Epigenetics: Science of Changes without Change in DNA Sequences

Abstract

The mechanisms for epigenetic modifications include modification of histone proteins or modifications of the DNA itself (not affecting the DNA sequence). These include acetylation, methylation, phosphorylation, SUMOylation, ubiquitylation, etc. For example, DNA methylation (cytosine methylation) or histone acetylation (lysine acetylation). Recent studies have indicated that the activity of non-coding RNAs, such as microRNAs, long non-coding RNAs, and small interfering RNAs also affects epigenetic mechanisms. In a genome, the collection of all the modifications that regulate gene expression is called its epigenome. Improper occurrence of the epigenetic mechanisms can lead to deleterious health and behavioral effects. For instance, the most studied epigenetic modification is DNA hypermethylation, which leads to the silencing of antitumorigenic genes, and this has been shown to cause cancer. Various techniques are employed for DNA methylation profiling such as pyrosequencing, bisulfite-PCR, ChIP seq (Chromatin Immunoprecipitation), bisulfite seq, and specialized RNA seq. This chapter will introduce epigenetics, describe the different epigenetic mechanisms, and discuss in brief how to study these mechanisms and their effects on the plant as well as human health.

1. Introduction

Epigenetics coined by Dr. Conrad H. Waddington is a branch of biology that studies the changes occurring in organisms resulting from changes in gene expression instead of the genetic sequence. Epigenetic mechanisms, some of which are reversible, can thus alter the phenotype without affecting the genotype. Epigenetic mechanisms regulate gene expression by affecting mainly the availability of the DNA for transcription by chemical modifications of the DNA base pairs without directly altering the DNA sequence, by affecting the architecture of the chromatin, and by the activity of non-coding RNAs. The DNA undergoes modifications such as methylation, whereas histones undergo modifications such as acetylation, phosphorylation, SUMOylation, ubiquitylation, etc. These modifications and other mechanisms govern the architecture of the chromatin. The architecture of chromatin determines which portion of the DNA can be expressed, and this depends on histones and non-histone chromatin-associated proteins such as the High mobility group (HMG) proteins [1]. Non-coding RNAs such as microRNAs (miRNAs), long non-coding RNAs, and small interfering RNAs have also been shown to affect epigenetic mechanisms [2, 3, 4]. In a genome, the collection of all the modifications that regulate gene expression is called its epigenome.

2. Factors affecting epigenetics

2.1 Histone modifications

2.1.1 Structure of chromatin

The negatively charged DNA, where the negative charge is due to the phosphate groups of its sugar-phosphate backbone, is electrostatically attracted to the positively charged lysine of the histone proteins. Two of each H2A, H2B, H3, and H4 histone proteins come together to form a histone octamer [5]. The DNA forms a complex with histone octamer to form a nucleosome. The nucleosome consists of about 146 base pairs of DNAs wrapped around the histone octamer in a superhelical fashion [6, 7]. Upon addition of H1 histone to the nucleosome, it forms a chromatosome, which consists of around 166 base pairs of DNAs wound around it. Two chromatosomes are connected by linker DNA [8]. The C-terminal domains of H2A and H2B, as well as the N-terminal domains of H2A, H2B, H3, and H4 extend from the globular nucleosome core and are called histone tails [9]. The region of chromatin where nucleosomes are densely packed is called heterochromatin. It is inaccessible to the transcription factors and polymerases and thus is a transcriptionally inactive region. However, the region of chromatin where nucleosomes are loosely packed is called euchromatin. The DNA in this region is accessible to the transcription factors and polymerases and thus it can be transcribed. Various modifications to the histone proteins allow the nucleosomes to be densely or loosely packed. These modifications shall be discussed below. There exist a variety of cross-talk among various modifications. This cross-talk is facilitated by “writers”, “readers”, and “erasers”. Writers are enzymes that add a modification to histones or DNA, similarly, erasers are enzymes that remove the modification. However, readers have a domain that recognizes and interprets the modified or unmodified site [10]. Histone modifications can be studied using chromatin immunoprecipitation assays (ChIP). In the presence of high-quality antibodies, ChIP assays can analyze even minute changes in histone modification and nucleosome structure [11]. Distribution and levels of endogenous histone H3 lysine modifications can be monitored using Fabs (fluorescently labeled specific antigen-binding fragments), without disturbing cell growth and embryo development [12].

