epigenetic control Flashcards
what is epigenetics
Processes that induced long term stable changes in gene activity without a change in gene sequence
describe the shift in genes expressed from fertilisation
Just after fertilisation you get expression of embryonic genes, this goes down for six weeks and get expression of alpha globin genes expressed throughout life. Foetal genes are expressed through foetal life which changes after birth in a switch from gamma globin to the beta globin gene expression.
genes are under ________ and ____________ control
Genes under temporal and tissue specific control
- via epigenetic regulation
what are the three main epigenetic processes
DNA methylation, Histone modification and non coding RNAs
what occurs in epigenetic DNA methylation
- Cytosine is methylated to 5-methyl cytosine – addition of CH3 group to the 5th carbon of the ring
(90% methylated cytosine fount next to G)
low levels of DNA methylation is associated with _________ of that gene
Low levels of methylation is associated with the activity of that gene
– low levels means RNA can bind and activate that gene.
high levels of methylation in the promoters ___________________ and the gene is __________
High levels in the promoters blocks RNA polymerase binding to the DNA and the gene is switched off
what are the 3 main DNA methyl transferases and what are their functions
- DNMT1: to maintain the methylation marks on the DNA every time a cell divides, it is copying them from parental strand onto the newly synthesised strands
- DNMT3a and 3 b: de novo, responsible for establishing the methylation marks in the first place
what enzymes demethylate DNA in the genes and how
• DNA demethylase: TET enzymes
Tet converts 5 methyl cytosine (5mC) to 5- hydroxymethylcytosine (5hmC)
5hmC is spontaneously oxidised or deaminated converting it back to cytosine.
how are histones able to interact with DNA
Histones are positively charged so they can interact with the negatively charged DNA – strong interaction
how do DNA and histones interact and pack
Nucleosome is DNA wrapped around core of histones – 8 units
Each nucleosome is further package to form a 30nm fibre/ solenoid
Then further looped and compacted to give 200nm fibre, this is how the DNA is stored in our cells
if the histone interactions with the DNA are loose it gives an ___________ structure that makes it __________________ and ________the gene
if the interaction is loose it gives an open relaxed structure that makes it easy for RNA pol to gain access to DNA and activate the gene
what determines whether interactions of histones and DNA is loose or tight/ active or repressed
the position and type of modification of the histones
what are the two main types of histone modification
histone acetylation
histone modification
what type of epigenetic control is histone acetylation associated with
- Histone acetylation: associated with open structure of DNA and so activation of the genes.
how does histone acetylation activate genes
positive amino acids on the tail like Lysine K when acetylated their charge is neutralised. - as a result, you reduce the affinity of the histone tail for DNA opening up the structure so the promoter and target genes are now accessible to transcription factors and RNA polymerase
the effect of histone methylation depends on what
which lysine is methylated
H3K4: methylation (to lysine 4) is associated with what change in the DNA
open structure & gene activity
what is methylation of H3K9 and H3K27 associated with
closed structure and gene silencing
what do histone writers do
proteins that add modifications to histones
give 3 examples of writers
Histone acetyl transferases (HATs); add acetyl groups
Histone methyl transferases (HMTs): add methyl groups
DNA methyl transferases (DNMTs): add methyl groups to DNA
what do histone erasers do
remove the modifications
give 3 examples of erasers
Histone deacetylases (HDACs): remove acetyl groups restoring the positive charge so it can have stronger interactions with DNA and more condensed form of the gene so RNA pol cant bind and the gene is then switched off
Histone demethylases (HDMs)
DNA demethylases – remove methyl groups from DNA
what are histone readers
read the epigenetic code and interpret it
give an example of a reader of histone acetylation
SWI/SNF clears nucleosomes from promoter region - Preferentially binds to acetylated histone, on binding it clears the nucleosomes away from the promoters of the gene making it easier for RNA pol to bind to the gene – make a region easy for it to bind and transcribe
give an example of a reader of histone methylation - what is its effect
Reader of histone methylation at K9/K27: HPI1 heterochromatin protein 1 - Binds preferentially to methylation at K9 or K27 coats dna inducing permanent silencing of that gene
give an example of an epigenetic writer
Polycomb repressive complex (PRC)
PRC1/2 work together. 2 thought to be most important because it has catalytic activity. Comprises 4 subunit, EZH2 of PRC2 has catalytic ability – a HMT (writer). Transfers 3 methyl groups to K27 (mediates trimethylation of H3K27). Key role in making sure the right genes are on it the right cell types.
