Epigenetics, DNA Methylation, Imprinted Genes Lecture Flashcards
Histone Modifications
Histone tails are sites of covalent modification
Acetylation
Acetylation of histone tails promotes loose chromatin structure that promotes transcription
Acetylation of the Lysines at the N terminus of histones removes positive charges, thereby reducing the affinity between histones and DNA.
This makes RNA polymerase and transcription factors easier to access the promoter region.
Histone acetylation enhances transcription while histone deacetylation represses transcription.
Histone modifications specify protein interactions
- Modified residues are “landing platforms” for transcriptional effector proteins.
- Covalent histone modifications determine the formation of euchromatin and heterochromatin.
Other types of Histone Modifications:
Lysine: Acetylation, SUMOsylation, Ubiquitination, Biotinylation
Arginine: Methylation, Citrullination, ADP-ribosylation
Glutamic Acid: ADP-ribosylation
Serine: Phosphorylation
Threonine: Phosphorylation
Proline: Cis-Trans Isomerization
DNA Methylation
Heterochromatin has more DNA methylation
Methylation leads to reduced transcription
Addition of CH3 (Methyl) group
4% of all Cytosines are methylated
70-80% CpGs methylated
CpG Islands
Majority unmethylated
- Regions (500 – 1 Kb) of higher G+C than genome average.
- Relatively devoid of methylation.
- ~ 60% mammalian RNA Pol II promoters found in CpG islands.
Epigenetics
Role of DNA Methylation, Histone Modification, Non-coding RNAs and Chromatin Remodeling in X-inactivation, Genomic Imprinting and Cancer
Study of heritable changes in genome function that occur without alterations to the DNA sequence.
They are difficult to reverse or irreversible.
They are inherited mitotically (somatic cells) or meiotically (transgenerationally).
Major Mechanisms of Epigenetic Change
Four important epigenetic factors that play clear roles in transcriptional regulation are known
Major Mechanisms of Epigenetic Change:
- DNA methylation
–A subset of cytosine (C) residues can be modified by methylation.
- Histone modifications
–Histones can be modified by acetylation & other types.
–Histone replacement/substitution
- Non-coding RNAs
–microRNAs (miRNAs).
–long non-coding RNAs (lnc RNAs).
- Chromatin remodeling
–Heterochromatin vs. Euchromatin
Euchromatin vs. Heterochromatin
Euchromatin: less compact; actively transcribed.
High acetylation, low methylation.
Histone 3 Lysine 4 methylation
Heterochromatin: more compact; transcriptionally inactive.
Low Acetylation, Dense methylation
Histone 3 Lysine 4 methylation
Heterochromatin can be either constitutive or facultative.
Constitutive: Regions such as centromeres- no gene expression, involved in chromatid separation.
Facultative: Developmental genes- can be opened or closed at specific times.
Mechanism of X-Inactivation
Creates Barr Bodies
Mechanism:
- Xist RNA Binding:
- Initiated at single site, the X inactivation center (XIC).
- At XIC is a gene for Xist RNA (X inactive specific transcript).
- Transcript is not translated (no open reading frame) – non-coding RNA.
- Expressed only from X chromosome to be inactivated.
- Binds in cis and then spreads. Spreads to other areas on same X chromosome.
- Binds independent of DNA sequence.
- Silencing pattern is global; it affects almost all genes on the X chromosome.
- Once established, the pattern of inactivation is maintained through mitosis.
- Expression of XIST on the active X chromosome is blocked by an antisense RNA called TSIX.
- Chromatin Alterations: Heterochromatin (proteins) and DNA Modifications
- Histones are under-acetlyated
- DNA is heavily methylated on cytosines of CpG sequences
ICE = Inactivation Element. When ICE is methylated, does not bind CTCF protein (usually insulates-blocks transcription).
Genomic Imprinting
An epigenetic modification that results in the unequal expression of the maternal and paternal alleles of a gene
A mechanism of epigenetic gene regulation through which the activity of a gene is reversibly modified depending on the sex of the parent that transmits it.
