relocalized129] [56,125] Relocalized 129] [56,125] 129][56,125] H3K4me3 KDM6BH3K4me3 (JMJD3) HeterochromatinH3K4me3 KDM6BRelocalized (JMJD3) RelocalizedHeterochromatin 45,102,103,127–[56,125] 45,102,103,127– H3K27me3H3K4me3 KDM2B H3K27me3KDM2Bactivation KDM2Bfocusedactivation area focusedactivation area focused 45,102,103,127–area focused area (MOF)KDM2B Histone DNA(MOF) repair,activation Histone chromatin DNA repair, chromatin IncreasedKMT2/KDM5D/(MOF) Histone activity DNAIncreasedHeterochromatinKMT2/KDM5D/(MOF)Euchromatin, repair, Histone activity chromatin gene geneDNAIncreasedHeterochromatin (MOF)Euchromatin, repair, Histone activity chromatin gene geneDNAHeterochromatin repair, chromatin gene 129][125,127] [125,127] 129] H4K16ac (MOF)H4K16ac Histone DNA(MOF) repair, Histone chromatin DNA(MOF) Relocalizedrepair, Histone chromatin DNARelocalized Relocalizedrepair, chromatin Relocalized [125,127][56,125][54,127] [125,127][56,125][54,127] [125,127][54,127] H4K20me3H3K4me3H4K16ac acetyltransferaseH4K20me3ofH3K4me3H4K16ac KDM2BSuv420h2 acetyltransferaseH4K20me3ofH4K16ac KDM2BSuv420h2compactionrepressionactivation of Suv420h2compactionrepressionactivation repression [125,127] [125,127] [125,127] H4K16ac Increasedacetyltransferase(MOF)H4K16ac Histone activity DNAHeterochromatinacetyltransferaseH4K16ac repair,compaction chromatin geneIncreasedacetyltransferase compaction activity Heterochromatincompaction gene Increasedacetyltransferase activity Heterochromatinacetyltransferasecompaction geneacetyltransferase compaction compaction [125,127][54,127] [54,127] H4K20me3H4K16ac acetyltransferase H4K20me3compaction [54,127] KMT2/KDM5D/(MOF)ofHAT/p300 Suv420h2 Histone DNAKMT2/KDM5D/(MOF)Euchromatin,HAT/p300Euchromatin, repair,repression Histone chromatin gene DNAKMT2/KDM5D/ Euchromatin,of Euchromatin, Suv420h2repair, chromatin gene Euchromatin, repression gene [55,125,127] [55,125,127] H3K9ac HAT/p300H3K9ac HAT/p300Euchromatin, HAT/p300Euchromatin,Relocalized RelocalizedEuchromatin,Relocalized Relocalized Relocalized [55,125,127][125,127][56,125] Relocalized [55,125,127][125,127][56,125] [55,125,127][56,125] H3K4me3H4K16acH3K9ac acetyltransferaseH3K4me3H4K16acHAT/p300H3K9acKDM2B acetyltransferaseH3K4me3HAT/p300H3K9acKDM2BEuchromatin,compactionactivation HAT/p300KDM2BEuchromatin,compactionactivation Euchromatin,activation [55,125,127] [55,125,127] [55,125,127] H3K9ac KMT2/KDM5D/HAT/p300H3K9ac Euchromatin,Euchromatin,H3K9ac gene KMT2/KDM5D/ Euchromatin, gene [55,125,127] H3K4me3H3K9ac H3K4me3 Relocalized Relocalized Relocalized[56,125] Relocalized [56,125] (MOF)HAT/p300KDM2B Histone DNA(MOF)HAT/p300Euchromatin repair,activation Histone chromatin DNA(MOF) KDM2BEuchromatin repair, HistoneNI chromatin DNA repair,activationNI NI chromatin NI [125,127] [125,127] H4K8ac HAT/p300H4K8ac HAT/p300Euchromatin,Euchromatin HAT/p300Euchromatin,EuchromatinNI EuchromatinNI NI NI NI [55,125,127][125,127] NI [55,125,127][125,127] [125,127] H4K16acH3K9acH4K8ac acetyltransferaseHAT/p300H4K16acH3K9acH4K8ac acetyltransferaseHAT/p300H4K16acH4K8acEuchromatincompaction acetyltransferaseHAT/p300EuchromatincompactionNI EuchromatincompactionNI NI NI NI [125,127] NI [125,127] [125,127] H4K8ac (MOF)HAT/p300H4K8ac Histone DNAEuchromatinH4K8ac repair, chromatin (MOF) HistoneNI DNA repair,NI chromatin [125,127] H4K16acH4K8ac H4K16ac [125,127] [125,127] H4K16ac acetyltransferase compaction acetyltransferase compaction H4K8acH3K9ac HAT/p300H4K8acH3K9ac HAT/p300Euchromatin,H3K9acEuchromatin HAT/p300Euchromatin,EuchromatinNI Euchromatin,NI NI NI [55,125,127][125,127] [55,125,127][125,127] [55,125,127] 5. The Epigenome5. The at DSBs Epigenome and5. during The at DSBs Epigenome DDR: and Early during at Epigenetic DSBs DDR: and Early Eventsduring Epigenetic inDDR: the SenescentEarly Events Epigenetic in Response the Senescent Events in Response the Senescent Response 5. The Epigenome5. The atHAT/p300 DSBs Epigenome and5. during The atEuchromatin, DSBs Epigenome DDR: and Early during atHAT/p300 Epigenetic DSBs DDR: and Early EventsduringEuchromatin, Epigenetic inDDR: the SenescentEarly Events Epigenetic in Response the[55,125,127] Senescent Events in Response the Senescent Response[55,125,127] 5. TheH3K9ac Epigenome atHAT/p300 DSBs and duringH3K9acEuchromatin, DDR: Early Epigenetic Events in the Senescent Response[55,125,127] HAT/p300 HAT/p300Euchromatin HAT/p300EuchromatinNI EuchromatinNI NI NI NI [125,127] NI [125,127] [125,127] 5. TheH4K8ac Epigenome 5. The atH4K8ac DSBs Epigenome and during atH4K8ac DSBs DDR: and Early during Epigenetic DDR: Early Events Epigenetic in the Senescent Events in Response the Senescent Response HAT/p300 Euchromatin HAT/p300NI EuchromatinNI NI [125,127] NI [125,127] H4K8ac H4K8ac

5. The Epigenome5. The at DSBs Epigenome and5. during The at DSBs Epigenome DDR: and Early during at Epigenetic DSBs DDR: and Early Eventsduring Epigenetic inDDR: the SenescentEarly Events Epigenetic in Response the Senescent Events in Response the Senescent Response 5. The Epigenome at DSBs and5. during The Epigenome DDR: Early at Epigenetic DSBs and Eventsduring inDDR: the SenescentEarly Epigenetic Response Events in the Senescent Response

