Stem cells constitute the source of cells that replenishes the worn out or damaged cells in our tissue and enable the tissue to carry out the destined function. Tissue-specific stem cells are compartmentalized in a niche, which keeps the stem cells under quiescent condition. Thus, understanding the molecular events driving the successful differentiation of stem cells into several lineages is essential for its better manipulation of human applications. Given the developmental aspects of the cell, the cellular function is greatly dependent on the epigenomics signature that in turn governs the expression profile of the cell. The stable inheritance of the epigenome is crucial for the development, modulation, and maintenance of the cell and its complex tissue-specific function. Emerging evidence suggesting that stem cell chromatin comprises a specialized state in which self-renewing genes and its downstream lineage-specific genes are kept paralleled poised for activation. Thus, the epigenetic regulatory network and pathway dictate lineage commitment and differentiation. It mainly modifies the chromatin landscape to facilitate euchromatin and heterochromatin architecture, which in turn alters the accessibility of transcription factors to the gene loci. DNA methylation and histone marks are the two widely studied epigenetic modifications regulating the transcriptome profile of a specific lineage. Abnormalities in the epigenetic landscape lead to diseases or disorders. Here, we emphasize the prominence of the epigenetic network and its regulation in normal tissue functioning and in the diseased state. Furthermore, we highlighted the emerging role of epigenetic modifiers in lineage differentiation and epigenetic markers as novel druggable targets for cancer therapy.
The concept of genomic equivalence states that all cells in an organism carry the same genetic material, but the expression profiles may vary according to its destined function. Further, in order to facilitate the differential expression profile, epigenetics plays a central role in modulating the sequential changes in the chromatin landscape that finally leads to the specific chromatin signature for a particular lineage. Epigenetics is classically defined as “the branch of biology which deals with the cross-talk between genes and its products that finally leads to the phenotype into existence” (
Chromatin is a multimeric complex made up of domains of histone proteins on which the DNA is tightly wound and well packaged within the cell. An epigenome is a complimentary term connected with the chemical changes occurring in the cytosine moiety of DNA base sequence in the chromatin without altering the sequence of the DNA. This change can be often transferred down to the progeny
DNA packaging and chromatin assembly are tightly organized phenomena that determine the transcriptome profile of cell type. Thus, the cellular identity and its homeostasis principally depend on the epigenetic modifications that widely occur throughout the genome. The conventional chromatin modifications that occur in the mammalian genome are methylation of DNA and modifications in the core histone residues. In DNA methylation, the methyl groups donated from S-adenosyl methionine (SAM) are covalently added cytosine or adenine residues of DNA by the DNMTase family of enzymes. Methyl group is added to the 5th position of cytosine residue in CpG dinucleotide is the most studied and highly heritable modification suggested to maintain the stable genomic integrity (
Histones are proteins made up of basic amino acids and involved in the formation of nucleosome core as octamer complex containing a C-terminal globular domain and N-terminal tail (
Presence of heavily acetylated histone residues (H3K9ac, H4K12ac (
The chromatin remodelers are ATP-dependent multi-enzyme complexes involving in the process of chromatin opening using an ATP dependent manner. According to the configuration and order of the ATPase subunits, the nucleosome remodelers are classified into four classes, such as the SWI/SNF family, the ISWI family, the CHD family and the INO80 family (
In the central dogma of molecular biology, the information flow from DNA to protein contributes through RNA, which functions to code for a protein with a destined specific function. However, a few exceptions to this paradigm are still present in which RNAs do not code for proteins, which in turn function in the regulation and processing of other RNAs (mRNAs, tRNAs, and rRNAs). It includes in a process such as splicing (snRNAs), nucleotide modification (snoRNAs), processing pre-tRNAs (RNase P, a ribozyme). Other small ncRNAs like miRNAs and siRNAs that involve in gene regulation by targeting mRNAs. LncRNAs are > 200 nucleotides to 2 kb sequence of non-coding transcripts (
1. DNA modifications (like acetylation or methylation mostly on promoter sites for inhibiting active gene transcription), 2. Histone modifications (like acetylation or methylation at lysine residues for activating or inhibiting gene transcription), and 3. miRNA biogenesis.
