EPIGENETIC REPROGRAMMING

(WikiLink: Epigenetics) - (Last Revision: 6/7/2022)

Epigenetics describes all reversible, environmental, psychological, physiological and heritable processes that regulate gene expression without altering the DNA sequence.

In the broadest sense, Epigenetics refers to processes that influence medium to long-term gene expression by changing the readability and accessibility of the genetic code. The source of the word “Epi” is Greek and means over or on top of. This process changes the readability by adding or removing “marks” to DNA and in some cases RNA. These marks consist primarily of methyl molecules. The absence of these markers leaves the reading frame of the DNA accessible to being transcribed, the addition of the methyl group blocks transcription. Methyl groups are deposited by DNA methyltransferases and removed by demethylases. Methylation of DNA controls our age state and as a direct consequence; diseases of aging. Emerging data from research in this arena is beginning to support an age regression strategy that would directly reprogram these methylation alterations of DNA and mRNA.

Mounting evidence indicates that specific modifications in epigenetic marks are responsible for cellular and organismal aging. These alterations are mediated by several enzymes that act as readers and modifiers, particularly histone methyltransferases (HMTs), demethylases (HDMTs), acetyltransferases (HATs), and deacetylases (HDACs). Short chain amino acids or peptides are also able to accomplish both addition and deletion of the marks. (see peptides) The cellular epigenetic state is a dynamic interplay of all these components and changes over time and with environmental stimuli. Overall, epigenetic variations are ubiquitous regulators of the aging process in various of organisms across several kingdoms. [8].

Horvath and his team looked at the DNA methylation changes with age in 59 different tissue types from 128 mammalian species to determine commonalities.

It was found that a highly conserved aging program driven by DNA methylation changes that downregulate genes that produced transcription factors by adding methyl groups to the promoter area of the genes. Thirty six genes (36) genes that were effectively shut down by DNA methylation and almost all of them were transcription factors that are involved in the differentiation of cells during development that have homeobox domains. Very few genes (12) that experienced the loss of methylation which was a surprise to me based on my predictions in my 1998 paper. apparently, most DNA methylation is uninvolved with direct aging control.

What can affect epigenetic marks? Quite literally everything. This is demonstrated throughout this page and all of the peer reviewed research available on the subject. Programatically altering these marks has been demonstrated to reverse disease processes and aging. The image below graphically demonstrates the association of environmental factors with these marks.

**Click [√] to Enlarge Source [10]**
**Clocks**
Studies clearly show that genome-wide DNA methylation levels are associated with chronological age throughout the human lifespan. Some age-related DNA methylation changes occur in specific regions of the genome and are directional, suggesting the existence of differentially methylated regions associated with age. Thus, biomarkers based on DNA methylation can accurately estimate age, as has been demonstrated by many investigations involving tissues, individuals, and populations.
**◉ The reversibility of DNA methylation is the most interesting feature of epigenetic clocks, which suggests that they can be used to measure the effectiveness of anti-aging interventions [19]**

Epigenetic Reprogramming and Rejuvenation

Epigenetic Dictionary

Epigenetic mechanisms

DNA methylation

DNA methylation is one of the most studied epigenetic mechanisms. It occurs at the level of DNA and regulates the expression of genes. DNA methyltransferases (DNMTs) are the enzymes responsible for DNA methylation and carry out the transfer of a methyl group onto the 5th carbon of particular stretches of DNA called CpG islands [2 (https://www.mdpi.com/20734409/12/8/1163)].

DNA methylation is reported to regulate transcriptional initiation activity while DNA hypomethylation is associated with higher transcriptional activity. The ten–eleven translocation (TET) carries out the catalyzation of the hypomethylation using α-ketoglutarate, Fe(II), and oxygen as substrates.

