Epigenetic Theory Flashcards
Cells vary in phenotype due to:
Unique genes (and thus protein) expression profiles. Different cells express/turn off different genes.
Phenotype vs genotype:
Phenotype is what it looks like/function and morphology
Genotype is the sequence of nucleotide
Gene expression is regulated by both (2):
Explain:
- Extrinsic and intrinsic cues
- Genes may be turned on or off or transcript levels can be increased/decreased
Extrinsic factors:
- Signals from the cell’s environment such as other cells/trophic/growth factors trigger intracellular signalling cascades: changes in transcription
Intrinsic factors:
- DNA modification
- Cell’s own machinery chemically modifies DNA in a way that affects gene expression
What is epigenetics?
Study of changes in gene expression (and thus phenotype) that occur due to chemical modifications of the genome rather than a change to the DNA sequence itself (not a mutation or change in gene sequence).
Explain the relationship between cell division and epigenetic modifications
Cells have the mechanisms to copy epigenetic modifications during divisions. If something causes modification, they can be passed on. These modifications are however, reversibles.
Way epigentic modifications affect offsprings (2):
- Mother experience something traumatic or positive in environment and that affects epigenome of offspring. (Epigenetic modification that affects her behaviour and put her in risky situations that also affect prenatal environment of offspring.
- Germline affected
Explain epigenetic example with human twins (3):
- There is a lack of concordance in pattern of disease despite both twins having 100% identical genome.
- 30-60% of concordance rate for a vast array of diseases such as schizophrenia, AD, MS, Crohn disease, asthma, diabetes, prostate cancer. There is only 10% concordance rate for breast cancer. These are fairly genetic disorders but there’s other things going on (epigenetic)
- When twin start to age they don’t look alike due to epigentic influence
Phenotypic variations can arise in abscence of:
changes to the nucleotide sequence of genes
Experiences and the environment can ——- gene expression
promote or inhibit
A lifetime of experiences can alter behaviour via:
the epigenome
What is gene expression?
Process by which cells convert the information encoded in our DNA into a functional product
Gene structure and expression
Genes are made up of —- and most genes code for ——
- DNA
- Protein products
Explain coding and noncoding regions:
Genes have coding and noncoding regions (introns) which need to be removed. Mature mRNA have introns removed and exons string together which gets translated to proteiin product.
Transcription is initiated at:
The promotor region which is upstream, more 5’ end of the gene.
Pre-mRNA has —–
introns
DNA consists of two polynucleotide chains: (2)
- Template strand (-) (aka antisense strand/noncoding strand): Read by RNA polymerase II to make mRNA
- Coding strand (+) (aka sense strand/coding strand): Sequence of mRNA ribonucleotide is identical to the nucleotide sequence. Not read by RNA poly but reported 5’-3’
RNA Polymerase have directionality in the —— direction. New nucleotides are added at the —– end.
- 3’-5’
- 3’
The template strand is —– and the coding strand is —– as the mRNA strand
- complementary
- the same
Control of transcription in eukaryocytes (3):
All cells + brain + neuron
- Each cell in the body contains the DNA for every gene but only express a subset of genes as RNAs
- The brain expresses more genes than any other organ
- Diverse populations of neurons have different gene expression profiles
—— regulatory regions, promotoers, enhancers and silences ensure that ——-
- Upstream
- The right gene is expressed in the right cells, at the right time
Unique complement of transcription factors interacts with:
- promoters and enhancer
Proximal regulatory region:
Promoter
Distal regulatory region:
Enhancers and silencers
Promoter consists of:
Core promoter region and a promoter proximal region
Core promoter vs Promotor proximal regions
Core promoter: closest to mRNA inititation site, ~30bp upstream
Promoter proximal region: Further upstream ~at least 100bp
Distal regulatory regions (3):
specificity + location + nessicity
- Specific to particular genes and particular tissues
- Even more upstream ~1000bp
- Can increase transcription but not required
Compared to enhancers and silencers, promotor regions are more:
invariable
Talk about the transcription initiation process in eukaryotes (3):
Regulatory regions are recognized and bound by protein complexes.
- Basal or General TF binds to TATA box. Absolutely necessary for transcription to occur.
- TF IIB binds and makes the pre-initiation complex. Recruits RNA Poly 2 + Basal TF that complex with RNA Poly 2.
- RNA Pol 2 binds to TFIIH, TFIIE, and TFIIF and is recruited to the pre-initiation complex. TFIIH is a kinase enzyme that phosphorylates RNA Pol2 to activate it.
How can special transcription factors attached to distal regulatory regions affect the core promotoer and it’s complex of basal TF? (3)
- Special TF called activators and repressors bind thousands of bp upstream of transcription start site. They can upregulate or downregulate to control expression of specific genes.
- Activators: Help basal TFs and/or RNA polymerase bind to core promoter.
- Repressors: May impede basal TFs or RNA polymerase such that they cannot bind to core promoter to initiate transcription.
CRE binding proteins (CREBs) (4)
What is it + mechanism of action (3)
- Special transcription factors
1. An extracellular signal arrives at the cell surface, activates the corresponding receptor, which leads to the production of a second messenger such as cAMP or Ca2+, which in turn activates a protein kinase. This protein kinase translocates to the cell nucleus, where it phosphorylates a CREB protein.
2. The activated CREB protein then binds to short lengths of palindromic DNA known as the CRE sequence.
3. It then brings a CBP (CREB-binding protein), which is an epigentic modifier that allows it to switch certain genes on or off.
Enhancers aid in
recruitment of the initiation complex (Basal TF and RNA ply 2)
Combinatorial regulation (2)
What + ex
- A mechanism where multiple transcription factors work together to regulate the expression of many genes, allowing for fine-tuned control of gene activity.
