The document is a summary of the lecuters of the course epigenetics and gene editing. Only lecture 5 and first half of lecture 6 on developmental epigenetics are missing.
The summary is really elaborate and with a lot of pictures
In disease, a lot of genes are sleeping, not mutated but not expressing enough product
Two sides of spectrum
1. Developmental epigenetics
a. Heritable
b. Not encoded in DNA seq
c. Accessibility of the gene
i. Level of expression
d. Determines
i. Differentiations and maintenance of cells
ii. Healthy aging - e.g. epigenetics clocks
iii. Disease
1. Diagnostics
2. Therapies
e. Reprogramming gene expression
2. Gene editing
a. CRISPR → nobel prize 2020
b. Changes in seq
c. Biomedical research
d. Rewriting gene seq → ethical issues
Epigenetics is not that far yet → now 8 companies
Induced pluripotent stem cells are nice example of epigenetic editing
Lecture 1
DNA, structure and organization
● Central dogma of biology only goes for 1.5% fo the human DNA
● Not all DNA is expressed in all genes
● Expression profiles are maintained during cell division
→ regulation of gene expression
Heritable diseases is NOT always due to changes in DNA seq, mendelian diseases is only a
small part of the diseases
Heritable susceptibility problems → runs in the family, can be genetics but often epigenetics
Most affected by environment (smoking, fast food) - epigenetics
Epigenetics = heritable, yet reversible changes in genome functioning, not encoded in the DNA
seq
DNA
- 4 building blocks, double helix, karyotype → geneticists
- From double helix to karyotypes → steps, really controlled → histones
- Organization of genes
- Brown = known genes
, - red= predicted genes → lot of the genes are still predicted
- gene= protein coded part of DNA
- Regulatory DNA seq→ determine not the kind of protein but the amount
- In a gene there are also introns, and small part is exon
- Some seq are not coding protein but are expressed→ not without function
- Genes that encode RNA only!
- NOT junk
- 40% of junk is unique
- Introns (regulation gene expression)
- Regulation gene expression (promotor)
- Non-coding RNAs (regulation gene expression)
- And this 1.5% of genes is not coded in every cell → epigenetics
- How is it regulated
- How to interfere
- DNA really long in a very small place
Functions of the DNA
- gene expression
- Replication
- Both in interphase: chromosomes are long and thin
-Cell division → mitotic chromosomes are compact (compacted form is only in m -phase)
- Often the chromo are in the interphase - chromatin leaking out of lysed nucleus during
interphase
- Mitotic chromosomes:
centromere even more
compacted part
- Chinetikore →
epigenetics
determine where it
binds
- Telomeres -
replication fo the
ends protection against “repair” -marked by
sequences and epigenetics
- Replication origins - where DNA duplication
starts
- centromere(1 per chromo): keeping 2
daughter chromatids together during mitosis
Organization
- DNA is not ‘naked’ double helix
- Histones
- Beats on a string formation → not how DNA likes to be → basic compaction of
DNA (step 1 of formation)
, - Chromatin fiber of packed nucleosomes
- Fiber in loops
- H2A, H2B H3 and H4 → majority of the histones
- Are subjective to post-translational modifications
- Nucleosome: contain 2x(H3+H4)+2x(H2a+H2B), 147 bp
are wrapped round and 1x linker DNA
- DNA + histones + non-histone protein = chromatin
- Fig4-21- experimental derived formation, (unpacked
stage)
- The length is x500 during interphase compared to
double helix formation
- 10 000x longer linear DNA compared to compact
form during mitosis
- Roughly two chromatin types: heterochromatin (closed)
vs euchromatin (open)--> easier to express
- This formation is dynamically organized → open
can become closed and vice versa
- How can it be maintained?
- It is highly controlled→ where and
when is it reprogrammed
Packing unit = nucleosome
- 8 histone protein
- 147 bp DNA,1.7x around
- These two form the nucleosome core
- Plus <80 bp linker DNA (little string between beats)
- Linker DNA is less protected - nuclease digest can
easy digest linker DNA → found out that the wrapped
DNA is 147 bp
- Histones are very well conserved between evolution
- All the four histones have similar histone folding
- H3+H4 dimerize and H2a and H2B
- N terminal tail differs between histones
Nucleosomal DNA
- less accessible than linker DNA
- Many H-bonds per nucleosome
- Many aa of histones are made of lysines of argenines→ + charged and
DNA is - charged
- Not a particular seq, location is affected by the DNA binding proteins
- Tight association
- Yet transcription / replication / repair is possible = breathing, chromatin
remodeling (some movement)
- It is stable yet is is dynamic
- Figure slide- gray is binding site →
unwrapped and rewrapped on and on = breathing
, (very short time)--> regulatory seq is exposed, regulatory proteins can bind in this small
time protein → cellular processes. When the protein does not bind the DNA keeps on
breathing
- ATP dependent (controlled)
- Chromatin remodeling
- Mostly by enzymes that are ATP dependent, bind histones and DNA,
more than 10 subunits, multiple rounds of ATP hydrolysis
- ATP dependent chromatin remodeling complex
can slide the DNA over the histone complex→
other seq is accessible
- Nucleosome sliding = pulling histone
octane along DNA double helix →
- Other
ways: making uses of
chaperones
2 ways
- 1. Histone exchange
- Chaperone bind H2A H2B dimers → replace
them by other H2A H2B dimers
- 2. Histone octamer
removal-Take the whole histone octamer and take
it out → transcribe→ put back the nucleosome
core again
~next slide is sort of summary~
Higher order
- 30 nm chromatin fiber: possible zigzag structure
Chromatin compaction: histone tail
- Compaction: interaction through trials (particular histone 4)
- Linker histone H1
- H1 is associated with the outgoing DNA → determining the direction
- 1 molecule present per nucleosome core
- Bind DNA and protein
- Larger than other individual core histones
- Less conserved → but it is functional (in cancer)
- Determines direction “nucleosome-outgoing” DNA
Chromatin structure and function
It is more than just the packaging of NDA → dynamic regulation
- Lot of non-histone proteins (e.g. TF, equal amount of histone vs non-histone protein)
- Histone proteins are highly conserved
- Chromatin structure can switch off genes
- Chromatin structure; cell type identity, cellular memory (epigenetic inheritance)
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