Epigenetics Recording Varied Environment and Complex Cell Events Is an Origin of 1 Cellular Aging 2 Xuejun Guo1*, Dong Yang2, X

1 Epigenetics Recording Varied Environment and Complex Cell Events is an Origin of

2 Cellular Aging

3 Xuejun Guo1*, Dong Yang2, Xiangyuan Zhang1

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5 1. State Key Laboratory of Environment Simulation, School of Environment, Beijing Normal

6 University, No. 19 Xinjiekouwai Street, Beijing 100875, China

7 2. Gene Engineering and Biotechnology Beijing Key Laboratory

8 College of Life Sciences, Beijing Normal University, No. 19 Xinjiekouwai Street, Beijing, 100875

9 China

10 *Corresponding author:

11 Xuejun Guo

12 Tel: 86-10-5880-7808

13 Fax: 86-10-5880-7808

14 Email: [email protected]

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22 23 Abstract Although the phenomenal relationship between epigenetics and aging phenotypic

24 changes is built up, an intrinsic connection between the epigenetics and aging requires to be

25 theoretically illuminated. In this study, we propose epigenetic recording of varied cell environment

26 and complex history could be an origin of cellular aging. Through epigenetic modifications, the

27 environment and historical events can induce the chromatin template into activated or repressive

28 accessible structure, thereby shaping the DNA template into a spectrum of chromatin states. The

29 inner nature of diversity and conflicts born by cell environment and its historical events are hence

30 recorded into the chromatin template. This could result in a dissipated spectrum of chromatin state

31 and chaos of overall gene expressions. An unavoidable degradation of epigenome entropy, similar

32 to Shannon entropy, would be consequently induced. The resulted disorder in epigenome,

33 characterized by corrosion of epigenome entropy as reflected in chromatin template, can be stably

34 memorized and propagated through cell divisions. Furthermore, hysteresis nature of epigenetics

35 responding to emerging environment could exacerbate the degradation of epigenome entropy.

36 Besides stochastic errors, we propose that epigenetics disorder and chaos derived from unordered

37 environment and complex cell experiences play an essential role in epigenetic drift and the

38 as-resulted cellular aging.

39 Keywords: Epigenetics; Environment; Cell events; Cellular aging; Epigenome entropy; DNA

40 methylation

41 1. Introduction.

42 Aging is the process of life becoming older characterized by debilitating losses of tissue or

43 cellular function. It refers to irreversible, progressive, and deleterious syndrome of changes that

44 occurs at molecular, cellular, tissue, and organismal levels (Johnson et al., 1999; Campisi, 2013). 45 The causes of aging can be assigned to all kinds of damage, which cause biological systems to fail.

46 These damages may be induced by toxic and nontoxic garbage accumulation, such as protein

47 cross-linking and aggregation, advanced glycation endproducts (AGEs), atherosclerotic and

48 amyloid plaques, inflammatory cytokines, lipofuscin, cortisol, metals, DDT, PCBs, etc

49 (Koschinsky, 1997). They are also derived from metabolic damage (i.e., free radicals, glycation),

50 telomere shortening, decline and inadequate antioxidant defense, defective cell cycle control,

51 declining efficiency of proteasomes, lysosomes, and heat shock proteins (Reiter, 2000; Yan, 1997).

52 Epigenetics refers to heritable changes in gene activity and expression without alterations in

53 the DNA sequence (Allis et al., 2015). Today, stable and long-term but not necessarily heritable

54 alterations in the transcriptional potential of a cell are also assigned to epigenetics (Calvanese et

55 al., 2009). Indexing the genome and potentiate signals from the environment, the chromatin in

56 eukaryotic organisms can be viewed as a dynamic polymer. This chromatin template is modified

57 by a variety of covalent and non-covalent modification. These modification processes include

58 post-translational histone modifications, chromatin-remodeling steps mobilizing or altering

59 nucleosome structures, the dynamic shuffling of histone variants, and the targeting role of small

60 ncRNAs. DNA itself can also be methylated usually at the cytosine residue of CpG dinucleotides

61 (Allis et al., 2015). All these mechanisms provide a set of interrelated pathways regulating the

62 accessibility of the chromatin template to the transcriptional machinery and ultimately determine

63 which genes are expressed and which are not (Pirrotta, 2015). These different patterns of gene

64 expression and silencing may be heritable through cell division and collectively contribute to

65 cellular phenotype (Allis et al., 2015).