non coding RNAs are divided into two groups what are they
• Small non coding RNAs
Less than 200 bases
eg miRNAs, piRNAs, tRFs
• Long non coding RNAs (LncRNAs)
Greater than 200 baseswhat are micro RNAS
- small single-stranded non-coding RNAs of 21-25 nts in length
bind to target messenger rnas and induce their degradation in the cell or inhibit the translation. - help control gene expression
how are microRNAs made
Transcribed first into a primary microRNA.
Then processed by drosha which creates a pre microRNA.
Transported into cytoplasm by exportin 5 where its cleaved by dicer into a mature microRNA. Then loaded onto miRISC complex which takes it to it messenger RNA.
what do microRNAs induce depending on where they bind to the messenger RNA
If the micro RNA binds to three prime end of mRNA it induces degradation of that messenger RNA in the cell.
If the five prime then it tends to inhibit the translation.
what mRNAs regulate embryo development early on
Maternal Messages (mRNAs) inherited from the oocyte (maternally inherited) regulate early embryo development until the first stage of cleavage wear they are degraded
when do you get transition from maternal mRNAs (MZT)
around the first stage of cleavage
what happens during the maternal zygote transition
Around the first stage of cleavage the maternal mRNAs are degraded and you get activation of zygotic genome
MZT: maternal-zygotic transition
ZGA: zygotic genome activation
Initially, the destruction of maternal mRNAs is accomplished by maternally encoded products – destabilise the mRNAs
Zygotic transcription leads to the production of miRNAs ( miRNAs-430/270)
when does compaction occur
when cells reach the morula stage
when do cells start to change from totipotent to pluripotent
when differentiation and cavitation occur
Zygote through to morula all cells are totipotent. From then the inner cells are pluripotent bcos can form all cells of adult but not placenta
what is the composition of the morula
- in the morula the cells in the middle are undifferentiated and the trophectoderm cell differentiate
what does CDX2 during preimplantation development
it activates all the genes necessary to form the Trophectoderm cells
downregulates Oct4 the pluripotency factor allowing the cells to differentiate
what happens in CDX2 null embryos
no TE cells, no implantation
what is Dapi stain
is a blue-fluorescent DNA stain that exhibits ~20-fold enhancement of fluorescence upon binding to AT regions of dsDNA
what are the two proposed models for the expression of CDX2 exclusively on the outer cells
inside/ out model
cell polarity model
describe the inside/ out model of proposed cdx2 signalling
Cells on outer edge of morula have reduced cell to cell contact, so hippo signalling pathway is inactive.
So Yap protein is able to translocate nucleus of cell and interact with Tead4 (sits on five prime end of control region of cdx2)
Inside the nucleus Yap recruits HATs which will acetylate histones to open up DNA which will then recruit SWI/SNF that moves nucleosomes out of the way allowing RNA polymerase to bind and activate the gene.
In the outer cells you now get expression of CDX2 which will downregulate Oct4 allowing them to differentiate into TE
The inner cells are now surrounded by cells, this increased cell to cell contact leads to activation of hippo pathway, yap is phosphorylated so can no longer move into the nucleus of the cell, so cdx2 stays off and these cells can continue to express Oct4 and stay in an undifferentiated state.
describe the cell polarity model proposed for CDX2 expression in TE cells
Proposes the outer cells are polarised with an apical and basal edge.
As a result, it allows AMOT to interact with polarity proteins which suppress the activity of protein LAT (kinase).
Means Yap is not phosphorylated so can move into the nucleus and allow transcription of CDX2.