- Certain combinations of activators may need to be present (in the absence of certain repressors) to induce gene expression.
Different cell types express:
characteristic set of transcription factors
Epigenetic modifications also determine wether a gene is turned onor off. Explain this during embryonic development:
When embryo is forming around blastocyst stage, prior to implantation, there is a global wave of gene demethylation and wipes away any epigenetic modifications. There is then a second wave of methylation that puts the modification that is needed back on.
Metaphase chromosomes cannot:
Express genes (buried)
Need to be in bow of spagetti form to express genes.
Explain how DNA is organized within the cell from least to most condensed (4):
- DNA (2nM)
- Nucleosomes (11nm)
- Chromatin fibre (30nm) Transcription inactive >30nM
- Metaphase chromosome (1400nM)
DNA packaging step 1(3) :
attract + structure + bp
- DNA has a negatively charged phosphate backbone. Histone are positively charged at the N-terminal tails so they attract.
- DNA will wrap around core histone octamer which is 8 different histone proteins (H2A, H2B, H3, H4) where H1 is the linker protein where DNA enter/exit to help fold in for compaction (same with linker DNA).
- 147 bp wrapped 1.7 times
Primary function of histone proteins is —-
transcriptional control
DNA packaging step 2 (3):
size + transcription + structure
- 30nm chromatin fibers
- Transcriptionally dormant because so compact
- Linker histones, linker DNA and N-terminal tails faciliate interactions between nucleosomes into high order structures
DNA packing is dynamic:
You can have one chromosome having parts that are loser (10nm) and parts that are more compact (30nm) depending on what are expressed.
DNA packaging steps 3 and 4 (2):
- 30nm fiber forms loops anchored to a protein scaffold (nuclear matrix), estabolishing a 300nm fiber compaction in chromosomes
- 300 nm fiber further folds in on itself forming a chromosome (metaphase)
Heterochromatin (4)
What + ex + function + size
Condensed or closed state. DNA is most organized.
- ex: Noncoding regions: centromeres + telomeres
Functions: Transcriptional repression, genome stability
30nm and above
Euchromatin (4)
state + transcription + size + ex
- Looser/uncoiled, “open” state
- Transcriptionally active: allows basal TF, RNA poly2, special TF acess to gene
- 10nm fibre (beads on a string)
- Ex: interphase chromosomes take up entire nuclear space
Chromatin remodeling complexes (4):
what + example protein + cooperate with + ex mechanism
- Large protein complexes (ATPases), require ATP to function
- SWI/SNF
- Cooperate with specific DNA-binding proteins to repress/activate gene expression.
- Ex: Remodeling complex A and DNA binding proteins can come in and DNA pulls away from histones making it less condensed (more space between each nucleosome) for transcription, DNA replication and repair. They can also slide the octomer around to expose previously inacessible sites or perform histone eviction. Remodeling complex B comes in and causes dissociation of DNA binding protein for restoration of standard nucleosomes.
What are the three categories of Chromatin remodeling complexes?
- Nucleosome assembly (Repress gene expression to promote gene silencing)
- Chromatin access (Promote transcriptional activation)
- Nucleosome editing (Replace 1 histone with another type (Ex: 1 H2A for another H2A), different variants, some may have specialized function
Chromatin access
Chromatin remodeling complexes can render chromatin more accessible to DNA-binding proteins, explain how this occurs (3):
Name + what they do
- Switch/sucrose non-fermentable (SWI/SNF) subfamily of remodellers
- Recognize histone modifications. Mobilize/unwrap or eject nucleosomes/histones.
- Transcriptional apparatus is granted access.
Chemical modifications of the nucleosome dynamically regulate transcription:
Posttranslational modification of histones (4) + DNA modification (1)
Histone acetyltransferases (HATs) (3)
location + mechanism of action + reversed
- Acetyl group added to lysine N-terminal tails (majority) and the internal globular domain to promote transcription
- Adding the acetyl group will cover the positive charge allowing chromatin structure to destabilize and recruits chromatin remodelling enzymes such as SWI/SNF
- Reversed by histone deacetylases (HDACs)
Explain transcription in relationship with histone acetyltransferase and HDACs
Methylation and demethylation of DNA (3)
what/target + transcription + exception
- Methyl groups added to cytosine nucleotides by DNA methyltransferases (DNMTs) especially targeted to CpG dinucleotides (cytosine next to guanine)
- Represses transcription: Prevents binding of TF + Recruitment of methyl-CpG binding domain (MBD) proteins (repressors) such as HDAC/chromatin remodeller repressors
- CpG sites are spread throughout genome and are usually methylated. Exception is CpG islands (most gene promoters), majority of gene promoters reside within CpG islands usually not methylated.
De novo methylation (2)
what + when
- Add methyl group to naked DNA. Both strand are not methylated.
- Embryonic development + environmental stimulation
Which enzyme for active demethylation, methylation maintainance and which for de novo methylation?
- Active demethylation: Gadd45 + TET
- Methylation maintainance (one strand already methylated so methylate the complememtary strand): DNMT1
- De novo methylation: DNMT3A and DNMT3B
What are the 2 general mechanisms for modulation of chromatin structure?
- Posttranslational modification (PTM) of histone tails (acetylation) and DNA methylation
- ATP-dependent chromatin remodelling enzymes (recruited when not enough)
Histone code hypothesis
DNA transcription is largely regulated by post-translational modifications to these histone proteins. Different combos at various AA residuces (N-terminal tails) may lead to either activation or repression of transcription.