Cells 2020, 9, 466 7 of 19

Table 1. Histone variants that characterize senescence. RS: Replicative senescence; OIS: Oncogene induced senescence; SIPS: Stress induced premature senescence; SASP: Senescence associated secretory phenotype; ↑: Increased expression; ↓: Decreased expression; →: No change; NI: Not Cells 2020, 9, 466 7 of 19 Cells 2020, 9, 466 Cells 2020, 9, 466 Cells 2020investigated., 9, 466 7 of 19 7 of 19 7 of 19

Table 1. Histone variants that characterize senescence. RS: Replicative senescence; OIS: Oncogene Table 1. HistoneTable variants 1. Histone thatHistones characterizeTable variants 1.and Histone that senescence characterize variantsRole. RS: duringthat senescenceReplicative characterize . RS:senescence;Changes senescenceReplicative OIS:. RS:senescence; Oncogene ReplicativeChanges OIS: senescence; OncogeneChanges OIS: OncogeneReferences induced senescence; SIPS: Stress induced premature senescence; SASP: Senescence associated induced senescence;induced SIPS: senescence; HistoneStressinduced inducedVariants SIPS: senescence; Stresspremature induced SenescenceSIPS: senescence; Stresspremature induced SASP: senescence;during prematureSenescence RS SASP: senescence;associatedduring Senescence OIS SASP: associated duringSenescence SIPS associated ↑ secretory↑ phenotype;↓ ↑: Increased↓ expression;→ ↓: Decreased→ expression; →: No change; NI: Not Cells 2020, 9,secretory 466 phenotype;secretory : Increasedphenotype;secretory expression; : Increasedphenotype; : Decreasedexpression; : Increased expression; : Decreasedexpression; : expression; No: Decreasedchange; NI:: expression;No Not 7change; of 18 NI:: No Not change; NI: Not Cells 2020investigated., 9, 466 Chromatin condensation NI [27]7 of 19 Cells 2020investigated., 9, 466 Cells 2020investigated., 9, 466 Cells 2020investigated.,H1 9, 466 7 of 19 7 of 19 7 of 19

HistonesTable 1.and Histone variants that characterizeIncreased senescence ratio . RS: Replicative senescence; OIS: Oncogene TableHistonesTable 2. Histone 1.and Histone post-translationHistonesTable variantsRole 1.and Histone duringthatHistones modificationscharacterizeTable variantsRole 1.and Histone duringthatChanges senescence (PMTs)characterize Chromatin variants Role observed. RS: coduringthatChanges ndensationsenescenceReplicativeChanges characterize in senescence. . RS:senescence;Changes senescenceReplicativeChangesChanges RS: OIS: . Replicative RS:senescence; Oncogene ReplicativeChangesReferencesChangesNI OIS: senescence; OncogeneReferencesChangesNI OIS: OncogeneReferences[15,40] HistoneinducedH2A Variants senescence; SIPS: Stress inducedH2A2/H2A1 premature senescence; SASP: Senescence associated senescence;Histoneinduced Variants OIS: senescence; oncogene Histoneinduced Variants SIPS: induced senescence; HistoneStressinduced senescence; inducedVariants SIPS: senescence; Stress SIPS:premature Stressinduced SenescenceSIPS: senescence; induced Stresspremature induced premature SASP: senescence;during prematureSenescence senescence; RS SASP: senescence;associatedduring SASP:Senescence OIS SASP: associated duringSenescence SIPS associated Senescence Senescenceduring RS Senescence↑ duringduring RS OIS duringduringduring↓ RS OIS SIPS duringduring OIS SIPS →during SIPS ↑ secretory↑ phenotype;↓ ↑: Increased↓ expression;→ ↓: Decreased→ expression; →: No change; NI: Not Senescencesecretory associated phenotype;secretory secretory : Increasedphenotype; phenotype;secretory expression; : :Increasedphenotype; Increased Chromatin: Decreasedexpression; expression;: Increased condensation expression; : : Decreasedexpression; Decreased : expression; No expression;: Decreasedchange; NI:: :expression;No NoNot change; NI:: No Not change; NI: Not[21] Cells 2020, 9, 466 Cells 2020, 9, 466 investigated.H3, H4↑ Chromatin condensation↓ → 7 of 19 NI7 of 19 [27] change;investigated. NI:H1 Not investigated. investigated.ChromatinH1 condensation investigated.ChromatinH1 condensation Chromatin condensation NI [27] NI [27]NI [27] HistoneTable 1. EnzymesHistoneTable variants 1. HistoneRole that duringcharacterize variants thatChanges senescence characterizeSensor .of RS: DNAChanges senescenceReplicative damage . RS:senescence;IncreasedChanges Replicative ratio OIS: senescence; Oncogene OIS: Oncogene [21] HistonesH2A.X and Increased Chromatin ratioRole Increased coduringndensation ratio IncreasedChanges ratio ReferencesChangesNI ChangesNI References[15,40] PMTsHistonesinducedH2A and Involvedsenescence; HistonesinducedChromatinH2A Role SIPS:and senescence; coSenescenceduring ndensationStressHistonesChromatin H2A induced Role SIPS:and coduringduring Changes ndensationStressprematureChromatin RS induced Role senescence; duringcoChanges ndensationChangesprematureNI OIS SASP: senescence;duringH2A2/H2A1Changes ChangesSenescenceChangesNI SIPSNI SASP: associated ChangesReferencesSenescenceChanges[15,40]NINI associatedReferencesChanges[15,40]NI References[15,40] Histone VariantsH2A2/H2A1 SenescenceH2A2/H2A1 duringH2A2/H2A1 RS during OIS during SIPS Histonesecretory Variants phenotype; Histonesecretory VariantsSenescence ↑: Increasedphenotype; Histone Variantsexpression;Senescence ↑: IncreasedduringRelocalized ↓ :RS Decreasedexpression; duringRelocalizedduring expression; ↓ :RS OISDecreased →duringduring: expression;No OISchange; SIPS →NI:during: No Not change; SIPS NI: Not SenescencePromote SASP during RS [43during,45,128 OIS, during SIPS [32] H2A.J ↓ Chromatin↑ condensation [21] H3K9me3investigated.KDM4A KDM4Binvestigated.ChromatinHeterochromatin Cellscondensation 2020ChromatinH3,, 9, 466H4 coinndensation focused Chromatin coinndensation focused [21] [21] [21]7 of 19 H3, H4 H3, H4 H3, H4 Chromatin condensation 129] NI [27] Chromatin condensationChromatinH1 condensationarea area NI [27]NI [27] H1 H1 H1 Chromatin condensation NI [27] RelocalizedRegulationRelocalized of CDKi NI [33,34] TableH2A.Z 1. Histone variantsSensor of DNAthat damage characterize senescence . RS:[43 Replicative–45,102 , senescence; OIS: Oncogene[21] Sensor of DNA damageH2A.XSensor of ↓DNA damage Sensor of ↓DNA damage [21] [21] [21] H3K27me3HistonesH2A.XKDM6B and (JMJD3)HistonesH2A.XRole and Heterochromatin duringH2A.X Role duringinChanges focusedChromatin coinChangesndensation focusedChanges IncreasedChangesChanges ratio ReferencesChangesNI ReferencesNI [15,40] Chromatin condensationinducedChromatinH2A senescence;Increased condensation ratio SIPS: Increased StressNI ratio induced Increased prematureNI ratioNI senescence;103,127[15,40]–129NI ] SASP: Senescence[15,40] associated HistoneH2A Variants HistoneH2A VariantsSenescence H2ASenescence duringareaChromatin RS coduringduringndensationarea RS OIS H2A2/H2A1duringduring OIS SIPS duringNI SIPS NI [15,40] Cells 2020, 9, 466 H2A2/H2A1Gene activation,↑ H2A2/H2A1 silencing, H2A2/H2A1 ↓ → [31,36,104]7 of 19 HeterochromatinH3.3,secretory H3.3cs1 phenotype; Promote: Increased SASP expression; : Decreased expression; : No change; NI: Not[32] Increased activityPromote SASP H2A.JPromote SASP chromosome Promote segregation SASP [32] [32] [32] H2A.J H2A.J investigated.H2A.J Chromatin condensation [21] H4K20me3 ChromatinChromatin cocondensationndensationgeneChromatinH3,Chromatin H4 cocondensationndensation NI [54,127[21][27]NI] [21][27] H3,H1 H4 of Suv420h2H3,H1 H4 TableH3, H41. Histone Chromatinvariants cothatndensation characterize senescence . RS: Replicative senescence; OIS: Oncogene[21] repression Gene activation, silencing, NI NI [21] H3.1 Regulation of CDKi NI [33,34] RegulationEuchromatin, of CDKiinducedH2A.Z Regulation senescence; of CDKi chromosome Regulation SIPS: segregationStress of CDKi induced premature NI senescence;[33,34] NI SASP: Senescence[33,34]NI associated[33,34] H2A.Z H2A.Z HistonesH2A.Z and Increased SensorratioRole of IncreasedDNAduring damage ratio Changes Changes Changes References[21] H3K4me3 H2A.XH2AKMT2 /KDM5DH2A.XChromatinSensorH2A/KDM2B of DNA condensation genedamagesecretoryH2A.XChromatinSensor of phenotype;DNA Relocalizedcondensation damage Sensor ↑ :of Increased DNARelocalized damage NI expression; ↓NI: NI Decreased [56 expression;,125[15,40][21] NI] →: No[15,40][21] change; NI: Not[21] HistoneH2A.X Variants H2A2/H2A1 SenescenceH2A2/H2A1 during RS during OIS during SIPS activation Gene activation, silencing, [31,36,104] Gene activation,H3.3, silencing,investigated.TableGene H3.3cs1 activation, 2. Histone silencing, Gene post-translation activation, silencing, modifications (PMTs) observed[31,36,104] in senescence.[31,36,104] RS: Replicative[31,36,104] H3.3, H3.3cs1 H3.3, H3.3cs1DNA H3.3, repair, H3.3cs1 chromosomePromote segregation SASP [32] (MOF) HistonechromosomeChromatinPromote co segregationndensation SASPchromosome ChromatinH2A.JPromote co segregationndensation SASP chromosome Promote segregation SASP [32][21] [32][21] [32] H4K16ac H3,H2A.J H4 H3,H2A.J H4 chromatinsenescence;H2A.JH1 OIS: Chromatinoncogene co inducedndensation senescence; SIPS: Stress[125 induced,127 ] premature senescence;NI SASP:[27] acetyltransferase Gene activation, silencing, ↑ NI ↓ NI → [21] Gene activation,compactionHistones silencing,SenescenceGeneH3.1 activation, and associated silencing, Gene Roleactivation, secretory during silencing, phenotype;NI Changes: Increased NINI expression;ChangesNI[21]NI : DecreasedChanges[21] expression;NI References : [21]No H3.1 H3.1 H3.1 chromosomeRegulation segregation of CDKi NI [33,34] chromosomeSensorRegulation of DNA segregationHistone of damage CDKichromosomeH2A.ZSensor RegulationVariants of DNA segregation of damage CDKi chromosome segregation Increased ratioNI [33,34][21]NI [33,34][21] H3K9ac H2A.ZH2A.XHAT /p300H2A.ZH2A.X Euchromatin,change;H2A.Z NI: Not investigated.ChromatinRegulationSenescence condensation of