2.1.2 Acetylation

Histone acetylation and deacetylation play a significant role in gene regulation. The N-terminal tail projecting from the histone core of the nucleosome contains positively charged lysine residues that undergo acetylation or deacetylation catalyzed by histone acetyltransferase (HAT) and histone deacetylase (HDAC), respectively. Acetylation removes the positive charges on histones, thus weakening the electrostatic attraction between histones and the phosphate-sugar backbone of the DNA, resulting in relaxed chromatin, which is associated with gene expression. Hence, histone acetylation is generally considered as an active histone marker. Generally, hyperacetylation leads to more relaxed chromatin whereas hypoacetylation leads to more condensed chromatin. Histone acetyltransferase CBP (cyclic-AMP response element-binding protein) acts in conjugation with p300, forming CBP/p300 complex, which is capable of recruiting other HATs, like PCAF (p300/CBP-associated factor) [13]. As many as 25 HATs have been identified so far and classified into five families—CBP/p300, SRC, MYST, TAFII250, and Gcn5-related N-acetyl-transferase. All HATs use acetyl-coenzyme A as an acetyl group donor [14]. Most active gene enhancers have been observed to show high levels of the H3K122ac mark (Table 1) [15].

Enzymes/Writers	Residues modified
HAT1	H4 (K5, K12)
CBP/p300	H3(K14, K18, K122) H4(K5,K8) H2A(K5) H2B(K12, K15)
PCAF/GCN5	H3 (K9, K14, K18)

Table 1.

Examples of HATs and their residues modified [15, 16, 17].

Humans show 18 HDACs, which are divided into four classes as shown in Table 2.

Class	Enzymes/Erasers	Properties
Class I (Rpd3-like proteins)	HDAC1 to 3; HDAC8	Catalyze zinc-dependent hydrolysis of acetylated histones
Class II (Hda1-like proteins)	HDAC4 to 7; HDAC9; HDAC10	Catalyze zinc-dependent hydrolysis of acetylated histones
Class III (Sir2-like proteins)	SIRT (sirtuins) 1 to 7	Utilize NAD^+ during deacetylation to form nicotinamide and 2’-O-acetyl-ADP-ribose (metabolite)
Class IV	HDAC 11	Catalyze zinc-dependent hydrolysis of acetylated histones

Table 2.

Various classes of HDACs and their general properties [13].

2.1.3 Application of acetylation

Inactivation of CBP, such as through chromosomal translocation or bi-allelic mutations, has been observed to be correlated with oncogenic effects, observed to be involved in leukemia [18]. However, inhibition of CBP/p300 has shown antitumorigenic properties in regard to gastric cancers [19]. TATA-box binding protein associated factor 9 (TAF9) increases fatty acid β-oxidation and reduces lipid droplet accumulation is reportedly deacetylated by HDAC1, which regulates the capacity of TAF9 to mediate fatty acid β-oxidation and lipid droplet accumulation in nonalcoholic fatty liver disease (NAFLD) [20].

2.1.4 Methylation

Methylation of lysine residues of histone proteins (usually H3 and H4) is catalyzed by lysine methyltransferases (KMTs) and reversed by lysine demethylases (KDMs). This modification occurs post-transcriptionally. KDMs and KMTs also have shown roles in the regulation of the cell cycle [21]. In plants, DNA methylation tends to occur as a heritable epigenetic mark at the C-5 position of cytosine in the context of CG, CHG, and CHH (where H is A, C, or T) to form 5-methylcytosine.

2.1.4.1 Lysine methyltransferase (KMT)

Lysine methyltransferases (KMT) transfer a methyl group from S-adenosyl-L-methionine (SAM) onto the epsilon amino group of lysine residues of histone proteins. There are two classes of KMTs based on their catalytic domains: the SET domain-containing enzymes, and the one lacking SET domain. The latter is represented by KMT4, which is also known as Dot1L in humans. Both enzyme classes use S-adenosyl-L-methionine (SAM) as the methyl group donor [21]. The lysine of histone can be monomethylated, dimethylated, or trimethylated. For instance, trimethylated lysine 9 of histone H3 is represented as H3K9me3 and its monomethylated form is represented as H3K9me1.