66 Epigenetics has emerged as an important subject area in aging biology (Calvanese et al., 2009; 67 Huidobro et al., 2013; Horvath, 2013; Brunet and Berger, 2014; Lardenoije, 2015). The

68 phenomenal relationship between epigenetic drift, a gradual change away from baseline, and age

69 was proposed many years ago (Martin, 2005; Teschendorff et al., 2013; Issa, 2014). The

70 mechanism of epigenetic drift is generally ascribed to stochastic errors and imperfect fidelity in

71 maintenance of epigenetic marks. It is proposed that the fidelity of transmission of epigenetic

72 patterns is variable across the genome (Issa, 2014). Epigenetic drift is related to many of the aging

73 phenotypic changes. For example, genomic global DNA methylation decreases with age

74 (Berdyshev et al., 1967), whereas a number of specific loci become hyper-methylated during aging

75 (Oakes et al., 2003). Other important epigenetic factors, such as histone modifications, also change

76 during aging (Narita et al., 2003). Although the phenomenal relationship between epigenetic drift

77 and aging phenotypic changes are built up, the intrinsic nature of epigenetics causing cellular

78 aging and ultimately the organism aging is not yet fully elucidated. An intrinsic connection

79 between the epigenetics and aging requires to be theoretically illuminated.

80 Epigenetics mediates the relationship between the genome and the environment (Toyokawa

81 et al., 2012; Cooney, 2007; Robert et al.,2011; Sutherland and Costa, 2003; Steves et al., 2012). In

82 fact, human being is started with a fertilized egg with a single genome. Accommodating a plethora

83 of environmental signals, intrinsic and external stimuli, genome is epigenetically programmed to

84 hundreds of different types of cells with a remarkable multitude of distinct phenotypes (Aguilera

85 et al., 2010; Allis et al., 2015). Epigenetics responses and records all the cell environment and

86 events, including all types of environmental signals and changes, and a wide variety of intrinsic

87 and external stimuli (Baccarelli and Bollati, 2009; Sutherland and Costa, 2003; Barros and

88 Offenbacher, 2009; Feil and Fraga, 2012). Here we present a theoretical assay how epigenetics, 89 which stands at the crossroads of genetics and environment, is essentially related to aging. With

90 respect to the basic relationship between epigenetics and environment, we aimed to explain why

91 epigenetics will inevitably and ultimately cause aging, a long-standing mystery.

92

93 2. Environment and cell events may induce the opening or closing of chromatin template

94 through epigenetic modifications, thereby shaping the DNA template into a spectrum of

95 chromatin states.

96 We first depict how environmental cues and cell (i.e., transcriptional) events induce an

97 opening state of the chromatin template through epigenetic modifications. When an environmental

98 signal (external or internal) causes a specific transcriptional event (Alterts et al., 2008), the

99 initiated transcriptional event can concomitantly induce the underlying chromatin template from a

100 native state to an active and open state (Cavalli and Paro, 1999; Struhl, 1998). Responding to

101 environmental cues and transcriptional events, a number of dynamic and elaborate epigenetic

102 mechanisms combine together and interact closely to bring about an opening state of chromatin.

103 This process is accompanied by a series of activated epigenetic modifications, including histone

104 modifications, nucleosome remodeling and the replacement of core histones with histone variants

105 (Allis et al., 2015). An example of activated modification is histone acetylation, which is proposed

106 to neutralize the positive charges of highly basic histone tails and generate a localized expansion

107 of the chromatin fiber, thereby enabling better access of the transcription machinery to the DNA

108 double helix (Hong et al.1993). Histone acetylation is closely associated with the Pol II machinery,

109 thereby providing a simple mechanism to account for the general correlation between

110 transcriptional events and histone acetylation (Struhl, 1998). Onset of transcription, RNA II 111 polymerase may recruit specific KMTs (histone-modifying enzymes) to set some specific histone

112 methylations, such as H3K4me3 around the transcriptional start site and H3K36me3 within the