CDKi during RS [55during,125NI,127 OIS] duringNI SIPS [15,40][33,34] H2A H2A2/H2A1 Gene activation, silencing, Table 2. HistoneChromatin post-translation ndensation modifications (PMTs) observed in senescence. RS: Replicative[31,36,104] H4K8acTable 2. HATHistone/p300TableGene post-translation activation, Promote2. HistoneEuchromatin SASP silencing,TableGene post-translationmodifications activation, Promote 2. Histone SASP silencing,NI Gene post-translation(PMTs)modifications activation, coobserved silencing,NI (PMTs)modifications in senescence. observed (PMTs) RS:in senescence.Replicative observed[125[31,36,104],127[32] ] RS: in senescence.Replicative[31,36,104][32]NI RS: Replicative[27] H3.3,H2A.J H3.3cs1 H3.3,H2A.J H3.3cs1 H3.3,HistoneH1 H3.3cs1 EnzymesChromatinchromosome co ndensationsegregationRole during Changes Changes Changes [31,36,104]References[21] chromosome segregationH3.3,senescence;chromosomeH3, H3.3cs1 H4 segregationOIS: chromosomeoncogene inducedsegregation senescence; SIPS: Stress induced premature senescence; SASP: senescence; OIS:senescence; oncogene inducedOIS:senescence;PMTs oncogene senescence; inducedOIS:involved SIPS:oncogene senescence; Stress induced induced SIPS:senescence senescence; prematureStress induced SIPS: senescence;during prematureStress RSinduced SASP: senescence;during premature OIS SASP: during senescence; SIPS SASP: Gene activation, silencing, ↑ ↓ → Gene activation, silencing,SenescenceGene activation, associated↑ silencing,Chromatin secretory co↑ndensation phenotype;↓ Increased ↑: Increased ratio↓ expression;→ NINI ↓: Decreased→ expression;NINI →[15,40]: [21]No Senescence associatedSenescenceRegulation secretory associated of CDKi Senescencephenotype;H2AH3.1 Regulation secretory associated of: Increased CDKi phenotype; Gene activation, secretory expression; : silencing,Increased phenotype; NI : Decreased expression; : Increased NI expression;NI : Decreased expression; :[33,34]NI [21]No expression;NI : Decreased :[33,34] [21]No expression;NI : [21]No 5. The EpigenomeH2A.ZH3.1 at DSBsH2A.ZH3.1 and duringchange;H3.1 DDR: NI: Early Not investigated.chromosome EpigeneticSensor of DNA segregation damage Events in theH2A2/H2A1 Senescent ↓ Relocalized Response in ↑Relocalized in [21] change; NI: Not change;investigated.chromosome NI: Not segregation change;investigated.chromosomeH2A.X NI: Not segregation KDM4A investigated.chromosome KDM4B segregation Heterochromatin [43,45,128,129] H3K9me3 focused area focused area Double-strand DNAGene breaks activation, cause silencing, massiveGene activation, epigenetic silencing,Chromatin changes. ndensation Immediately, at the damaged sites, H3, H4 co [31,36,104] [31,36,104] [21] H3.3, H3.3cs1 H3.3,chromosome H3.3cs1 segregation Tablechromosome 2. Histone segregation post-translationPromote SASP modifications (PMTs)↓ observed ↓ in senescence. RS: Replicative[43–[32] the DNAHistone unwrapsTable 2. Histone fromHistoneTableEnzymes thepost-translation histones2. Histone HistoneTableEnzymes and H2A.J post-translationRolemodifications the2. during Histone chromatin Enzymes post-translation(PMTs)RolemodificationsChanges undergoesduring observed (PMTs)Rolemodifications Changes ainChanges de-structuration, duringsenescence. observed (PMTs)Changes ChangesRelocalizedRS:inChanges senescence.Replicative observed thus inReferences losingChangesRelocalized Changes RS:in senescence.Replicative in ReferencesChanges RS: Replicative References senescence; OIS:KDM6B oncogene (JMJD3) induced Heterochromatin senescence; SIPS: Stress induced premature senescence; SASP: senescence; OIS:senescence; oncogene inducedOIS:H3K27me3senescence;PMTs oncogene senescence; inducedOIS: involvedSIPS:oncogene senescence; Stress induced induced SIPS:senescence senescence; prematureStress induced SIPS: senescence;focusedduring prematureStress area RSinduced SASP: senescence;duringfocused premature areaOIS SASP: during senescence; SIPS SASP:45,102,103,127– compactnessPMTs [138 ]. ThisPMTsinvolved responseGene activation, is achievedsilencing,PMTsinvolvedGenesenescence activation, mainly silencing,involved Sensor throughsenescenceduring of DNA theRS damage chaperone-mediatedNI senescenceduring during OIS↑ RS duringNINI during during SIPS nucleosomesOIS RS during[21] NIduring ↓ SIPS OIS during[21] SIPS → [21] H3.1 H3.1 SenescenceH2A.X associated↑ Regulation secretory↑ of CDKi phenotype; ↓ : Increased ↓ expression;→ : Decreased→ expression;NI 129][33,34]: No Senescence associatedSenescencechromosome secretory associated segregation Senescencephenotype;H2A.Zchromosome secretory associated segregation: Increased phenotype; secretory expression; : Increased phenotype; : Decreased expression; ↑: Increased expression;↓ : Decreased expression; : No expression;↑ ↓ : Decreased : No expression; →: No disassembly [139]. Functionally, it allows the recruitment↓ of proteins↑ ↓ involved in↑ ↓ DDR.Relocalized Structurally in ↑Relocalized in change; NI: NotIncreasedKDM4A