2.1.4.2 Lysine demethylase (KDM)

Lysine demethylases remove the methyl group from the methylated epsilon amino group of lysine residues of histone proteins. KDM1A (also known as LSD1)—the first demethylase to be discovered—contains a flavin adenine dinucleotide-dependent monoamine oxidase domain that has been known to catalyze the demethylation of H3K4me2 and H3K4me1. Another class of KDMs employs jumonji (jmj) C domain to catalyze demethylation by oxidizing methyl groups. The cofactors of JmjC proteins are alpha-ketoglutarate, molecular oxygen, and Fe (II) [22]. Formaldehyde is one of the products of demethylase reactions (Table 3) [26].

Nature of methylase	Enzyme(s)/Writer(s)	Histone residue(s)
Mono-demethylases	KDM1B	H3K4
Di-demethylases	KDM8	H3K36
Tri-demethylases	KDM5A	H3K4

Table 3.

Examples of demethylases with their target histone residues [23, 24, 25].

2.1.5 Phosphorylation

Histone phosphorylation is a posttranslational modification instigated by DNA damage, entry into mitosis, or extracellular signals. It can trigger the binding of reader proteins and change the affinity of reader or writer proteins of other histone modifications [27]. Serine (S), threonine (T), and tyrosine (Y) are the sites of phosphorylation on histones. The mammalian 14–3-3 family of readers of the H3S10ph mark is composed of seven members that have been demonstrated to show interaction with around 700 different factors [28], including many chromatin-modifying proteins and transcriptional regulators, for instance, p53 [29]. 14–3-3 show increased affinity for the H3S10ph mark when the nearby lysine residues K9 or K14 are acetylated [30]. H3S10 is phosphorylated during mitosis by the action of Aurora B kinase, where data has suggested that this phosphorylation may function by displacing HP1 (Heterochromatin protein 1) from H3K9me, which otherwise plays a role in the compaction of chromatin. H3T3 phosphorylation catalyzed by Haspin kinase is required for appropriate metaphase chromosome alignment [31] (Table 4).

Histone residue (phosphorylated)	Kinase(s)/Writers	Function(s) of the phosphorylation mark
H1T18ph,	CDK2	—
H2AS1ph	Ribosomal protein S6 kinase alpha-5	Transcription inhibition.
H2AT119ph	NHK-1, Aurora B	Mitotic regulation of chromatin structure and function.
H2BS32ph	Protein kinase C (PKC)	Probable role in apoptosis-related nucleosomal DNA fragmentation.
H2BS36ph	AMPK	Direct transcriptional and chromatin regulatory pathways resulting in cellular response to stress.
H3T3ph	Haspin	Proper localization of chromosomal passenger complex (CPC) at centromere.
H3T11ph	Death associated protein-like kinase (Dlk)	Regulation of kinetochore assembly (during prophase to early anaphase) [34].
H3T6ph	PKC beta 1	Hormone dependent gene activation: Phosphorylation-dependent on androgen prevents LSD1-mediated H3K4demethylation.
H3S10ph	Aurora B	Dissociates HP1 from chromatin and prevents formation of condensed heterochromatin. Assists in condensation during cell-division; involved in transcription of certain genes.
H3T41ph	JAK2	Involved in hematopoietic differentiation.
H3T45ph	Protein kinase -C, S-phase kinase Cdc7-Dbf4	DNA replication, apoptosis, function in DNA damaged cells when DNA is nicked.
H3Y41ph	Tyrosine-protein kinase JAK2
H4S1	CK II	Repair of DNA damage, chromatin assembly, and mitosis.
H4H18 & H4H75	Unknown	Destabilization of histone octamer to facilitate DNA replication.

Table 4.

Some mammalian histone sites of phosphorylation [32, 33].