113 coding sequences (Sims et al., 2004; Smith and Shilatifard, 2013). Such histone modifications in

114 place are often represented as transcriptionally active chromatin (Sims et al., 2007). They are also

115 read by subunits of nucleosome remodeling complex, inducing the recruitment of nucleosome

116 remodeling machines and resulting in looping, twisting, and sliding of nucleosomes (Wysocka et

117 al., 2006). In concert with activated histone modifications, these nucleosome remodeling

118 mechanisms are particularly important for chromatin opening. Finally, the replacement of specific

119 core histones with histone variants may further facilitate the unraveling of chromatin template

120 upon transcriptional events (Weber and Henikoff, 2014).

121 We then consider how cell environment and its historical events induce the underlying DNA

122 sequence into a closed state. Repressive chromatin modification on a DNA sequence can be

123 specifically targeted by transcription factors, such as de novo DNA methylation (Brenner et al,

124 2005). This mechanism appears to be directly determined by environmental stimulus (external or

125 internal), which usually induce the on/off of transcription factors through a wide variety of

126 molecular pathways. Alternatively, in absence or at low frequency of specific environmental

127 signals and inductions, a related segment of DNA-sequence would not be frequently visited by

128 transcriptional factors and transcribed by RNA II polymerase. We propose the DNA segment in

129 this condition is inclined to be closely packaged by the nucleosomes, and gradually silenced by

130 another series of combinational epigenetic modifications. This consumption, although at a

131 molecular level, is similar to ‘use it or lose’ theory as put forward by Lamarckian. Nevertheless,

132 our hypothesis is reasonable since the major enzymatic systems catalyzing histone modifications 133 and DNA methylation have their counterpart enzymatic systems reversing the modifications. In

134 fact, much of evidence supporting such hypothesis has come from work on the de novo DNA

135 methylation. In presence of transcription factor and transcriptional events, de novo methylation of

136 CpG sites are abolished (Straussman et al, 2009; Gebhard et al, 2010; Lienert et al, 2011; Brandeis

137 et al, 1994; Macleod et al, 1994). When the binding sites to transcription factor are mutated, CpG

138 islands become to a methylated state (Brandeis et al, 1994; Macleod et al, 1994). Similarly, when

139 transcription factors binding to specific gene promoters are down regulated, the now-exposed CpG

140 sites can be targeted for DNA methylation (Lienert et al, 2011). Besides the establishment of DNA

141 methylation in CpG-islands from gene promoters and body regions, silencing epigenetic pathways

142 also involve histone tail de-acetylation, methylation of specific histone lysine residues

143 (particularly H3K9), recruitment of heterochromatin associated proteins (e.g., HP1).

144

145 3. Environment and cell history are in turn stably imprinted and propagated on chromatin

146 template through epigenetic modifications, and strongly determine the pattern of gene

147 expression.

148 As discussed above, cell environment and the history of a living cell (i.e., series of

149 transcriptional events) can induce a chromatin segment between on and off state through

150 epigenetic modifications, thereby shaping the DNA template into a spectrum of chromatin states .

151 These epigenetic signatures as-imprinted on chromatin template, in turn record a varied cell

152 environment and its complex historical events. Epigenetic modifications offer a molecular

153 explanation for the memorization and inheritance of acquired traits induced by the environment

154 and the past. They actually mirror the historical cell events and environmental changes, 155 significantly contributing to phenotypic variation. Epigenetic signatures in chromatin can be

156 viewed as marks of epigenome recording the cell environment and the antecedent events, and in

157 turn strongly determined the accessibility and expression potential of a chromatin region. When

158 these marks are stably recorded onto the template, they can be memorized, propagated, and

159 transmitted over many somatic cell divisions (Allis et al., 2015；Nakayama et al., 2001；Margueron

160 et al., 2009；Song et al., 2011；Kaati et al., 2012). Some acquired traits could even be

161 trans-generational transmitted in the sense of Lamarckian evolution (Kaati et al., 2012).

162 Classical genetics considers a panel of TFs is responsible for activation and initiation of gene

163 expression. But the availability and binding of TFs is transient and will be lost immediately.