investigated. RelocalizedKDM4B activity Heterochromatin inHeterochromatin RelocalizedRelocalized ingene in RelocalizedRelocalized in in Relocalized in [43,45,128,129] change; NI: NotKDM4A change;investigated. KDM4B NI: Not KDM4AH3K9me3 change;investigated.Heterochromatin KDM4B NI: Not KDM4A investigated. Heterochromatin KDM4B Heterochromatin focused area[43,45,128,129] focused area [43,45,128,129] [43,45,128,129][54,127] it causesH3K9me3 significant topologicalH3K9me3 alterationsH4K20me3H3K9me3 in the DNA fiber.focusedPromote These area SASP modificationsfocusedfocused area area focused arefocused limited area area byfocused the area [32] H2A.J ofGene Suv420h2 activation, silencing,repression [31,36,104] subsequentTable heterochromatinization 2. HistoneTable post-translation 2. HistoneH3.3, upstream post-translation H3.3cs1modifications and downstreamchromosome (PMTs)modifications observed segregation from (PMTs) the in damagesenescence. observed site,↓ RS:in which senescence.Replicative restrains↓ RS: Replicative [43– Histone KMT2/KDM5D/Enzymes↓ Euchromatin,↓Role↓ during gene ↓ ↓RelocalizedChanges [43–in ↓RelocalizedChanges in[43– Changes [43–References Histonesenescence; OIS:Histonesenescence;Enzymes oncogene inducedOIS:Enzymes oncogeneRole senescence; during inducedKDM6B SIPS:Role senescence;Relocalized(JMJD3)Changes Stress during induced in SIPS:Heterochromatin RelocalizedChangesRelocalized Changes prematureStress inducedin in senescence;ChangesRelocalizedChangesRelocalizedRelocalized premature inSASP: in References senescence;ChangesRelocalizedRelocalized inSASP: References [56,125] relaxation [140]. KDM6B (JMJD3) KDM6BH3K27me3H3K4me3HistoneHeterochromatin (JMJD3) KDM6BEnzymes HeterochromatinRegulation (JMJD3) of CDKi HeterochromatinRole during focusedChanges area focusedChanges area NIChanges References[33,34]45,102,103,127– H3K27me3 H3K27me3 H3K27me3PMTsH2A.Z involvedGeneKDM2B focusedactivation, area silencing, senescencefocusedactivationfocused area area focusedfocusedduring area area RS 45,102,103,127– duringfocused areaOIS 45,102,103,127–during SIPS 45,102,103,127– SenescencePMTs associatedSenescencePMTsinvolved secretory associated PMTsinvolvedphenotype;senescence secretory ↑: Increased involvedphenotype;senescenceduring expression; ↑ RS: Increased senescenceduring ↓during: Decreased expression; OIS RS during expression;during ↓during: Decreased SIPS OIS RS → :NIduring No expression; during SIPS OIS → :during NoNI SIPS 129][21] The phosphorylation by PIKK kinasesH3.1 (ATM, ATRchromosome and DNA-PK) segregation of ser 139 of H2AX (γH2AX)129] is the 129] 129] (MOF) Histone DNA repair, chromatin ↓ ↑ change; NI: Not change;investigated. NI: Not investigated.Increased Gene↓ activation, activity silencing,Heterochromatin↑ ↓ gene↑ Relocalized in Relocalized in [125,127] widespread histone modificationIncreased activity that IncreasedH4K16acHeterochromatin extends activity for megabases IncreasedKDM4A geneHeterochromatin RelocalizedKDM4B activity around geneHeterochromatin inHeterochromatin a Relocalized DSBs.RelocalizedγH2AX ingene in Relocalized↓Relocalized acts as a in platformin ↑Relocalized in [31,36,104][43,45,128,129][54,127] KDM4A KDM4B KDM4AH3.3,H4K20me3H3K9me3Heterochromatin H3.3cs1 KDM4B KDM4Aacetyltransferase Heterochromatin KDM4B Heterochromatincompaction focused area[43,45,128,129] [54,127]focused area [43,45,128,129][54,127] [43,45,128,129][54,127] H4K20me3H3K9me3 H4K20me3H3K9me3 H4K20me3H3K9me3 ofchromosome Suv420h2focused segregationarea focusedrepressionfocused area area focused area for the recruitment of MDC1of Suv420h2 and 53BP1.ofTable Suv420h2repression This2. Histone recruitment of Suv420h2post-translationrepression is sustained modificationsrepression by the accumulation(PMTs)focused observed area atfocused in the senescence. area RS: Replicative Histone HistoneEnzymes EnzymesRole during KMT2/KDM5D/ HAT/p300GeneRole activation,Changes during silencing, Euchromatin,Euchromatin,Changes Changes gene Changes↓Changes References↓Changes References [43–[55,125,127] damaged site of DDR RNAsKMT2/KDM5D/ [141]. TheKMT2/KDM5D/H3K9acsenescence; generalEuchromatin, relaxation OIS: geneKMT2/KDM5D/ Euchromatin,oncogene ↓ of the induced chromatin geneEuchromatin, ↓ ↓senescence; is achieved gene SIPS: ↓ ↓Relocalized throughRelocalized Stress induced di[43–in NIff ↓ erentRelocalizedRelocalized premature in[43– senescence;NI SASP:[43–[21][56,125] H3K4me3H3.1 KDM6B Relocalized(JMJD3)Relocalized in Heterochromatin RelocalizedRelocalizedRelocalizedRelocalized in in RelocalizedRelocalizedRelocalizedRelocalized in in [56,125]RelocalizedRelocalized in [56,125] [56,125] H3K4me3PMTs KDM6BH3K4me3PMTsinvolved (JMJD3) KDM6BH3K27me3H3K4me3involvedHeterochromatinsenescence (JMJD3) KDM6B chromosomeHeterochromatinKDM2Bsenescence (JMJD3)during segregation RS Heterochromatinduringactivationduring OIS RS during duringfocused SIPS OIS area during focused SIPS area 45,102,103,127– mechanisms,H3K27me3 which