2.1.6 SUMOylation

Chromatin structure and gene expression are also regulated by small ubiquitin-like modifier (SUMO) conjugation. Along with altering substrate-protein or substrate-DNA interactions, SUMO can also block ubiquitin attachment sites [35]. The reversible attachment of mature SUMO proteins to the lysine (K) side chains of substrate proteins are regulated by an enzyme pathway analogous to the ubiquitin pathway. SUMO is expressed in all eukaryotes and is evolutionarily conserved. Humans express five SUMO paralogs, SUMO-1, −2, −3, −4 and − 5. Saccharomyces cerevisiae expresses only one SUMO ortholog—Smt3, which is similar to human SUMO-1.H4K12 in humans undergoes SUMOylation, which results in the recruitment of HDAC1 and heterochromatin protein 1 (HP1) – affecting transcription. The C-terminal of SUMO undergoes activation by Aos1/Uba2 SUMO-activating enzyme (E1), post which, it is transferred to the Ubc9 SUMO-conjugating enzyme (E2). SUMO ligase (E3) often aids in ligating SUMO to one or more lysine residues of the substrate. This modification is reversible by the action of SUMO proteases [36]. Unlike the initial idea of SUMOylation exclusively affecting transcription negatively, recent studies have shown that histones of many active genes are SUMOylated. Therefore, SUMO conjugation can have either negative or positive effects on transcription [37, 38].

2.1.7 Ubiquitination

Ubiquitination is the reversible process of transfer of ubiquitin to the histone core proteins (H2A, H2B, H3, H4). It is also known as ubiquitylation. Histone ubiquitination is involved in nearly all DNA-related processes such as DNA replication, transcription, and repair. Ubiquitin moiety consists of the 76-amino acid polypeptide, and hence is a bulky modification. In humans, ubiquitination of histone mainly occurs on the H2AK119ub1 and H2BK120ub1 catalyzed by an isopeptide bond formation between the carboxy-terminal glycine of ubiquitin and the epsilon-group of a lysine residue on the carboxy-terminal tail of histones. Ubiquitin transfer is an ATP-dependent process. The first step is adenylation of the C terminus of ubiquitin catalyzed by E1. Two of the known human ubiquitin E1 enzymes are UBA1 and UBA6. It was observed that UBA1 associates with DNA break by interacting with poly-ADP ribosylated proteins [39]. UBA1 might be the preferred nuclear E1 [40]. E2 enzyme receives ubiquitin moiety from E1 enzyme and conjugates it to the respective substrate. It has been observed that in vitro, E2 is capable of E3-independent ubiquitination [41]. E3 ubiquitin ligase acts as a scaffold by positioning the E2-ub complex close to the target lysine. This target lysine is nucleophilic toward the C-terminus of ubiquitin, resulting in bond formation [40]. There are about 500–1000 E3 enzymes in humans [42]. The lysine can be both poly or monoubiquitinated. Polyubiquitylation is irreversible and a signal for proteasomal degradation, however, monoubiquitination results in a regulatory signal, which is reversible upon the action of deubiquitinating enzymes (DUBs), which are ubiquitin-specific proteases—USPs/UBPs. Although histone ubiquitination has largely been correlated with open chromatin and active genes, it can have an inactivating effect as well. Another instance of chromatin cross-talk can be observed in Ref. to histone ubiquitylation. H2A ubiquitylation mediated by PRC1 usually represses gene expression, on the other hand, H2B ubiquitylation can activate as well as repress gene expression. Ubiquitinated H2B is required for H3K4 methylation, however, H2A blocks it and thereby results in chromatin compaction.