164 According to classical genetics, persistent gene expression requires persistent availability of TF.

165 However, the current epigenetics fully recognizes that the transcriptional state (repressive or active)

166 is strongly determined by epigenetic modifications. Gene expression pattern is actually controlled

167 epigenetically rather than genetically (Calvanese et al., 2009; Laurent et al., 2005). Epigenetics

168 strongly impacts the gene expression by regulating the accessibility of the underlying DNA

169 template to the transcriptional machinery. When a primary signal from environment (external or

170 internal) and a historical transcriptional event induce the opening the underlying DNA sequence,

171 this opening of local structure can be stably memorized through several cycles of cell divisions

172 even when the initial signal and TFs are not in presence anymore (Allis et al., 2015). The

173 chromatin template recorded with historical events and cell environment thereby significantly

174 impact the pattern of gene expression and greatly determine the further response to emerging

175 environment.

176 177 4. Varied environment and complex cell events would inevitably result in a dissipated

178 spectrum of chromatin state and lead unavoidably to the chaos of overall gene

179 expressions.

180 The environment related to a living cell prefers to all internal and external factors influencing

181 its survival, growth, division and differentiation. Cells in either unicellular or multicellular

182 organisms live in an unpredictably and variable environment. They are exposed to all kinds of

183 environmental factors in the whole living history (Fig. 1a). These varied environmental

184 factors/conditions can either be biotic and abiotic. Abiotic factors include temperature, pH, redox,

185 ionic concentrations and nutrient availability, etc. Biotic factors can be physiological, including

186 intracellular or extracellular signaling molecules, energy and metabolism homeostasis, chemotaxis,

187 healthy and aging states, and so on. Biotic factors can also be pathological, including oxidative

188 stress, toxic compounds, UV radiation, osmotic pressure, all types of wounding, inflammatory

189 cytokines, pathogen infection, and so on (Baccarelli and Bollati, 2009; Sutherland and Costa, 2003;

190 Barros and Offenbacher, 2009; Feil and Fraga, 2012; Yang et al., 2018). The historical events of a

191 cell prefer to all the cellular processes have been occurred (Fig. 1b). The history of a cell life is

192 extremely complex and involves a variety of cellular events. They include information processing

193 (i.e., transcription, translation, and replication) and cell signaling, growth and differentiation,

194 metabolism, division, protein synthesis, and so on. Throughout the whole life, cells are doomed to

195 undergo a variety of exogenous stresses and pathological attacks (Alberts et al., 2008; Feng et al.,

196 2016). 197

198 Fig. 1 (a) Cells live in an unpredictably environment comprising many diversified

199 environmental factors and (b) undergo a wide variety of cellular historical events.

200 Generally in a eukaryotic cell, each regulatory gene regulates the expression of a number of

201 genes. Meanwhile the expression of this regulatory gene is usually regulated by many other

202 regulated proteins. Tens of thousands of interactions between genes are organized into very

203 complex networks that help to coordinate the cell's activities and relay signals into the cell from

204 the cell's environment. Many a time, a regulated gene is involved in a multiple of cellular

205 pathways. Through these molecular cell pathways, cells can response and adapt to a wide variety

206 of environmental factors. To answer a change of specific environmental factor such as EF I, we

207 assume a specific collection of gene variations are involved. This collection of gene assembly,

208 through elaborate up-regulation or down-regulation of gene expression, collaborates and

209 coordinates together to cope with the variation of a specific environmental parameter (Fig. 2, a). 210 But actually, cells are exposed simultaneously to some other different environmental factors. The

211 expression pattern of a specific gene, that is up-regulated by one environmental factor, could be

212 either down-regulated antagonistically or up-regulated synergistically by another environmental

213 factor (Fig. 2, b). These environmental factors are usually characterized with diversity and in

214 conflicts. In fact, unpredictable variation with diversity is an intrinsic nature of different

215 environment factors. The assembly pattern of gene expression must answer all of these different

216 environmental factors, and has managed to reconcile between those diversity, discordance and

217 conflicts (Fig. 2, c). Imprinted by these varied environmental factors, a chaos of epigenetic states

218 would be generated in the related regions of chromatin. An irreconcilable conflict between the

219 ideal genetic regulation and the suboptimal epigenetic state would be conveyed to chromatin

220 template in answering each environmental factor (Fig. 2, d). A perfect match between the actual

221 epigenetic states and ideal genetic regulations in answering each specific environmental factor

222 (here refers to EF I) actually does not exist (Fig. 2, e). This awkward situation can even be

223 generated by overlying of environment factors with temporal difference, considering the

224 memorable (also hysteresis and irreversibility) property of epigenetic modifications. An

225 environmental factor changing in the opposite direction at different time can also produce such

226 discrepancy between the real epigenetic states and ideal genetic regulations.