includeH3K27me3KDM2B (i) the RNF8H3K27me3SenescenceKDM2B/Ubc13activation / HUWE1 associated KDM2Bactivation ubiquitylationfocused secretory area phenotype; focusedactivationfocused of Histone area ↑ area : Increased H1focused [142 expression; area,143 ],45,102,103,127– (ii) ↓ the: Decreased45,102,103,127– expression; →45,102,103,127–: No focused area focused area 129] change; NI: Not(MOF) investigated.HAT/p300↓ HistoneRelocalized DNA in ↑Euchromatin Relocalizedrepair,↓Relocalized chromatin in in ↑ RelocalizedNI in129] NI 129] 129][125,127] RNF168 mediated K63-ubiquitylationKDM4A(MOF) Histone KDM4B KDM4A(MOF)DNAH4K8ac ofHeterochromatin H2Arepair, Histone KDM4B /B chromatin and (MOF)DNA H2AXHeterochromatin repair, Histone [ 144chromatin ],DNA and repair, (iii) thechromatin PARylation-dependent, [43,45,128,129] [43,45,128,129] [125,127] H3K9me3 H3K9me3 H4K16acTable 2. HistoneIncreasedacetyltransferase post-translation activity Heterochromatin modificationscompaction gene (PMTs) observed[125,127] in senescence. [125,127] RS: Replicative[125,127] proteasomalH4K16ac degradation IncreasedacetyltransferaseH4K16ac of H1.2 activity [145 IncreasedacetyltransferaseH4K16acHeterochromatin]. compaction activity Increasedacetyltransferase geneHeterochromatin compactionfocused activity area geneHeterochromatin compactionfocusedfocused area area gene focused area [54,127] H4K20me3 H4K20me3 H4K20me3 of Suv420h2 repression [54,127] [54,127] [54,127] of Suv420h2 H4K20me3ofsenescence; Suv420h2repression OIS: of oncogene Suv420h2repression induced repressionsenescence; SIPS: Stress induced premature senescence; SASP: In addition to these huge chromatinHistone remodeling, EnzymesHAT/p300↓ localRelocalized histone in ↓RoleEuchromatin,Relocalized↓ modificationsRelocalized during↑ in in ↓Relocalized surroundingChanges in [43– Changes↓ the [43–Changes →References[55,125,127] KDM6BHAT/p300 (JMJD3) 5. TheKDM6BSenescenceH3K9acHAT/p300 HeterochromatinEpigenomeEuchromatin, (JMJD3) associated KMT2/KDM5D/ atHAT/p300 Heterochromatin DSBsEuchromatin, secretory and during phenotype;Euchromatin, Euchromatin, DDR: gene: EarlyIncreased Epigenetic expression; [55,125,127]Events : Decreased in the [55,125,127]Senescent expression; Response : [55,125,127]No DSBs takeH3K27me3H3K9ac place in cells KMT2/KDM5D/H3K27me3H3K9ac undergoing KMT2/KDM5D/ NHEJH3K9acPMTsEuchromatin, or HR, geneKMT2/KDM5D/ thusinvolvedEuchromatin, sculpturingfocused areageneEuchromatin, ansenescencefocusedfocused epigenetic area areagene patternfocusedduringRelocalized area specificRS 45,102,103,127–duringRelocalized for OIS 45,102,103,127–during SIPS [56,125] H3K4me3 H3K4me3 H3K4me3change; NI: Not investigated.KDM2BRelocalized RelocalizedactivationRelocalized RelocalizedRelocalized [56,125]Relocalized [56,125] [56,125] KDM2B H3K4me3KDM2Bactivation KDM2Bactivation activation 129] 129] each of the two repair mechanisms [138]. The activationHAT/p300 of NHEJ is supportedEuchromatin by a↓ generalNI chromatin↑ NI [125,127] HAT/p300 H4K8acHAT/p300Euchromatin KDM4A(MOF) HAT/p300Euchromatin HistoneKDM4B NI DNA HeterochromatinEuchromatin repair,NINI chromatin Relocalized NINI in [125,127]Relocalized NI in [125,127] [125,127] H4K8ac Increased(MOF)H4K8ac Histone activity IncreasedH3K9me3(MOF)DNAHeterochromatinH4K8ac repair, Histone activity chromatin DNA geneHeterochromatin repair, chromatin gene [43,45,128,129][125,127] expansion around the DSBs, by the depositionH4K16ac of the(MOF) histone Histone variants DNA H3.3 repair, [ 146 chromatin ] and H2AZfocused [ 147area] and[125,127][54,127]focused by area [125,127][54,127] H4K20me3H4K16ac H4K20me3H4K16ac H4K16acHistone acetyltransferaseEnzymes Rolecompaction during Changes Changes Changes References[125,127] the monoubiquitylationacetyltransferaseof Suv420h2 H2BK120. acetyltransferase Thisof Suv420h2compactionrepression monoubiquitylation acetyltransferase compactionrepression promotes compaction the appearance of H3K4me3 PMTs involved senescence ↓Relocalizedduring RS in ↓duringRelocalized OIS in during SIPS [43– KMT2/KDM5D/ KMT2/KDM5D/Euchromatin, geneHAT/p300Euchromatin, gene Euchromatin, [55,125,127] and the binding of the SWI/SNF remodeling5. The Epigenome complex.KDM6B at DSBs Moreover, (JMJD3)Relocalized and during the Heterochromatin TIE2-mediatedRelocalizedRelocalized DDR: Early Relocalized Epigenetic phosphorylation Events[56,125] in the Senescent [56,125] Response 5. TheH3K4me3H3K9ac Epigenome 5. TheH3K4me3 H3K9acatHAT/p300 EpigenomeDSBs and 5. TheH3K27me3 during H3K9acatHAT/p300 EpigenomeDSBsEuchromatin, DDR: and Earlyduring atHAT/p300 DSBsEuchromatin, Epigenetic DDR: and Earlyduring EventsEuchromatin, Epigenetic DDR: in the Early SenescentEvents Epigeneticfocused in area theResponse [55,125,127]SenescentEventsfocused in area theResponse [55,125,127]Senescent Response45,102,103,127–[55,125,127] KDM2B H3K9acKDM2Bactivation activation ↓Relocalized in ↑Relocalized in of H4Y51 [148], as well as the RNF20H3K9me3/40-mediated