Metabolism and Epigenetic Transgenerational Epigenetic Inheritance Flashcards

Question

What happens if the male experiences stress during spermatogenesis?

Answer 1

Stress will be transmitted to the next generation (only one generation)

Answer 2

The altered activity of chromatin modulating enzymes could have a significant impact on the differentiation of tissues in the developing embryos Hence: if the mother is exposed to adverse conditions, the epi-genome of the future child will be altered Sometimes, these adverse exposures of the pregnant mother can be passed to grandchildren and grand-grand children, which is known as epigenetic memory, the same can apply to fathers

Answer 3

Responsible for the synthesis of Phaeomelanin The deposition of pigments in the hair by melanocytes depends on the activity of the Agouti gene (A)

Answer 4

Oscilates between on and off, which means the gene cycles between active and inactive in the adult organism

Answer 5

Agouti OFF: Black pigments are made Agouti ON: Yellow pigments are made Black is the recessive allele Yellow is the dominant allele

Answer 6

Well, it would be yellow and yellow mice are obese and prone to diabetes

Answer 7

Agouti expression oscillates 1) Oscillating OFF: BLACK 2) Oscillating ON: YELLOW, so a single piece of hair has yellow and black stripes 3) Oscillating OFF: BLACk So, the overall colour of the wild type mouse is brown

Answer 8

Agouti allele: Normal or Wildtype Oscillating ON/OFF: Brown (agouti) Agouti- (a): No functional protein made, so it is always OFF, meaning a is black Ayellow (Ay): Oscillating doesn't work, so always ON, meaning the mouse is yellow, obesity and diabetes Avy allele: Oscilation works only in some cells

Answer 9

It contains a transposable element, IAP which is upstream of the Avy promoter IAP is a transposon LTR, which means it has its own promoter IAP has its own promoter that can override the normal oscillating Agouti promoter The epigenetic state of the IAP promoter can determine the level of expression of the yellow pigment

Answer 10

Normal A allele (brown fur): The promoter alternates/oscilates between on and off Mutant a allele (black fur): The promoter is always OFF Mutant Ay allele (yellow fur): The promoter is always ON Avy allele: The Avy contains an IAP, which has its own promoter, and the promoter can override the normal promoter. The IAP promoter only overrides the normal promoter in some cells and not others, which is what results in the patches of yellow fur and patches of brown fur, as such the normal promoter works in some cells, and not in others, and in ones where the IAP overrides, the normal promoter does not work. The IAP overrides normal promoter to produce yellow coat when IAP promoter is unmethylated

Answer 11

Some Avy mice are yellow, some are brown, but most have brown and yellow patches Hence, the Avy allele is in different states in genetically identical mice and within the cells of the same individual All mice have the exact same genotype (Avy), but different phenotypes, (yellow, brown, agouti)

Answer 12

There are two potential epigenetic states of Avy that can occur within cells of Avy/a mice First off all, the a allele is not functional, because agouti gene is mutated The IAP which has a promoter, lies upstream of the agouti gene If the IAP promoter is unmethylated, there is ectopic expression of the gene from the IAP, which results in a yellow coat colour If the IAP promoter is methylated, this means that the gene is expressed under its normal development controls, leading to a brown coat colour.

Answer 13

1) Cross Avy/a (male) with a recessive a/a female mice *2 2) Supplement diet with folic acid/VitB12: rich diet in one of the groups (abundance of SAM), leave other group with a normal diet 3)

Answer 14

Diets of female a/a mice are supplemented with methyl donating substances: folic acid, choline, vitamin B12 and give diet 2 weeks before mating with Avy/a males and throughout pregnancy and lactation

Answer 15

In the un-supplemented mother, there is less methylation of the IAP, leading to more yellow phenotypes In the supplemented mother, there is plenty of SAM in the diet, which means IAP is more methylated here than in the normal diet, which produces more brown phenotypes Therefore, the folic acid content in the mother's diet leads to differential methylation of the IAP promoter in progeny, determining the coat colour. It was seen that a folic acid/VitB12 rich diet reduces the frequency of yellow phenotypes, which suggests that exposure during pregnancy has epigenetic consequences for the progeny