2.2 DNA methylation

DNA methylation includes the addition of a methyl group to the DNA at the 5′ position of the pyrimidine ring of cytosine residues. This results in 5-methylcytosine (5mC). DNA methylation usually takes place on CpG dinucleotide sequence. The region of the genome where CpG residues are concentrated is known as a CpG island. CpG islands are located on more than half of human gene promoters. Most CpG dinucleotides are methylated [43] whereas most CpG islands are unmethylated, especially those located in the promoter region of transcriptionally active genes. These CpG islands, upon undergoing methylation can lead to gene silencing through various mechanisms such as inhibiting or promoting the recruitment of regulatory elements to their respective binding sites. Cancer cells usually show hypermethylated CpG islands causing the silencing of tumor suppressor genes. The role of 5-mC does not merely depend on its abundance but also on its genetic context or surroundings, and its location within the different regions of a gene. Non-CpG methylation can be found in a context where CHH or a CHG are present (H being T, A, or C), which is found in plants and embryonic stem cells. Other DNA methylations such as N⁶-methyladenine is being studied as potential epigenetic mark [44]. 5mC is converted to 5hmC (5-hydroxymethyl cytosine). This has been observed to be catalyzed by ten-eleven translocation family proteins [45]. DNA is methylated by the action of DNA methyltransferases (DNMTs), of which DNMT 1 is ubiquitously expressed. It uses S-Adenosyl-L-methionine as a methyl group donor. Cytosine methylation patterns are inherited through cell division. This involves DNMT 1 having hemimethylated CpG dinucleotide specificity. Hence, based on the presence of methylation on the CpG dinucleotide in the complementary template strand, DNMT 1 can methylate CpGs in the newly synthesized DNA strand [43]. Studying DNA methylation is centered on three major approaches: (i) bisulfite conversion-based, (ii) methylation-sensitive-enzyme-restriction based (MSRE), and (iii) affinity enrichment based. The methylation signal generated by these assays is then analyzed by either DNA hybridization or sequencing. Bisulfite converted DNA is most commonly analyzed by microarray or Next Generation Sequencing [46]. Various techniques are employed for DNA methylation profiling such as pyrosequencing, bisulfite-PCR, ChIP seq (Chromatin Immunoprecipitation), bisulfite seq, and specialized RNA seq. Illumina sequencing of total genomic DNA known as whole-genome bisulfite sequencing (WGBS), is a high-throughput for DNA methylation analysis [47]. Since bisulfite sequencing results in the alteration of unmethylated cytosine into uracil, which upon PCR amplification is replicated as adenine, bisulfite-free approaches have gained traction attributing to their noninterference with the DNA sequence. Several bisulfite-free methods for the detection of methylation have been developed recently, such as TAPS (TET-assisted pyridine borane sequencing) [48] and cfNOMe (cell-free DNA-based Nucleosome Occupancy and Methylation profiling) [49]. Some key factors to be considered when choosing a method for DNA methylation analysis have been comprehensively reviewed in [50]. Additionally, in a recent study, it has been shown that minor experimental variations can significantly impact epigenome outcome measures and data interpretation [51].

DNA methylation is capable of altering chromatin structure and by extension gene expression. Histone modifications, transcription factors, ncRNAs, etc. in concert with DNA methylation affect chromatin and regulate gene expression [52].

3. Effect of epigenetics on health

Although plants and mammals have significant morphological dissimilarities and a long evolutionary history, the similarities on a fundamental level are striking. Epigenetic mechanisms discovered in mammals or plants are mostly relevant to both [53]. Nutrition and environment play a crucial role in the development of phenotypic characters, from prenatal development to later on in life. The most widely studied effect of epigenetics on health is in terms of cancer biomarkers that are studied in the form of DNA methylation. However, epigenetics has a broader impact on health. Epigenetics also play a major role in plant growth, development, and reproduction, especially in plant breeding. Epigenetics of human health has gained traction in complex disorders such as allergies, autoimmune diseases, memory, cancer, behavior plasticity, and psychological and neurodegenerative disorders. Some epigenetic marks can be reversible, and this has funneled researchers’ interest in epigenetic therapy. Epidrugs are drugs that target epigenetic marks responsible for epigenetic alterations. An example of these is histone deacetylase inhibitors [54]. Histone deacetylase inhibitors are being used as cancer therapeutic agents, all while some have received U.S. F.D.A. approval for treatment of multiple myeloma, cutaneous and peripheral T-cell lymphoma. Additionally, HDAC inhibitors are being used as antifibrotic, anti-inflammatory, and antidiabetic agents.