The inner cells are apolar meaning the Lat kinase is active, and the AMOT protein is localised to adherence junction. As a result Lat will phosphorylate AMOT which will phosphorylate yap, and so it cant translocate into the nucleus so CDX2 is off and Oct4 is continually expressed.
When there is a differentiation signal, oct4 expression will go down and you get expression of developmental control genes
when are the methylation marks on the maternal and maternal genomes removed
Upon fertilisation - paternal genome marks removed within 12-24 hours of fertilisation
Followed by eraser of DNA methylation marks on the maternaly inherited DNA usually complete after 48 hours.
demethylation of the maternal and paternal genomes is mediate by what enzyme
TET
what is the state of the genome in totipotent cells (in terms of developmental control genes)
they’re not methylated but not active either said to be in a poised state
Promoter regions of these genes they are marked by both active and repressive marks – trimethylation of lysine 4 and RNA pol bound. Trimethylation of lysine 27 and binding of polycomb complex – referred to as bivalent domains.
when totipotent cells receive differentiation signals what happens to the genome
When they receive signals, the bivalent domains are dissolved.
Loses repressive marks retain active if that particular gene is needed. Visa versa for requirement
Retains repressive marks then Binding of protein HP1 and DNMT – genes are permanently switched off
as development continues you get remethylation of DNA in a cell specific manner, what enzymes do this
Re-methylation is mediated by DNMT3a and 3b then maintained by DNMT1
this marking is then thought to be stable and maintained, locking these cells into their differentiated form
when is the epigenome most susceptible to changes in the early life environment
in fetal and early embryonic life
what parts of the early life environment contribute to alterations of the epigenome
parental body composition, maternal nutrition, maternal exercise, pollution, social adversity, diet, body composition and exercise
how does in vitro culture affect the mRNA expression in blastocysts
Expression of 5/11 genes involved in epigenetic and chromatin regulation was reduced
how does in vitro culture affect the male offspring phenotype
insulin resistance
aberrant testes gene expression
less sperm with reduced motility
reduced fertility
what is large offspring syndrome (farm)
Abnormally large offspring observed after in vitro production or manipulation of farm animal embryos.
which rare human diseases increased 3.5 fold as a result of invitro fertilisation
Beckwith-Wiedemann and Angelmans syndrome
how does smoking affect the epigenome
alters methylation of very specific genes eg AHRR that plays a role in tumour progression
As a whole increased metabolic stress, decreased genomic stability, heightened risk of mutation and altered brain development.
how can nutrition/ the maternal diet affect the epigenome
can alter the methylation of key metabolic genes within the offspring
(eg restriction can induce changes in PPARα methylation in fetal liver)
maternal protein restriction in rats lead to what changes in the offspring
change in the ratio of TE/ICM, hypertension, dyslipideamia and insulin resistance
what was the Dutch Hunger Winter
a period of food shortage in the Netherlands in 1944
energy intakes dropped from 1800 to between 400 and 800 kcal a day
Alterations in DNA methylation has been shown in adult children from mothers who were exposed to famine during pregnancy compared to their non exposed siblings
what epigenetic changes occurred as a result of the Dutch Hunger Winter
Change in methylation of igf2 and lep genes (involved in appetite control) – these changes measured 60 years after the famine.
pregnant rats fed a high fat diet led to what epigenetic chagnes
to altered methylation of appetite control genes reduced locomotor activity, altered food preferences and obesity
maternal over nutrition leads to what
Altered methylation of appetite control genes, as well as altered food preference and obesity.
eg changes in DNA methylation and increased adiposity in the child
how does the paternal environment affect the child
Paternal diet can affect the non coding RNAs within the sperm
( sncRNAs deposited in fertilised eggs may influence early development specifically transfer RNA-derived RNA fragments tRFs (Sperm mediated epigenetic inheritance))
what are tRFs
tRNA-derived small RNA fragments
Trfs act as microrna and bind to 3’ end of mrna and helped gradation or inhibit translation of rna as well
what parts of the paternal environment can effect epigenetics of offspring
paternal body composition or diet