KDM4A H2BK120ub KDM4B Heterochromatin [149] and the RNF168-mediated 129][43,45,128,129] (MOF) Histone (MOF)DNA repair, Histone chromatin DNAHAT/p300 repair, chromatinEuchromatin focusedNI area focusedNI area H2AK15ub [144] act asHAT/p300 platforms toHAT/p300 recruitEuchromatin DDRIncreased proteins,Euchromatin activityNI like Heterochromatin ABL1NI NI [ 148 gene], Ku70 NI/ 80 [149[125,127]] and [125,127] [125,127] H4K16ac H4K16ac H4K8ac HAT/p300 Euchromatin NI [125,127]NI [125,127] [125,127][54,127] H4K8ac acetyltransferaseH4K8ac H4K20me3acetyltransferaseH4K8accompaction compaction 53BP1 [150,151]. The latter modification and theof successful Suv420h2 recruitmentrepression of 53BP1,↓Relocalized which controlsin ↓Relocalized in [43– H3K27me3 KDM6B (JMJD3) Heterochromatin 45,102,103,127– end re-sectioning, dictateHAT/p300 the choice ofHAT/p300 theEuchromatin, NHEJ KMT2/KDM5D/ pathwayEuchromatin,in spite Euchromatin, of HR [gene152 ,153focused]. H4K20me2 area [55,125,127]focused is area [55,125,127] H3K9ac H3K9ac 5. TheH3K4me3 Epigenome at DSBs and during DDR: Early EpigeneticRelocalized EventsRelocalized in the Senescent Response129][56,125] another5. The anti-resection Epigenome5. The modification at EpigenomeDSBs and5. The that during at Epigenome reinforceDSBs DDR: and 53BP1 Earlyduring atKDM2B DSBs bindingEpigenetic DDR: and Earlyduring to theEvents activationEpigenetic chromatin DDR: in the Early SenescentEvents [154 Epigenetic]. However,in theResponse SenescentEvents the in theResponse Senescent Response Increased activity Heterochromatin gene [54,127] co-presence of H2AK15UbHAT/p300 and H4K20me2 H4K20me3HAT/p300Euchromatin in the proximity(MOF) Euchromatin HistoneNI of theDNA DSBs repair,NI bu NIchromatin ff ers NHEJ NI processing [125,127] by [125,127] H4K8ac H4K8ac H4K16ac of Suv420h2 repression [125,127] recruiting the HAT Tip60/KAT5, which triggers H2AK15acacetyltransferase and 53BP1compaction displacement. An activity that KMT2/KDM5D/ Euchromatin, gene H3K4me3 Relocalized Relocalized [56,125] 5. The Epigenome5. The at EpigenomeDSBs and during H3K9acat DSBs DDR: and EarlyduringHAT/p300KDM2B Epigenetic DDR: Early EventsEuchromatin,activation Epigenetic in the SenescentEvents in theResponse Senescent Response [55,125,127] (MOF) Histone DNA repair, chromatin H4K16ac [125,127] H4K8ac acetyltransferaseHAT/p300 Euchromatincompaction NI NI [125,127]

H3K9ac HAT/p300 Euchromatin, [55,125,127] 5. The Epigenome at DSBs and during DDR: Early Epigenetic Events in the Senescent Response H4K8ac HAT/p300 Euchromatin NI NI [125,127]

5. The Epigenome at DSBs and during DDR: Early Epigenetic Events in the Senescent Response

Cells 2020, 9, 466 8 of 18 favors the establishing of HR [155]. 53BP1 displacement seems to occur during S/G2 transition, as in G1 cells 53BP1 stably occupying the HR sites [156]. The general loss of occupancy achieved in proximity to DSBs is counterbalanced by the distal accumulation of [156]. This PTM can be used as a scaffold for the binding of HDACs, thus ensuring the transcriptional silencing of genomic loci affected by DSBs [157,158]. A different epigenetic environment is associated to the activation of the HR pathway [158]. In this context, macroH2A is found to be more abundant than H2AZ, the acetylation of H2BK120 overcomes ubiquitylation in a SAGA complex-dependent manner [159] and γH2AX is more spread [156]. As explained above, Tip60/KAT5 acetylates H2AK15, thus displacing 53BP1 [155]. Tip60/KAT5 also maintains the acetylation of H4K16, which is required for keeping an open chromatin status [160] at the damaged site. An opposite epigenetic mark, H3K9me2, is required for the BARD1/HP1γ mediated retention of BRCA1 [161] and to allow the loading of the pro-resection factor CtIP [162]. In active genes affected by DSBs, the loss in and of H4 acetylation levels [156], as well as the recruitment of macroH2A [163] and of repressive protein complexes (HP1,KAP1,SUV39H1,PRDM2,HDACs), seems to compensate for the general loss of histones that characterizes the damaged sites [164]. The spatio-temporal regulation of chromatin remodeling at DSBs is achieved and sustained also by the deposition of other histone variants, like the HIRA-assisted H3.3 and the CAF-1-mediated H3.3 [23]. In summary, from the literature emerges a bleeding/vasoconstriction model in which the chromatin expands and becomes flexible at the damaged sites to accommodate the repair complexes, but heterochromatinization and the creation of a transcriptional repressive environment is required later on to allow the repair before restarting the transcription. The successful prediction of DSBs by looking at the histone positioning and PTMs [165], the emerging roles played by epigenetic regulators in DDR as well as the impact of epigenetic drugs on DDR [166] confirm that the epigenetic status of the chromatin not only identifies sites of genome fragility, but it is also causally linked to the successful repair [23].