Answer 16

Dietary supplements high in SAM lead to the IAP upstream of agouti gene to be more methylated than the mothers fed an unsupplemented diet, More methylation:Brown Less methylation: Yellow

Answer 17

SAM is a donor group of methyl, so when it gives methyl group, Mts can methylate histone/DNA which promotes formation of heterochromatin So, therefore: building heterochromatin depends on the amount of SAM individuals have

Answer 18

Genomic transmission and altered cellular environment do not have the potential for epigenome reprogramming, which means that they will not be transmitted to progeny Non genomic transmission during development can be transmitted to progeny , leading to epigenome reprogramming, and therefore altered gene expression in progeny

Answer 19

Development of neurons takes longer than the development of other tissues, so prolonged stress on the mother has a higher chance of affecting neuronal differentiation development Neurons affect behaviour

Answer 20

methylation of DNA in the sperm can transmit epigenetic marks to progeny, but not well understood

Answer 21

Paternal transmission of epigenetic defects to either sons or daughters can occur if an epimutation is introduced into the sperm epigenome in the form of altered DNA methylation or histone modification patterns on the nucleosomes that package the genomic DNA Such alterations can be induced by aberrant genetic or environmental effects, and if the epimutations escape germline reprogramming, they can be transmitted to multiple subsequent generations, even in the absence of any further exposure to the original inducing effect

Answer 22

Set up a male mice control group, male group exposed to acetophenone, and a male group exposed to propanol (aversion??) Males were mated with naive females (no exposure) Separate males from female F1 offspring born and observe effects Mate F1 males with naive females F2 offspring born

Answer 23

Removed so that behaviour of avoidance cannot be taught by males to progeny To avoid teaching mice babies to avoid the smell F1 progeny not taught behaviour by father

Answer 24

In F1 whose fathers were exposed to actophenone, there was a strong aversion to the smell, suggesting trait was acquired as the behaviour was not taught. In F1 whose fathers where exposed to propanol, there was a strong aversion to the smell, suggesting this trait was acquired as the behaviour was not taught EPIGENETIC INHERITANCE!!

Answer 25

Looked at differential methylation of olfactory centre in the brain of these mice, to show behaviour is not genetic, but rather epigenetic because it is an accquired trait that cannot be explained by Mendelian genetics as fathers were not born with aversion trait

Answer 26

If DNA replication is impaired, there is a loss of repression of the construct Daf-21 drives expression of GFP in this construct (normally repressed) Experiment was performed with licensing factors and DNA polymerases to produce progeny following a Mendelian distribution If experiment is performed at 20 degrees in wildtype worms, worm is not very green If experiment is performed at 20 degrees in mutant worms, worm is green The worm is green because there is a loss of repression of Daf-21 If there is a loss of repression of Daf-21, heterochromatin cannot be maintained Overall, worms with KO genes of replication factors are mated to mCHerry array worms Replication deficiency lasts only during embryonal development Impaired replication de-represses mCherry array

Answer 27

DNA replication correlates to both H3K27me3 and H3K9me3 The transmission of H3K27me3 is dependent on DNA replication, and PRC2 catalyzes the methyl of H3K27 Behaviour correlates with the inability to maintain both epigenetic marks (H3K9me3 and H3K27me3)

Answer 28

Associated with mutations of cell cycle regulating genes in somatic cells We observed genome instability and impaired DNA replication (mutations in checkpoints, p53, progression through damaged DNA, etc..)