3.1 Effects of epigenetics on human health

Epigenetics play a significant role in various diseases such as cancers, autoimmune diseases, neurodegenerative diseases, congenital diseases, etc. HATs and HDACs modulate the transcriptional activity of nuclear factor-κB that results in downstream inflammatory gene expression levels that have been identified in the regulation of several diabetic key genes [55]. Cancer cells usually show hypermethylated CpG islands preceding promoters, and this leads to the silencing of tumor suppressor genes. This silencing allows cells to grow rapidly, leading to tumorigenesis. Imprinting, in genetics, delineates a condition where one of the two alleles for a gene pair is not expressed due to certain epigenetic modifications. This can lead to complications if the expressed allele is impaired, causing phenotypes such as susceptibility to certain microbes or chemical substances. Compared to healthy cells, malignant cells show decreased monoacetylated (H4ac) and trimethylated form of H4 (H4me3) [56]. DNA methylation patterns show a change in response to inherited genetic polymorphisms, exposures to environmental chemicals, and diet [57, 58, 59]. Histone acetylase inhibitors are a class of epidrugs. An epidrug Panobinostat, a non-selective histone deacetylase inhibitor, has been approved by the U.S. F.D.A. for the treatment of multiple myeloma [60].

Nutrition, being one of the most studied factors, has been understood to play an important role in epigenetics. Adverse antenatal nutritive conditions and postnatal health all have been observed to be correlated. Nutrients can either act directly by inhibiting epigenetic enzymes such as DNMT, HDAC, or by altering the substrate availability necessary for those enzymatic functions. Low dietary levels of folate, methionine, or selenium (all involved in methyl group donation or transfer) can lead to hypomethylation, which has been observed in neural tube defects, atherosclerosis, and cancer [61, 62, 63, 64, 65]. It has been observed that prenatal as well as early postnatal stress exposure have impacts on disease susceptibility [66]. DNA hypomethylation and histone acetylation are involved in the induction of gamma-globin expression [67]. A clinical trial is underway that deals with the down-regulation of BCL11A gene, which suppresses the production of fetal hemoglobin (HbF), resulting in an increase in the level of HbF, which has been shown to be therapeutic in patients with beta-hemoglobinopathies [68].

Endocrine Disrupting Chemicals (EDC), man-made chemicals known to alter endocrine functioning, that has been correlated with lower birth weight in children induce Adipogenesis. The epigenome is susceptible to the generation of new phenotypes in response to changes in environmental stimuli (Tables 5 and 6).

Year	Name of the scientist(s)	Conclusions drawn/discoveries made
1996	Korenke et al.	Studied monozygotic identical twin for x-linked adrenoleukodystrophy (ALD) gene and concluded that some non-genetic factors might be responsible for the difference in ALD phenotype. [69]
2005	Fraga, M. et al.	Epigenetic variations arise during the lifetime of monozygotic twins. [70]

Table 5.

A few twin studies that led to the foundation of twin studies in epigenetics [69, 70].

Genes/diseases/disorders	Epigenetic observation	Note
Diabetes	HATs & HDACs modulate transcriptional activity of nuclear factor-Κb.	Show downstream inflammatory gene expression levels identified in the regulation of diabetic key genes.
Cancer	Hypermethylated CpG islands preceding promoters.	Leads to the silencing of tumor-suppressing genes and hence tumorigenesis.
Cancer	Decreased acetylation at H4ac and decreased methylation at H4me3.	Seen in malignant cells. [56]
Neural tube defects, atherosclerosis, cancer	Hypomethylation	Caused due to Low dietary levels of folate, methionine, or selenium (all involved in methyl group donation or transfer) [61, 62, 63, 64, 65]
Immunity	Alterations in levels of acetylation and methylation.	Required to alter DNA accessibility to allow recombination for antigen specific responses. [71]
Endocrine Disrupting Chemicals (EDC) that have been correlated with lower birth weight in children induce Adipogenesis	DNA methylation variance was also observed along with adipogenesis in human. Mesenchymal stem cells [72] exposed to EDC.

Table 6.

Summary of effects of epigenetics on human health from the text [56, 61, 62, 63, 64, 65, 71].