227 228

229 Fig. 2 Varied environment factors produce irreconcilable conflict between the ideal

230 genetic regulation and the suboptimal epigenetic state. (a) a specific collection of gene variations,

231 up(↑) or down(↓) regulated, are involved to answer a specific environmental factor (EF I) ; (b) But actually,

232 cells are exposed to and inevitably have managed to answer some other different environmental factors (i.e.,

233 EF II, EF III, EF IV, EF V, …); (c) both the synergistic (↑ or ↓, vertical arrows) and antagonistic (↔,

234 horizontal arrows) effect can be generated in the assembly pattern of gene expression in answer to all of these

235 varied environmental factors; (d) Imprinted by these varied environmental factors, a spectrum of epigenetic

236 states could be generated in the related regions of chromatin (‘A’, activated epigenetic state; ‘HA’, highly

237 activated epigenetic state; ‘R’, repressive epigenetic state; ‘HR’, highly repressive epigenetic state; ‘M’,

238 medium epigenetic state); Consequently, the irreconcilable conflict between the ideal genetic regulation (a)

239 and the suboptimal epigenetic state (d) would be conveyed in answering each environmental factor; (e) a

240 perfect match between the ideal genetic regulations and actual epigenetic states in answering each specific

241 environmental factor (here refers to EF I) actually does not exist.

242

243 The dilemma conditions as resulted make cells always in sub-optimal situations in response 244 to the complex environment and become more sensitive to all aspects of damage sources and

245 threaten factors. More importantly, when cells are exposed to superposition of such varied

246 environmental factors and complex cell events for a sustained period of time, all traits of their

247 disorder would be recorded onto chromatin template through various epigenetic mechanisms as

248 depicted above. The irreconcilable conflict between the desired genetic regulation and the

249 suboptimal epigenetic state would be conveyed to the chromatin template. When the wide

250 diversity of environment factors and the complex history events are mapped together to the

251 chromatin template by the combinational epigenetic modifications, a dissipated spectrum of

252 chromatin state could be produced unavoidably. Mapping ultimately to the chromatin template,

253 such chaos and disorder would inevitably lead to a gradual degradation of “epigenome entropy”.

254 We now consider building a model of epigenome entropy. Epigenome is a semiosis system

255 with many similarities with languages and computer system. Thereby, the information (or

256 Shannon) entropy can be used as reference for the definition of epigenome entropy. Since

257 Shannon entropy is negatively related to thermodynamics entropy, cells at age of zero are

258 consumed to have the maximum value of epigenome entropy. The resulted disorder in chromatin

259 template with age growth actually reduces epigenome entropy. For a specific epigenetic

260 mechanism such as DNA methylation, the reduced epigenome entropy in a single cell is proposed

261 to be represented as equation (1):

1 1 1 262 ∆퐸푛푡푟표푝푦 = − ∑푛 푊 ∗ 푙표푔 ( ) = ∑푛 푊 (1) 퐷푁퐴 푚푒푡ℎ푦푙푎푡𝑖표푛, 푠𝑖푛𝑔푙푒 푐푒푙푙 𝑖=1 𝑖 2 2 2 2 𝑖=1 𝑖

263 Where i refers to the sequence number of CpG sites, where the status of DNA methylation,

264 demethylation or methylation, has changed in response to changing environment and cellular

265 events. Because the gene regulation impact of each change in DNA methylation at different 266 chromatin site is different, we introduce a weighting coefficient Wi to the model. For the sake of

267 computability, the values of Wi can be assigned to a few fixed values based on the genome regions

268 of CpG sites (i.e., gene promoter, gene body and inter-genetic region). The reduced epigenome