Histone modiﬁcations

Packaging of the eukaryotic genome into a complex structure known as chromatin ensures the compactness of the genome as well as the accessibility of RNA polymerases and regulatory proteins to the DNA. The nucleosome is the fundamental unit of the chromatin which is made up of an octamer of histones comprising two copies each of histone H2A, H2B, H3, H4, and other variants of histones and surrounded by 147 DNA base pairs [2 (https://www.mdpi.com/20734409/12/8/1163)]. A histone linker H1 binds to the DNA that moves in and out of its 1.65 turns around the nucleosome. Histones undergo several modiﬁcations such as acetylation, methylation, phosphorylation, ubiquitination, ribosylation and sumoylation. These modiﬁcations help in the existence of various spatial chromatin rearrangements which in turn inﬂuence the interaction of the DNA with RNA polymerase II and other transcription factors.

Histone methylation

Histone methylation is involved in the silencing or activation of gene transcription depending on the number of methyl groups attached to speciﬁc amino acid residues (lysine and arginine). Histone methyltransferases (HMTs) carry out the methylation of the residues where lysine can be mono-, di- and trimethylated and arginine can be monomethylated, or asymmetrically/symmetrically dimethylated. Although initially methylation was assumed to be irreversible, the identiﬁcation of two families of histone demethylases proved it to be otherwise.

Histone acetylation

Histone acetylation promotes a more relaxed chromatin structure and initiates transcriptional processes by removing positive charges from lysine residues at histone tails. Histone deacetylation promotes a closed chromatin structure which suppresses the transcriptional processes. Histone acetylation is carried out by histone acetyltransferases (HATs) and deacetylation by histone deacetylases (HDACs).

Histone phosphorylation

Histone phosphorylation is a post-transcriptional modiﬁcation carried out by various enzymes. Histone phosphorylation is associated with gene expression, transcription regulation, and DNA damage response (DDR).

Histone ribosylation

Histone H1 as well as all the core histones can undergo ribosylation. Identiﬁcation of About 22 members of the ADP ribosyltransferase (ART) superfamily, as well as several members of the sirtuin (SIRT) family, has taken place that possess mono-ADP-ribosylation properties. Histone ribosylation has been reported to play important roles in transcription and replication.

Histone ubiquitination and sumoylation

Histone ubiquitination takes place with the help of E1-activating enzymes,

E2 conjugation and E3 ligases. These result in the addition of ubiquitin (Ub) to a lysine (Lys) residue on proteins or Ub itself, leading to the formation of polyUb chains. On the contrary, deubiquitinating enzymes (DUBs) carry out the removal of Ubs from the residues.

Histone sumoylation is another post-traditional modiﬁcation that has been discovered recently. Their effects on gene expression and chromatin organization are not as well known as other posttranscriptional modiﬁcations

Since epigenetic modifications are transient, epigenetic reprogramming, adding or deleting marks is a promising approach to increase healthspan and lifespan.

**Click [√] to Enlarge Source:[9] [2020]** **Epileptic Disorders: International Epilepsy Journal with Videotape**
**Keywords: aging, CpG, epigenetics, longevity, methylation. marks**

Ten-Eleven-Translocation (TET)

Recently discovered, TET (ten-eleven translocation) enzymes have been found to aid in the de-methylation process or the removal of methyl groups from cytosine. This reversal of cytosine methylation is promising in age regression as well as disease control.

Changes to DNA methylation patterns over time form the basis of 'aging clocks', but whether old individuals retain information to reset the clocks and, if so, whether it would improve tissue function is currentely one of aging researches most intense areas of focus. . Of all the tissues in the body, the central nervous system (CNS) is one of the first to lose regenerative capacity. Using the eye as a model tissue, we show that expression of Oct4, Sox2, and Klf4 genes (OSK) in mice resets youthful gene expression patterns and the DNA methylation age of retinal ganglion cells, promotes axon regeneration after optic nerve crush injury, and restores vision in a mouse model of glaucoma and in normal aged mice. This process, which we call the reversal of information loss via epigenetic reprogramming or REVIVER, requires non-global, active DNA demethylation by TET enzymes and the downstream enzyme TDG, indicating that alterations in DNA methylation patterns may not simply indicate age, but participate in aging. Thus, old tissues retain a faithful record of youthful epigenetic information that can be accessed for functional age reversal. [1]