DNA methylation occurs mainly in the CpG dinucleotide region, which is found in clusters called CpG islands. 60% of human gene promoter has CpG islands, which is unmethylated in stem cells and get methylated, leads to tissue-specific expression during early embryonic development or in different specific tissue types in adults. Stem cells unveil a unique gene expression profile that governs cell fate during lineage commitment and differentiation (
These genes were highly methylated during lymphoid differentiation demonstrating DNA methylation may not only interfere with gene expression but also block the binding of unwanted myeloid transcription factors, which are of oncogenic nature. DNA methylation parallelly inhibits the activation of TF gene loci, as well as block the binding of TFs across the whole genome to make sure the two-tier epigenetic barrier in regulating the expression of myeloid TFs in lymphoid cells. Thus, a high-resolution global DNA Methylation mapping proved the gain of DNA methylation and its direct correlation in loss of gene expression during the course of differentiation (
The differentiation potential of MSCs to adipocytes was directly correlated with its long-term culture and demonstrated the methylated status of
The promoter methylation status of NKX 2.5 and sFRP4 in umbilical cord MSCs displayed that both promoters underwent demethylation and further validated with upregulated expression at mRNA level upon cardiac stimulation (
A specific mark on histone protein contributes to its post-translational modifications governs gene expression patterns and differentiation potential in stem cells (
PcG protein complex maintains the gene expression of many cells during development. PcG proteins like EZH (EZH1/EZH2), EED and SUZ 12 along with methyltransferase, PRC2 acts on histone H3 lysine 27 (H3K27), are very essential for maintenance and control of pluripotency. PRC2, along with jumonji protein, acts as a master regulatory switch by which this protein complex rapidly reprograms the epigenome either by repression or subsequent activation
The master regulatory pluripotency triads such as SOX2, OCT4, and NANOG determine the stemness and differentiation potential of embryonic stem cells (
Histone lysine demethylase (KDM2A) was shown to regulate MSCs proliferation and osteo-/dentinogenic differentiation. Knock-in/Knock out studies on KDM2A has enhanced the SCAP differentiation potential into the adipogenic and chondrogenic lineage. Also demonstrated knockdown of KDM2A, showed cofactor BCOR has considerably increased expression of Sox2 and Nanog by depositing H3K4Me3 marks in the Sox2 and Nanog loci (
Comprehensive epigenomic profiling between CD44+ and CD24+ in breast epithelial cells showed the crosstalk between K27 and DNA methylation correlated with higher expression of CD24+ genes independent of the K27 mark. But CD44+ cells showed CD44 high dependence of K27 marks. Thus, suggesting a presumed strategy independent of gene body methylation and gene expression and further correlated with an increase of promoter K27 marks (
WGBS of large partially methylated domains study showed signature similar to DPSCs as compared to 30–40% with ICM. DPSCs showed a similar methylation profile with neuronal stem cell lines and placenta-derived cells, as demonstrated by principal component analysis (
The ChIP-on-chip assay revealed that promoters of RUNX, MSX2, and DLK5, early mineralization genes provided with H3K4Me3 active marks whereas repressive marks H3K9Me3 or H3K27Me3 augmented in OSX, IBSP, and BGLAP gene promoters. It also mediated the suppression of dental family genes (DSPP and DMP1 genes) in dental follicular (DF) cells and not in dental pulp (DP) cells. (
Precise chromatin configuration leads to appropriate gene expression which ensures proper stem cell and progenitor differentiation, lineage commitment. miRNAs are demonstrated to act mainly in RNA silencing and post-transcriptional via base-pairing with complementary sequences within mRNA molecules, which triggers the degradation of mRNA strand (
miRNA studied | Target cells | Interpretation | Reference |
---|---|---|---|
miR122 | hADSCs | overexpression triggers hepatogenesis | ( |
Let 7f | hADSCs | negatively regulates hepatogenesis | ( |
miR137 | hADSCs | induce adipogenesis via targeting CDC42 | ( |