At least 131 genes in humans.
Characteristics:
- Located in clusters
- Noncoding RNAs and antisense transcripts
- Differential methylation
- Histone modifications
- Heritable
Challenges two assumptions of Mendelian genetics:
Maternal allele = paternal allele.
Two working copies are associated with normal function
Imprinting and cancer: Beckwith-Wiedemann syndrome
Beckwith-Wiedemann syndrome
- Chromosome 11
- Fetal overgrowth disorder:
Gigantism
Macroglossia
Viceromegaly
embryonic Wilms tumors
Condition results from an excess of paternal or loss of maternal contribution of genes or both.
IGF2 = Promotes embryonic growth (paternal)
CDKN1C: H19 = Tumor suppressor RNA (maternal)
Chromatin Remodeling
Chromatin Remodeling
Positioning of histones along DNA mediated by ATP-dependent nucleosome - remodeling complexes
Use the energy of ATP hydrolysis to:
Noncovalently reposition histone octamers or
Generate nucleosome free or dense chromatin.
Writers: Establish a mark on DNA or histone tail, such as methylation or acetlyation
Editors: Recognizes mark on DNA or histone and further modifies or removes
Readers: Mediates interaction of mark and protein complex to effect transcription
Chromatin remodeling can lead to cancers when an oncogene is acetylated or a tumor suppressor is methylated.
Epigenomics
Study of the effects of chromatin structure, including the higher order of chromatin folding and attachment to the nuclear matrix, packaging of DNA around nucleosomes, covalent modifications of histone tails (acetylation, methylation, phosphorylation, ubiquitination), and DNA methylation.
Importance of Epigenetics
Importance of Epigenetics:
- DNA methylation
- Non-coding RNAs & heterochromatin
- Histone modification
- X-chromosome inactivation
- Imprinting
- Development/reprogramming of somatic nucleus
- Cancer
* Developmental plasticity:
Environmental exposure produces a broad range of adult phenotypes from a single genotype by epigenetically altering gene expression.
Prenatal and postnatal environmental factors, nutritional supplements, xenobiotic chemicals, behavioral cues, reproductive factors and low-dose radiation can result in altered epigenetic programming and subsequent changes in the risk of developing disease.
- Fetal basis or developmental origins of adult-onset of disease
X-Chromosome inactivation
Imprinting
- Potential Targets for New Therapies:
Stem cells – developmental reprogramming of somatic nucleus
Cancer
Non-Coding RNAs
miRNAs
lncRNAs
MicroRNAs: Small non-coding RNAs (less than 200bp in length)
Long/Large intergenic non-coding RNAs (over 200bp in length)
microRNAs
Small non-coding RNAs (less than 200bp in length), High quality supporting data, Specific role in carcinogenesis.
Aberration in Cancer: Amplification, Deletion, Methylation, Gene Expression
Mechanism of Gene Regulation:
MicroRNAs (miRNAs) regulate gene expression through multiple pathways.
MiRNA finds target in messenger RNA
1. Endonucleolytic Cleavage: Perfect pairing between an miRNA and its target site induces endonucleolytic cleavage and the leads to degradation
2. Deadenylation and degradation: Loss of Poly-A tail leads to degradation
3. Inhibition of translation initiation: Block ribosomal entry
4. Inhibition after translation initiation: Translational repression such as promoting ribosome drop-off or stimulating proteolysis of the nascent peptide.
5. Stimulation of translation: miRNAs have also been shown to upregulate target expression under certain conditions.
long/large non-coding RNAs
Long/Large intergenic non-coding RNAs (over 200bp in length):
High quality of supporting data, specific role in carcinogenesis.
Aberration in cancer: Gene expression, Translocation
Mechanisms of Gene Regulation:
- Flexible scaffold for chromatin modifying complexes.
- Enhancer RNAs
- Tumor Suppressor Signaling
- RNA Processing: inhibiting or changing splicing pattern of mRNA
- RNA-RNA interactions: direct lncRNA-mRNA interactions can lead to cleavage
- MiRNA Sequestration