269 entropy derived from DNA methylation for a cell assembly with J cells can be represented as

270 equation (2):

1 푛1 1 푛2 1 푛푗 271 ∆퐸푛푡푟표푝푦 퐷푁퐴 푚푒푡ℎ푦푙푎푡𝑖표푛, 푐푒푙푙 푎푠푠푒푚푏푙푦 = ∑ 푊𝑖 + ∑ 푊𝑖 + ⋯ + ∑ 푊𝑖 (2) 2 𝑖1=1 1 2 𝑖2=1 2 2 𝑖푗=1 푗

272 For a homogenous cell assembly, equation (2) can be simplified to equation (3):

1 273 ∆퐸푛푡푟표푝푦 = 퐽 ∗ ∑푛 푊 ∗ √(퐷 − 퐷 )2 퐷푁퐴 푚푒푡ℎ푦푙푎푡𝑖표푛, 푐푒푙푙 푎푠푠푒푚푏푙푦 2 𝑖=1 𝑖 𝑖,푡 𝑖,0

274 (3)

275 Di, t is the percent of DNA methylation for site i at time point t; Di, 0 is the percent of DNA

276 methylation for site i at original time.

277 The quantification of reduced epigenome entropy from some other epigenetic modifications,

278 i.e., all forms of histone modifications, can be obtained using the similar discipline as above. The

279 “epigenome entropy” should be inevitably decreased. The resulted chaos would gradually

280 accumulate in a living cell, or even pass to its offspring cells, since the epigenetic modifications

281 can be memorized, propagated, and transmitted over many somatic cell divisions. Note that the

282 environmental factors and history events here prefer to those lasted for a time period and

283 occurring at a certain intensity which can produce distinct chromatin alternations.

284

285 5. Irreversibility and hysteresis of epigenetics in response to emerging environment and

286 new events may aggravate the dissipated spectrum of chromatin state with increased loss

287 of epigenome entropy 288 One defining characteristic of epigenetics is relative irreversibility and hysteresis (Laurent et

289 al., 2005; Nagaraj et al., 2014). Many epigenetic marks may persist through several rounds of cell

290 division, and a few could even be inherited as germ line modifications. A typical example is

291 shown in the reprogramming field. In recent years, one of the most influential discoveries is that

292 somatic cells can be induced to become pluripotent stem cells in tissue culture (Takahashi and

293 Yamanaka 2006). However, the efficiency of reprogramming was actually very low <0.1%.

294 Certain somatic epigenetic modifications, such as repressive H3K9me3 and DNA methylation are

295 the major obstacle, which are very difficult in reprogramming. These epigenetic modifications are

296 stably transmitted through somatic cell divisions and some even resist reprogramming in the

297 oocyte.

298 In fact, hysteresis occurs ubiquitously in biology in different spatiotemporal scales.

299 Epigenetics has many characteristics of a non-linear bistable system, exhibiting distinct hysteresis

300 effects and the associated bifurcation diagram (Noori, 2014). Darlington ever discussed hysteresis

301 in his classic works on genetics, which occurs as failure of the external form of the chromosomes

302 to respond immediately to the internal stresses of the chromosomes (Darlington, 1937). In cells

303 with distinct epigenetic modifications, gene expression is actually controlled by the combinational

304 functions of both epigenetics (external form) and genetics (internal stresses). Epigenome carries

305 the chromatin signatures as memory of historical environment and events, and subsequently affect

306 expression pattern of genes responding to new environment and emerging events. The hysteretic

307 nature of epigenetics makes the current state of underlying chromatin, which has been recording

308 the historical environment and events of a cell, a key determinant of gene expression pattern.