TET-mediated oxidation of 5mC. TET enzymes successively oxidizes 5mC to 5hmC, 5fC, and 5caC. The oxidation depends on the presence of Fe(II), molecular oxygen, and α-KG as cosubstrates . Vitamin C and ATP enhance the enzymatic activity of TET. TET enzymes and 5mC oxidation derivatives are involved in various molecular events (DNA demethylation and regulation of gene expression) and therefore play key roles in embryonic development and oncogenesis. Patient-derived mutations of enzymes in TCA cycle, including IDH1/2, SDH, and FH lead to accumulation of metabolites 2-HG, succinate, and fumarate. These α-KG analogs competitively inhibit the activities of TET enzymes and JmjC-containing histone demethylases and therefore may contribute to oncogenesis

Epigenetic control of Biological Age

⫸ 1) Age can be accurately estimated by epigenetic clocks based on DNA methylation profiles from almost any tissue of the body.

2) Since such pan-tissue epigenetic clocks have been successfully developed for several different species, it is difficult to ignore the likelihood that a defined and shared mechanism instead, underlies the aging process.

Subsequent successes in developing similar pan-tissue clocks for other species hint at the universality of the aging process.

3) We identified and characterized specific cytosines, whose methylation levels change with age across mammalian species.

The human genome contains approximately 28 million CpG dinucleotides, in which a cytosine precedes a guanine in the 5'-3' direction. Each of these cytosines can be modified through the addition of a methyl group to create 5-methylcytosine, termed DNA methylation (DNAm). In most contexts, a majority of these sites are indeed methylated, with the specific pattern of methylation varying across cell types, individuals, and conditions[1–3]. [21]

Cytosine (symbol C or Cyt) is one of the four nucleobases found in DNA and RNA, along with adenine, guanine, and thymine (uracil in RNA).

Cytosine can also be methylated into 5-methylcytosine by an enzyme called DNA methyltransferase or be methylated and hydroxylated to make 5-hydroxymethylcytosine.

The CpG sites or CG sites are regions of DNA where a cytosine nucleotide is followed by a guanine nucleotide in the linear sequence of bases along its 5' → 3' direction. CpG sites occur with high frequency in genomic regions called CpG islands (or CG islands).

4) Genes associated with these cytosines are greatly enriched in mammalian developmental processes and implicated in age associated diseases.

5) Collectively, these new observations support the notion that aging is indeed evolutionarily conserved and coupled to developmental processes across all mammalian species.

6) To investigate this, we sought to i) identify and characterize cytosines whose methylation levels change with age in all mammals and ii) develop universal age-estimators that apply to all mammalian species and tissues (universal epigenetic clocks for mammals).

7) Consistently across all tissues, there were more CpG’s with positive correlations with age than negative ones (Extended Data Fig. 1) and most of them were located within CpG islands, which are known to become increasingly methylated with age.

8) While many of these cytosines were either specific to individual organs or shared between several organs, 54 potential universal age-related CpGs were shared among all the five organs (Fig. 1e, Extended Data Table 1). Strikingly, the overwhelming majority of the 36 genes that are proximal to these 54 CpGs encode transcription factors with homeobox domain, and are involved in developmental processes.

9) Genes that are proximal to age-related CpGs, overwhelmingly represent those involved in the process of development. A large body of literature connects growth/development to aging starting with the seminal work by Williams 1957 11.

10) Therefore, methylation regulation of the genes involved in development (during and after the developmental period) may constitute a key mechanism linking growth and aging. The universal epigenetic clocks demonstrate that aging and development are coupled and share important mechanistic processes that operate over the entire lifespan of an organism.

⫷[20]

(TET) TEN-ELEVEN-TRANSLOCATION

Know Supplemental Epigenetic Modifyers

Vitamin C

Uridine

AKG?

GHK-Cu?

di, tri, and tetra peptides

Thymic Regeneration: https://onlinelibrary.wiley.com/doi/10.1111/acel.13028