6. An Anti-Apoptotic Response Sustains the Survival of Damaged Cells Exposed to Irreparable DNA Damage Senescent cells display an increased resistance to pro-apoptotic stimuli that is achieved through the up-regulation of the pro-survival gene Bcl2 and Bcl-XL and the down-regulation of the pro-apoptotic genes Bid and Bax [127]. The TP53-dependent up-regulation of the CDKi p21 plays a key role in sustaining the survival of damaged cells [167], while DNMT3a and HDAC1 are recruited on the p21 to switch off its expression during [168]. The increase in p21 promotes the cell-cycle arrest allowing the activation of DDR; cellular proliferation is subsequently permanently locked through the activation of INK4 CDKi (mainly p16 and p15) [169]. Many of the anti-apoptotic functions of p21 are achieved through the modulation of TFs (p300, STAT, JUN and TP53) [170] and are sustained by an epigenetic reprogramming [127]. In senescent cells, high levels of H4K20me3 and low levels of H4K16ac keep down the transcription of pro-apoptotic genes, while the increased acetylation of H4K16 characterizes pro-survival loci [127]. The heterochromatinization that surrounds and borders DSBs enhances these pro-survival responses and any relaxation of these structures, obtained for example through HDAC inhibition, triggers apoptosis [6]. Finally, the mTORC1 and PGC-1β dependent metabolic reprogramming [171] observed in senescent cells and in different models of aging [5] ensures energy supply. Since the epigenetic regulators require cofactors that are produced in a large majority in mitochondria [172,173], any mitochondrial dysfunction can affect both the transcriptional and the metabolic fitness of the cells and lead to senescence even in absence of DNA damage (MiDAS, mitochondrial dysfunction-associated senescence) [171]. Severe and acute stresses induce cell death, while prolonged and mild insults lead to cellular senescence and survival, probably because the cells have the time and a not completely compromised ability to mount the epigenetic, transcriptional and metabolic responses that characterize them [174]. Cells 2020, 9, 466 9 of 18

7. Final Considerations: The Link between Senescence, Aging, Epigenetics and DDR The accumulation of DSBs is a general hallmark of senescence and aging [6]. The main endogenous sources of DSBs are telomere attrition and replicative stress. Replication stress is commonly observed in RS, OIS and aging. In all these conditions cells slow down DNA synthesis and replication fork progression. However, the reduced replication fork speed activates dormant origin to preserve replication timing during replication stress [175]. This adaptive response allows the maintenance of an unaltered replication timing also in cells entering senescence [176]. On the other side it exposes common fragile sites (CFSs), which are genomic loci more prone to breakage after DNA polymerase inhibition, and the accumulation of genomic alterations. CFS alterations are typically observed in pre-neoplastic lesions [177]. Similarly, cells exposed to genotoxic agents (e.g., chemotherapeutic agents, pollutants and toxins) or to oxidative stress activate the DDR that frequently leads to cell-cycle arrest. Whatever the origin, the accumulation of irreparable DNA damage gives rise to a univocal response characterized by the activation of tumor suppressors and CDKi and by the release of pro-inflammatory cytokines [5]. Global histone loss, as well as the focused deposition of histone variants (H2AX, H2AZ, H2AJ, H3.3 and macroH2A) and the redistribution of H3K4me3, H3K27me3 and H3K36me3 characterize both DDR, DSBs and senescence. The chromatin remodeling observed in different senescence models seems to represent a temporal and spatial evolution of what is observed after a short-time treatment of the cells with DNA damaging agents. Although it is clear that an altered epigenome can expose cells to DSBs [134] and that epigenetic regulators control the fate of damaged cells [15,55,66,115,178–180], investigations on the epigenetic inheritance in daughter cells coming from DNA-damaged cells are still in their infancy [177]. Cancer cells appear as forgetful cells that have lost the epigenetic memory of a healthy genome. Aging seems to be predisposed to this memory loss. One of the major challenges of the future regarding the treatment of aging and cancer, will be the identification of the framework of epigenetic changes that can restore this memory.

Author Contributions: H.P., writing—original draft preparation; E.D.G., writing—original draft preparation; C.B., conceptualization, writing—original draft preparation, writing—review and editing, visualization, supervision, project administration, and funding acquisition. All authors have read and agreed to the published version of the manuscript. Funding: This study was supported by PRIN 2017JL8SRX “Class IIa HDACs as therapeutic targets in human diseases: new roles and new selective inhibitors” and Interreg Italia-Osterreich rITAT1054 EPIC to C.B. Conflicts of Interest: The authors declare no conflict of interest.

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