309 Recorded with the past environment and cell history, cells modify the chromatin template to 310 accommodate the new environment and emerging events. It means that epigenetic modifications

311 are not entirely responsive to the present environment stress, but seems to make compromised

312 epigenetic modifications between the existing epigenetic records and genetic stresses derived from

313 emerging environment. This is similar to two component forces (epigenetic records and genetic

314 stresses) producing a resultant force (emerging epigenetic modifications) (Fig.3, a). As shown in

315 Fig. 3, b, epigenetic hysteresis may potentially result in a range of epigenetic states when

316 confronted with a specific genetic stress, which is dependent on historical cell events and

317 environment changes. Hysteresis thereby results in significant epigenetic drift for different cells,

318 where the possible pattern of epigenetic states at different gene locations varies greatly for

319 different cells corresponding to environment change (Fig. 3, c). To answer one changing

320 environment, the epigenetic state for the collection of genes involved would be modified, but with

321 different hysteretic paces. As shown in Fig. 3, d, the epigenetic state of cells at each gene location

322 is first at their beginning position. After a series of round-trip change in cellular events or

323 environmental parameters, dissipated patterns of epigenetic states could be generated due to

324 different hysteretic degree of epigenetic modifications at different gene locations. Although

325 epigenetic hysteresis makes gene expressions more resistant to noise, it inevitably leads to a

326 dissipated spectrum of chromatin state with degradation of epigenome entropy as an inevitable

327 side effect. 328

329 Fig. 3. Irreversibility and hysteresis of epigenetic modifications in response to new

330 environment and emerging events would inevitably lead to a dissipated spectrum of

331 chromatin state with increasing loss of epigenome entropy. (a) cells make compromised epigenetic

332 modifications between the epigenetic records and stresses derived from environment; (b) Epigenetic hysteresis

333 may produce a range of epigenetic states (from E1 to E2) when confronted with a specific genetic stress (G1); (c)

334 Hysteresis thereby results in epigenetic drift, where the epigenetic states ( Eps1, Eps 2, Eps3, …, EpsX) of

335 different cells at each gene locations (Gn1, Gn2, Gn3, …GnX) vary greatly in a specific environment (Gn: gene;

336 Eps: epigenetic state); (d) The epigenetic state at each gene location (symbol of five-point stars) is first at their

337 beginning position. After a series of round-trip change (R1, R2, R3, …, RX) in specific cellular events or

338 environmental parameters, dissipated patterns of epigenetic states are generated due to the different degree of

339 hysteresis for different gene locations.

340 6. Epigenetics modifications on single copy chromatin lack the mechanisms of

341 error-epigenetic checking and erasing. 342 To maintain in a homeostatic and health state, life has a myriad of checkpoints, error

343 correcting mechanisms and immunities to defend against all kinds of damages (Johnson et al.,

344 1999; Alberts et al., 2008). Combating metabolic damage such as free radicals and glycation, life

345 can create fewer free-radicals by more efficient mitochondria. They may use less energy to live,

346 have more effective antioxidant defenses, better DNA protection and DNA repair. Stem and germ

347 cells contain telomerase to prevent telomere shortening. Animals can have a better immune system

348 and detoxify more effectively in the liver tissue. To avoid garbage accumulation, damaged and

349 misfolded proteins are eliminated by the enzymatic and proteolytic proteasomes. Lysosomes are

350 responsible for degradation of aging mitochondria.

351 Unlike the error correcting mechanisms combating all aspects of damages as listed above, so

352 far there is no evidence in mechanisms of error checking for epigenetics modifications. Epigenetic

353 modification is a dynamic process and not sustained over an indefinitely long term. Most of the

354 characteristic epigenetic marks can be reset during the course of differentiation (Kohli and Zhang,

355 2013). However, all of the mechanisms of epigenetic modifications, including those putting

356 chromatin marks in place, maintaining and responding to them, are firstly based on the existing

357 chromatin states, which have been imprinted with past events and environment. It’s the past

358 environment and cellular events superposed with ongoing genetic stresses that determine the new

359 assembly pattern of epigenetic modifications. Except for germ cells, most somatic cells lack the

360 molecular mechanisms of checking and erasing the epigenetic modifications. There are no cellular

361 pathways to recover the lost epigenome entropy with age growth. Although the DNA double helix

362 provides potential mechanisms for DNA replication and repair, and for the maintenance and

363 propagation of DNA methylation (Song et al., 2011), it does not provide mechanisms for 364 error-checking or erasing of epigenetic modifications in either histone or DNA sequence level.

365 Epigenetic disorders are eliminated only in the germ cells. The renewing of epigenetic

366 modifications is carried out in embryos of new life, but not in any adult animal cells.