miR103a-3p | hADSCs | induce osteogenesis by CDK6 and DICER pathway | ( |
miR26a | hADSCs | induce osteogenesis by targeting SMAD1 TF | ( |
miR196 | hADSCs | induces osteogenesis via targeting HOXC8 | ( |
miR125b, miR26a | MSCs | inhibits osteogenesis | ( |
miR21 |
mES cells | targets Sox2 and declines pluripotency | ( |
miR133 |
Mesodermal progenitor cells | interact with serum response element and enhance myogenesis |
( |
miR223 |
Common myeloid or lymphoid progenitor cells | induces B-lymphocyte lineage | ( |
miR221, miR222 | HSCs | blocks erythropoiesis by targeting c-Kit | ( |
miR105, miR155, miR221, miR222 |
HSCs | downregulated during erythropoiesis |
( |
miR144, miR451 | ES cells | requires GATA1 for inducing erythropoiesis | ( |
miR1 | hES cells | promote mesoderm formation by repressing notch ligand DLL-1 induces cardiac mesoderm formation | ( |
miR181 | Hepatocellular carcinoma | reduction of EpCAM+ CSCs and tumor initiating potential | ( |
miR200c | Breast cancer cells | targets BMI1 and inhibits the expansion of embryonal carcinoma cells | ( |
miRNA188-5p | Bone marrow derived cells | targets MMP1/13 and mediates matrix degeneration of chondro neovascularization development | ( |
miR144-5p | Non-small cell lung carcinoma cells | enhanced radiosensitive by targeting ATF2 | ( |
miR140-5p | 3T3-L1 | induces adipogenesis by targeting TGF-β | ( |
miR135a-5p | 3T3-L1 | inhibits adipogenesis via canonical Wnt/Beta catenin pathway | ( |
miR24 | C2C12 myoblast cell line | targets TGF-β and inhibits myogenesis | ( |
miR124, miR128 | Neuron stem cells | promotes neuronal differentiation and suppresses astrocyte differentiation | ( |
miR203 | Skin stem cells | promotes skin cells differentiation by inducing cell cycle exit | ( |
Let-7 | Breast CSCs | suppress CSCs self-renewal | ( |
miR451, 486, 425, 16, 103, 107, 185 | Glioblastoma CD133+ve population | declines the CSCs number, and inhibits neurospheres formation | ( |
MiRNA-MicroRNA, hADSCs-Human Adipose Derived Stem Cells, MSCs-Mesenchymal Stem Cells, mES-Mouse Embryonic Stem Cells, HSCs-Hematopoietic Stem Cells, hES-Human Embryonic Stem Cells, CSCs-Cancer Stem Cells, CD-Cluster of Differentiation, 3T3-L1-Mouse embryo fibroblast cell line, CDC42-Cell Division Control protein 42 homolog, CDK6-Cyclin Dependent Kinase-6, TF-Transcription Factor, MEF2C-Myocyte-specific Enhancer Factor-2C, GATA1-GATA binding factor-1, DLL1-Delta Like canonical Notch Ligand-1, TGFβ-Transforming Growth Factor-β, EpCAM-Epithelial Cell Adhesion Molecule, BMI1-B cell-specific Moloney murine leukemia virus Integration site-1, ATF2-Cyclic AMP-dependent transcription factor-2.
Each mammalian cell differs from the other in the differentiated state but still retains a similar genome, which was inherited from the common precursor ESC. These cells have the potential to de-differentiate and acquire their totipotent character in a specific milieu. However, this process is determined by the expurgation of diverse epigenetic states in the chromatin through various covalent modifications in DNA and histone leads to change of fate by reprogramming. The initiation of CSCs also involves the parallel route during cancer triggering might be hypothesized based on epigenetic reprogramming in which downregulation of differentiation-specific genes and upregulation of stemness property, thereby eventually escapes the natural cell death process. The major cellular event that drives the carcinogenesis is reprogramming of the epigenome initiated by a series of cellular signaling cascades, finally culminating in gaining and maintenance of stem cell properties (
The CSCs are a subpopulation of cells present in the tumor niche, which undergo changes in Methylome and chromatin signature that finally transform into CSCs. During the initial phase of cancer initiation, the epigenetic modifiers might facilitate opening up target oncogenic DNA sites by requisite over-expression of oncogenic factors.
Through epigenetic analysis, various druggable targets are identified and targeted. Some of which are currently in clinical trials. In
Pictorial illustration showing the panel of epigenetic markers involving in regulating the homeostasis in ESCs, any aberrant imbalance in the expression of the epigenetic markers is often correlated with cancer progression and prognosis of the cancer treatment.