Answer 29

Epigenome plays a large role in cancer progression In cancer, epigenetic aberrations, including methylation 100-1000, histone modifications > 1000, miRNA

Answer 30

It shows genetic inheritance because the mutation that confers resistance to streptomycin was not acquired during exposure to streptomycin, it was spontaneous, so we need to mutate gene to see transmission

Answer 31

Neurons engage in multiple connections via synapses Avoid the generation of synapses with themselves, known as self-avoidance

Answer 32

In humans, by the Pcdh genes The selection of the pcdh varian is epigenetically regulated The selection involves the differential methylation of CTCF binding sites The methylation of CTCF determines a complex 3D structure and the selection of the Pcdh variant Pcdh genes have precise control over which neurons interact with other neurons (regulate cell to cell contact)

Answer 33

A mechanism for self-recognition Neurons need to recognize other neurons, cannot recognize themselves, so polycadherins make sure that neurons do not interact with themselves.

Answer 34

alpha, beta, gamma clusters span a genomic region of 1 Mb The alpha cluster has 12 monoallelically variable exon and two biallelically expressed variable exons (C1 and C2) All variable exons splice to three constant expense that code for the intracellular region

Answer 35

Pcdhalpha alternate genes are expressed by long range DNA contacts between their promoters and the HS5-1 enhancer These contacts are mediated by the CTCF and Cohesion protein complexes The choice of Pcdha alternate promoters is also regulated by histone and DNA methylation (H3K9me3 and 5mC) and the activity of the Smcdh1 protein

Answer 36

DNA methylation of CTCF binding sites and the DNA sequence between them, correlates with CTCF occupancy of CBS sites, engagement with HS5-1 enhancer, and transcriptional activation of the unmethylated promoter

Answer 37

Neuroblast cell expresses only three variable Pcdh-alpha exons There are two CTCF binding sites at each active variable expense (CSE and eCBS sequences) at two at the HS5-1 enhancer (HS5-1a and HS5-1b)

Answer 38

Regions outside the central transcriptional hub are thought to be packaged in a repressive chromatin conformation Suggested by the DNA methylation of in active pcdh-alpha promoter regions Active genes are thought to be concentrated in a transcriptional hub, aided by a double clamp binding of CTCF, between the variable exons and the HS5-1 enhancer and cohesion

Answer 39

Methylation Splices out exons downstream of chosen exon

Answer 40

It reveals that 14 generations display stress changes, even though all generations after the F0 were not exposed to the stress

Answer 41

It reveals that 14 generations display stress changes, even though all generations after the F0 were not exposed to the stress

Answer 42

mCHERRY expresses red fluorescence, and when working red fluorescence will be seen mCHERRY is repressed at 20 degrees Celsius

Answer 43

In F0 generation, there was exposure to high temperature, 25 degrees celcius and this induced expression of mCHERRY In F1 to F14, the temperature was decreased to 20 degrees celcius, but mCHERRY was still expressed, so there was abnormal expression even in the normal temperatures

Answer 44

The worms contained cloned repressed mCHERRY array of genes mCHERRY is normally repressed, but derepressed if the worm is grown at 25 degrees - See epigenetic inheritance here because at 20 degrees mCHERRY is still expressed Transmission occurs through oocytes and sperm and in cis with the locus

Answer 45

PRC2 is not involved Worms are grown at 25 degrees and then grown for 15 generations at 20 degrees The de-repression of mCHERRY persists for 15 generations Great example of a multigenerational epigenetic transmission in a model organism

Answer 46

FISH to detect expression Looking for an overlap between mCHERRY and H3K9me phenotype associated with loss of H3K9me, because trait is not dependent on PRC2

Answer 47

H3K9me3 is involved in mCHERRY exposure Changes in H3K9me3 accompany de-repression and is transgenerationally inherited The development at high temperature from embryo to adult results in reduced H3K9me3 on the array in the germline nuclei of adults

Answer 48

H3K9me3 is depleted from transgene locus in F2 descendents of worms grown at 25 degrees celcius F2 embryos were extracted Reduced H3K9me3 on the array of germline nuclei of adults was extracted at moved from 16 degrees to 25 degrees during development and compared with worms kept constantly at 16 degrees celcius