367 Through deposition of various epigenetic modifications, the DNA double helix records the

368 cell history and responds to the changing environment. As illustrated above, epigenetic

369 modifications are regulated by an array of delicate molecular machines, including DNA-binding

370 interactions, histone modifications, histone variants, nucleosome remodeling, DNA methylation,

371 and ncRNAs (Allis et al., 2015). On one hand, this highly organized and dynamic polymer can be

372 viewed as a single molecule because for each specific DNA sequence only one-copy is existed in

373 cell nucleus. On the other hand, the process of epigenetic modifications is extremely elaborate and

374 intricate, requiring the assembly of so many multi-protein complexes. It’s the low-affinity

375 associations of hundreds of multi-proteins along a DNA sequence. Thereby, one would expect that

376 stochastic factors should play a substantial role in depositing epigenome disorder and chaos onto

377 the chromatin. The stochastic errors and imperfect fidelity in maintenance of epigenetic marks are

378 generally thought to be the main mechanism of epigenetic drift, a gradual change away from

379 baseline, and aging (Martin, 2005; Teschendorff et al., 2013; Issa, 2014). Besides stochastic errors,

380 in this assay we propose that epigenetics disorder and chaos imprinted by varied environment and

381 complex cell events play an essential role in epigenetic drift and the as-resulted cellular aging.

382

383 7. Cellular aging as resulted and its implications on organismal aging and cell life

384 expectancy

385 Mechanisms behind cell aging are extensively addressed, which include telomere shorting, 386 genomic and epignenomic damage, oxidative stress, unbalanced mitogenic signals, and so on

387 (Johnson et al., 1999; Campisi, 2013). In this assay, we depicted the inner nature of epigenetics

388 recording historical events of a living cell and its associated environment. We then propose that an

389 inevitable dissipated spectrum of chromatin state with degraded epigenome entropy as imprinted

390 by complex history of a living cell and variable environment factors will ultimately cause cellular

391 aging. From this point of view, we propose cellular aging is inherently rooted in epigenetics, not

392 requiring any specific hormonal signaling and transcriptional programing, although we recognize

393 specific mitogens and proliferation-associated genes are involved.

394 Cellular senescence/aging is thought to play an essential role contributing tissue and

395 organismal aging (Ben-Porath and Weinberg, 2005; Tchkonia et al., 2013). Senescent cells with

396 degraded epigenome entropy could ultimately induce tissue and organismal aging. We believe it is

397 epigenetic recording of unpredictable environment and complex cell events that determine the

398 inevitable aging of cells. In this epigenetic point of view, one could predict that life expectancy of

399 a living cell in animals is strongly impacted by stability of its living experience and surrounding

400 environment, the plasticity of its epigenetic mechanisms, and the complexity of its physiological

401 function. A living cell in animals is expected to have relatively longer life expectancy if its living

402 experience and surrounding environment are stable, if its epigenetic modification is more plastic,

403 and if its physiological functions are relatively simple and narrow.

404 Cell life expectancy ∝ stability of its living experience and surrounding environment ×

405 plasticity of its epigenetic modification ÷ complexity of its physiological function

406 As an example, the environment of living niche for stem cell is very stable. Its epigenetic

407 state is quite plastic (Hemberger et al., 2009). The physiological function of stem cell is majorly 408 division and renewal, which is supposed to be relatively simple and specific. Thereby, stem cell

409 usually has a longer life expectancy. Similarly, neurons in human brain have a long life expectancy

410 likely due to its highly stable living environment and its highly specified function as processing

411 and transmitting electrical and chemical signals. However, hepatocyte lives in an unpredictable

412 environment with all types of variable stresses. Hepatocyte in the human body is responsible for a

413 very comprehensive function, including protein synthesis, detoxification, and carbohydrate and

414 lipid metabolism (Klaassen, 2008). Epigenome entropy in hepatocyte is likely to degrade more

415 rapidly, as imprinted by its multi- physiological processes and variable environment stresses.

416 Therefore, hepatocytes in animals often have a relatively short life expectancy.

417 Acknowledgment

418 This work was funded by the National Key R&D Program of China (2017YFA0605001), the

419 National Natural Science Foundation of China (91547207), and the Fund for innovative Research

420 Group of the National Natural Science Foundation of China (51721093).

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