The expression of repressed tumor-promoting factors and silencing tumor-suppressing genes were correlated with the downregulation of DNMT enzyme (
Wnt pathway was epigenetically regulated by Brahma-related gene 1 (BRG1), which is the main tumor-initiating factor in triggering intestinal cancer, and its downregulation prevents adenoma development and decreased TIC population. Also, BRG is involved in leukemia maintenance, as BRG-AML cells are more sensitive to treatments than the BRG+ cells (
BMI-1 maintains the stemness of CSCs and also intricate in triggering various types of cancers. Overexpression of BMI-1 induced stemness property in CSCs which in turn augmented tumor initiation (
Efficient silencing of appropriate chromatin remodeling complexes in differentiated cell types induces pluripotency. Any modifications in the transcriptome profiling of chromatin remodelers are proficient in initiating tumor genesis. Hence, regulating chromatin complexes modifies the capability to induce CSCs phenotype. Ectopic expression of these complexes could cause repression of tumor suppressors or expression of oncogenic promoter genes. EZH2 expression was found higher in side-populations as observed in breast and pancreatic cancer lines than in non-CSC populations. Also, a recent report showed that knockout of EZH2 resulted in decreased CSCs incidence, which supplementary endorses EZH2 as a useful CSC marker and targeting protein for therapeutic purposes. Regulatory genes are often silent in ES cells (
A chip on-chip studies showed the overlapping of more H3K27 trimethylated (repressed state) regions with few such specific modifications where both active and inactive marks co-exist in the particular gene promoter region is called “Chromatin Bivalency,” which is considered as a unique feature frequently found in the domains of developmental regulatory genes poised for induction. In human primary T cells, both H3K27 and H3K4 methylation co-exhibit in the HOXB7 promoter region, which is as compared analogy to the proposed role of bivalent chromatin in ES cells (
The well-known PcG group of proteins has two main histone modifiers, PRC1 and PRC2, respectively. PRC1 complexes (BMI1, RING1A, RING1B, PHC) are capable of catalyzing mono-ubiquitination of lysine residues on Histone2A proteins. Any aberrations in PRC1, specifically RING1B, exhibited embryonic lethality in
High mobility nuclear proteins regulate the expression of many developmental genes
Active DNA demethylation is the prerequisite obligation for cells to recover self-renewal property and reverts to their pluripotent state. This can be achieved progressively by modification of 5-methyl cytosine to thymine or 5-hydroxy methylcytosine by Ten-Eleven Translocation proteins (TET) and activation-induced deamination (AID), respectively (
The characteristic property of pluripotency in ESCs is highly dependent on the PcG proteins, where it tends to maintain the balance between repressing markers and pluripotent specific markers by repressing the early differentiation marker genes and maintains the pluripotency genes. Initial days of fate commitments, deposition of PCR2 mediated histone H3K27Me3 makes these cells trigger the early differentiation marker genes, but still, PcG proteins suppress the late differentiation genes for a specific lineage. Consistent high expression of EZH2 in ESCs and early mouse development, which determines the pluripotent state upon declining its level differentiation is triggered. Also, EZH2 is abundantly expressed in progenitor cells of the epidermis region; nevertheless, its level declines upon commitment. Additionally, EZH2 was shown to maintain the multipotency in Mesoderm derived stem cells like myeloid and lymphoid progenitors, muscle progenitors, and neural progenitors.