3.2 Effects of epigenetics on plant health

Epigenetic change of plant genomes resembles that of mammals in that there is an analogous profile of histone marks and the DNA can be methylated at cytosine residues. Still, plant epigenomes are more susceptible to environmental influence than those in animals. Transgenerational epigenetic inheritance has a requirement that the epigenetic marks can be passed to the progeny. The variation in methylation of the same gene among different plants is known as epialleles [73]. Stable and heritable stress-induced modifications that cannot be reversed are being referred to as the epigenetic “stress memory”. Epigenetic marks that are heritable may affect the inheritable phenotypic variation of plants, impacting fitness, and hence are subject to natural selection. However, unlike inheritable inheritance, the epigenetic changes show unstableness and are affected by the climate [74, 75]. DNA hypomethylation induced by pathogen infections acts as a part of plant defense response in many species including the model plant Arabidopsis thaliana (Tables 7 and 8) [76].

Epigenetic observation	Note
DNA hypomethylation induced by pathogens infections	Part of plant defense response.
Hypermethylated genome regions in Arabidopsis accession Columbia-0	Tend to preferentially occur in shoots than in roots. [77]

Table 7.

Few epigenetic observations and their role in plant health [77].

Scientists, Year	Observed effect	Probable Cause
Sano et al., 1990	Induction of dwarf plants in rice	Demethylation of rice genomic DNA
Burn et al., 1993	Induction of flowering initiation	Vernalization treatments cause a reduction of DNA methylation levels.

Table 8.

Few observations having underlying epigenetic mechanisms [78, 79].

4. Conclusion

Epigenetic mechanisms play a crucial role in the phenotype of an organism. Epigenetic mechanisms include DNA modifications such as methylation, histone modifications such as SUMOylation, methylation, acetylation, phosphorylation, etc.—and action of non-coding RNAs. Recent technological advancements have made and will progressively make studying such modifications easier, more accurate, and cost-effective. Studying epigenetic modifications has provided insights into the inter-individual differences that genetics alone could not account for. Many phenotypes and diseases in humans and plants show underlying epigenetic marks at play from early on in the life of the organism, and some conditions or diseases can even manifest later on in life depending on their nutrition and environment. Histone modification reactivates gamma-globin gene expression in adults. Down-regulation of gamma-globin suppressing genes, which suppresses the production of fetal hemoglobin (HbF), results in an increase in the level of HbF, which has been shown to be therapeutic in patients with beta-hemoglobinopathies. Histone deacetylases are being used to treat various diseases such as multiple myeloma, cutaneous and peripheral T-cell myeloma. Epigenetics can be used for selective breeding of crops with desirable traits. As more would be understood about the various regulatory pathways involved in epigenetic mechanisms and more epigenetic modifications, it could revolutionize human disease prevention.

Acknowledgments

The authors would like to thank Dr. B.A. Mehere, Principal, and Dr. Utpal Dongre, Head of the Department of Biochemistry and Biotechnology, Dr. Ambedkar College, Deekshabhoomi, Nagpur, India, for providing research space and facility.

Conflict of interest

The authors declare no conflict of interest.

Fundings

No fund was received for this work from any funding agencies.

Abbreviations

CBP	Cyclic-AMP response element-binding protein
CDK2	Cyclin-dependent kinase 2
cfNOMe	Cell-free DNA-based Nucleosome Occupancy and Methylation profiling
ChIP seq	Chromatin Immunoprecipitation
CPC	Chromosomal passenger complex
Dlk	Death associated protein-like kinase
DNMTs	DNA methyltransferases
DUBs	Deubiquitinating enzymes
HAT	Histone acetyltransferase
HbF	Fetal hemoglobin
HDAC	Histone deacetylase
HMG	High mobility group
HP1	Heterochromatin protein 1
jmj C	Jumonji
KDMs	Lysine demethylases
KMTs	Lysine methyltransferases
miRNAs	microRNAs
MSRE	Methylation-sensitive-enzyme-restriction based
NAFLD	Non-alcoholic fatty liver disease
ncRNAs	Non-coding RNAs
PCAF	p300/CBP-associated factor
PKC	Protein kinase C
SUMO	Small ubiquitin-like modifier
TAF9	TATA-box binding protein associated factor 9
TAPS	TET-assisted pyridine borane sequencing
USPs/UBPs	Ubiquitin specific proteases
WGBS	Whole-genome bisulfite sequencing