EZH2 cover-expressed in the HSCs preserves the long-term self-renewing potential, which prevents HSCs depletion after serial transformation. Increased EZH2 expression blocked muscle differentiation from myoblasts due to histone lysine methyltransferase (HKMT) activity in its SET domain. NSCs also expressed high EZH2 level, and further commitment to astrocytes its level declined. Reduced differentiation potential into astrocytes in NSCs on ectopic expression of EZH2 further substantiated the role of EZH2 in preserving pluripotent or multipotent property of stem cells (
It is well demonstrated that complex coordinated networking between epigenetic mediators and chromatin landscapes facilitates the expression of Glial gene expression and favors glial fate determination in Neuronal Stem Cells (NSCs). Whole-genome bisulfite sequencing analysis data demonstrated that distinct epigenetic signatures leading to three different neuronal sub-populations in NSCs, such as self-renewing neurons stem cells, progenitors consistently expressing neuronal markers actively, and cells switched from neurogenic to gliogenic phase, respectively (
Molecular events associated with lineage decisions are the contemporary area of research in normal stem cells for its better manipulation in clinical use. A large number of evidences is available to depict chemical-based approaches, as a versatile tool in controlling the stem cell properties and their fate, such as stemness, lineage differentiation, reprogramming and regeneration. Our previous study about methylation profiling of cardiac-specific gene (CSG) promoter in human Wharton’s jelly derived MSCs at single-nucleotide resolution mapping (
CHIR 99021 is one such molecule that is an agonist of the Wnt pathway, widely used to sustain pluripotency in ESCs, induce reprogramming in somatic cells along with few Yamanaka factors, and lineage differentiation in MSCs (
From the above-cited references and reports, it is well apparent that epigenetics is either directly or indirectly involved in the lineage commitment, identity, differentiation potential of the cell. On the other hand, it is playing a predominant role in cancer initiation, tumor progression and dissemination. We found much of the work has been extensively done from 2010 to 2016, and this accelerated us to understand the contribution of the epigenetic regulatory network in the above-mentioned areas of research. Due to this, many avenues of cancer research and basic fundamental research in stem cell biology are started employing epigenetic modifiers in disease management and
A study by
Interestingly screening of small molecules having the potential to rescue MSCs senescence-related concerns under
The generation of iPSCs has revelutionized the avenues of stem cell biology, still the concept of reprogramming is not fully understood. Hitherto, the two well-known proposed models of reprogramming such as the elite and stochastic were widely accepted. However, it is still under debate about the mechanism of reprogramming. The elite model proposes that not all the cells were conducive for reprogramming, this often correlated with the reprogramming efficiencies of the source cells employed for reprogramming studies. Whereas the stochastic model proposes that every cell inherently has the potential to undergo the process of reprogramming and become iPSCs (
Epigenetics plays a significant regulatory role in determining stem cell linage and cellular differentiation. Its predominant function has also been recognized in recruiting the appropriate transcriptional machinery during embryonic development and adult tissue homeostasis. Specialized chromatin structure dictates the unique expression profiles of stem cells intact and regulates its differentiation into various downstream lineages. Numerous epigenetic modifications occur concomitantly during the differentiation of MSCs to respective cell types. However, the knowledge about the epigenetic regulatory mechanism in relation to differentiation towards specific cell lineage is limited. Further investigations of the epigenetic profiling may help us in better understanding the systematic derivation of the physiologically competent cell types for exploitation in the field of various other regenerative therapy pursuits.
The MSCs have been used in preclinical models for various bone and cartilage tissue engineering. The development of tissue-engineered products has given considerable promising use for rebuilding damaged or diseased tissues. Epigenetic regulatory mechanisms are likely to enhance scientific hold on transcriptional regulation, especially critical for stem cells, their potential for self-renewal and differentiation. Classification based on gene expression profiles could help us in segregating stem cells into pluripotent stem cells, multipotent stem cells, and multipotent adult stem cells. Analysis of histone modifications mediated by PcG proteins and promoter histone methylation of a gene have demonstrated that certain marks on the histone bodies are necessary for the self-renewing stem cell populations, and its subsequent loss could deliberately lead to differentiation of a specific lineage. DNA methylation may often correlate with the restricted differentiation potential towards the specific lineage.
Research on the chromatin signature and cellular behavior would be more useful in fishing out the long-term self-renewing potential cells for transplantation and regenerative therapy. Overall, a combination of DNA methylation at gene promoter region and histone core modification marks in gene promoter and gene body contributes to the epigenetic regulations in stem cell state and determines the degree of differentiation impending from pluripotent stem cells to multipotent stem cells and progenitors. Similarly, the epigenetic road map will give us a clear picture of normal and cancerous chromatin organization or architectural difference, which will contribute to identifying new potential druggable targets for cancer treatment regime in the future.
KG is thankful to UGC, PKG and BZE to MHRD for their fellowships. All authors are grateful to IIT-Madras, Kumaun University, Mahatma Gandhi Central University, and Department of Life Sciences, SBSR, Sharda University for infrastructure and facility.